Dr Boris Gouilleux

Dr Francois-Marie Moussallieh

Dr Philippe Lesot
email: philippe.lesot@universite-paris-saclay.fr

Prof Aharon Loewenstein

Anisotropic 1 H STD-NMR Spectroscopy: Exploration of Enantiomer-Polypeptide Interactions in Chiral Oriented Environments

Keywords: 1 H STD-NMR, PBLG-based Liquid Crystals, Enantiodiscrimination, Molecular interactions, Epitope mapping

We explore and report for the first time the use of 1 H saturation transfer difference NMR experiments (STD-NMR) in weakly-aligning chiral anisotropic media to identify the hydrogen sites of enantiomers of small chiral molecules interacting with the side-chain of poly-γ-benzyl-Lglutamate (PBLG), a helically chiral polypeptide polymer. The first experimental results obtained on three model mono-stereogenic compounds outcomes are highly promising and demonstrate the possibility to track down possible differences of spatial position of enantiomers at the vicinity of the polymer side-chain. Anisotropic STD experiments appear to be well suited for rapid screening of chiral analytes that bind favorably to orienting polymeric systems, while providing new insights into the mechanism of enantio-discrimination without resorting to the time-consuming determination of molecular order parameters.

Introduction

NMR spectroscopy in polymer-based, lyotropic chiral liquid crystals (CLC) is a powerful analytical tool to address many issues of molecular enantiodiscrimination, enantiopurity determination and stereochemistry. [1,2,3] These aligned media, which are formed by organic solutions of helically chiral homopolymers oriented with their main axes parallel to the static magnetic field B 0 , interact differently with enantiomers of a guest chiral molecule, so that they exhibit a different orientation, on average, relative to B 0 . Investigating and evaluating the interaction processes is therefore a key step to understand the whole enantiodiscrimination mechanisms in these orienting systems. [4,5] The molecular orientation may be represented by a Saupe matrix, 𝑆 !" !,! , or an alignment tensor, 𝐴 !" !,! = ! ! 𝑆 !" !,! , which are different for both enantiomers when the enantiodiscrimination occurs as demonstrated from 1995. [4] The bracket symbol corresponds to an ensemble average. This orientation difference leads to a spectral discrimination of enantiomers based on variations of residual anisotropic observables, 𝑂𝑏𝑠 !"#$% , namely dipolar coupling (RDC), chemical shift anisotropy (RCSA) or quadrupolar coupling (RQC). [6] All can be simply expressed as the product of constant value -depending on the nature of the interaction -and the Saupe tensor elements (Eq. 1). [1] 𝑂𝑏𝑠 !"#$% ! !" ! = 𝐾 𝑆 !"

! !" ! (1)
These NMR interactions have been successfully exploited in structural/conformational studies, in the determination of 3D structures, relative configurations [7,8,9,10] or in the measurement of enantiomeric excesses, … [1,2] However, the underlying principles governing the enantiodiscrimination mechanisms, i.e. the difference between 𝑆 !" ! and 𝑆 !" ! , are still under investigation. [11,12,13] This understanding is of major importance for: i) determining the absolute configuration of stereogenic centers from NMR residual anisotropic data (RDC, RCSA, RQC) supported by computational simulations (molecular dynamics), [14,15,16,17,18] ii) designing the structure of new efficient polymeric-based lyotropic liquid crystal, e.g. choice of the polymer, [19,20,21,22] or iii)

optimizing the choice of co-solvent (polarity), the sample component concentration, the presence or not of additives, [23] to enhance the enantio-discrimination abilities.

Previous studies have assessed and compared the enantio-discrimination either for a given chiral solute in regards to various polymeric-based CLC, or rather for a collection of analytes dissolved in similar mesophases. This can be evaluated by measuring the local differential ordering effect (DOE) using selectively deuterated analytes [24] or at a more global level by comparing 𝑆 !" !,! for both enantiomers (variation of principal axis direction, 9D angle, …) determined using anisotropic data such as ( 1 H-1 H)-RDC, ( 13 C-1 H)-RDC and more recently 2 H-RQC. [1,2,6,11] While more comprehensive and reliable, this approach is sometimes time-consuming and requires a sufficient amount of anisotropic data to determine the Saupe's matrices, 𝑆 !" !,! , (over 5 for C 1 -symmetry chiral molecules). [START_REF] Emsley | NMR Spectroscopy Using Liquid Crystal Solvents[END_REF] Moreover, it is worth noting the average nature of these calculated tensors, which encompasses the orientational behavior of the chiral analyte both at the vicinity of the polymer ("bound" analyte)

and in the bulk of the mesophase ("free" analyte). In a fast exchange dynamic regime, this situation may be represented by a simple two-site model, as expressed by Eq. 2: [START_REF] Lesot | [END_REF] 𝑆

!" !"#$% ! !" ! = 𝑓 ! 𝑆 !" ! ! !" ! + 1 -𝑓 ! 𝑆 !" ! ! !" ! . (2)
In this equation, 𝑓 ! is the fraction of "bound" analytes, 𝑆 !" ! !,! and 𝑆 !" ! !,! are the Saupe matrix corresponding to the "bound" and "free" analytes, respectively. In this modeled view, we may safely assume that the term 1 -𝑓 ! is close to 1, while the elements in order matrix 𝑆 !" ! !,! are close to zero, namely the order of the analytes is negligible when they are mainly surrounded by the organic solvent (bulk).

From Eq. 2, the enantio-discrimination phenomenon leading to 𝑆 !" ! ≠ 𝑆 !" ! mainly arises from: i) enantioselective diastereomorphous interactions between enantiomers and the chiral polymer side-chains, i.e. 𝑆 !" ! ! ≠ 𝑆 !" ! ! , and ii) from affinity differences between R or S isomers and the chiral polymer involving a difference of bound fraction, i.e. 𝑓 ! ! ≠ 𝑓 ! ! , or iii) both of them.

Therefore, the direct investigation of the chiral analyte-polymer interactions is of major interest to further understand the enantio-discrimination mechanisms. [START_REF] Lesot | [END_REF] To this aim, we propose to characterize directly the oriented transient polymer-analyte array by saturation transfer difference (STD) NMR spectroscopy. [27,[START_REF] Gairí | Saturation transfer difference spectroscopy[END_REF] This is a well-established method to probe and characterize small molecules in weak interaction with a macromolecule based on intermolecular magnetization transfers. It has been widely used to assess ligand-protein interactions both in terms of affinity (determination of dissociation constants K D ) and geometry (epitope mapping). [27,[START_REF] Gairí | Saturation transfer difference spectroscopy[END_REF]29,30,31] Interestingly, this tool has also been employed for investigating polar solutes interacting with soft matters (as stationary phases used in HPLC) by 1 H high-resolution magic angle spinning (HR-_______________________________________________________________________________________________ at low speed) techniques, [32] as well as applied successfully to characterize enantiomersmacromolecule interactions in achiral isotropic liquids. [33,34,35,36] However, STD-NMR approach has never been tested so far to investigate the case of chiral lyotropic liquid crystals, to the best of our knowledge.

In this exploratory work, we report the first 1 H STD-NMR experiments performed in a polypeptide CLC (PBLG), a well-known and effective enantiodiscriminating polymer, in presence of various chiral secondary alcohols (R 4 -C*H(OH)(CH 3 )): hexan-2-ol (HL), 1-phenethyl alcohol (PL) and 3-butyn-2-ol (BL) (see Figure 2 and SI-1). These analytes are interesting chiral model compounds with common features: a marked shape anisotropy around the stereogenic center as well as an active site promoting H-bonding, but with different types of substituent that may either allow π-π or van-der-Waals interactions. Under these conditions, we anticipate that a STD-NMR approach may deliver valuable insights on difference of enantio-orientations at the vicinity of the polymer side-chains (difference in 𝑆 !" ! !,! ) and/or affinity (𝑓 ! ).

After describing the method, this article reports: i) a qualitative 1 H STD-NMR study to screen model chiral secondary alcohols that bind to the PBLG polymer as well as the identification of solute interacting sites; ii) a quantitative STD-NMR analysis carried out with an acetylenic chiral alcohol to highlight and characterize differential epitope mapping of the two enantiomers with respect to PBLG. This analysis is performed both on enantiopure samples and on a racemic mixture.

Results and Discussion

Anisotropic 1 H STD-NMR to screen chiral analytes interacting with PBLG. The basic principle of 1 H STD-NMR in anisotropic media is formally identical to that applied to isotropic samples. [27,[START_REF] Gairí | Saturation transfer difference spectroscopy[END_REF]29,30,31] Hence, from a practical point of view, the anisotropic STD-NMR experiments can be performed using the routine STD-NMR pulse sequence developed in isotropic liquids, with no peculiar modification (see Figure SI-2).

In anisotropic 1 H STD-NMR experiments, an adequate spectral region of the oriented polypeptide (PBLG) is selectively irradiated (saturated) while leaving those of the enantiomers unaffected. During the saturation process, 1 H magnetization is mainly transferred throughout spindiffusion to the polymer-side chain nuclei (most accessible region of the polymer due its high flexibility) under interaction with the oriented solutes. The intermolecular NOE between the polymer and enantiomer spins leads to a transfer of magnetization from the polymer to solute molecules in a "bound" state, which is then shared with those in the bulk by a rapid exchange between the "bound" and "free" states. A scheme of this process is shown in Figure 1a. As this magnetization transfer impacts the signal intensity of the solute, substracting the 1 H spectra recorded in the absence ("off-experiment") and presence ("on-experiment") of polymer saturation leads to a differential spectrum, called the "anisotropic STD spectrum", which is non-zero or zero for interacting or non-interacting enantiomers, respectively. Amplification factors, 𝑆𝑇𝐷 ! , can be then determined as the intensity (or area) ratio between the remaining signals on the STD spectrum and on the reference spectrum (off-experiment) as:

𝑆𝑇𝐷 ! !,! (%) = ! !"" !,! (!) !! !" !,! (!) ! !" !,! (!) ×100 (3)
In Eq. 3, the terms, 𝐼 !"" !,! (𝑖) and 𝐼 !" !,! (𝑖), correspond to the signal intensity (or area) of a site 𝑖 of the analyte measured with the "off-experiment" and "on-experiment", respectively (see Figure 1b).

The superscripts R and S highlight the fact that enantiomers potentially lead to different STD amplification factors for a given chiral molecule dissolved in an enantiodiscriminating mesophase. As they are based on an intermolecular NOE, the STD amplification factors are more important if the interacting nuclei are spatially closed, reflecting the spatial proximity of the chiral solute towards the polymer side-chains in the "bound" state. Thus, the distribution of the STD values (i.e. epitope mapping) for each enantiomer may deliver new and valuable insights on the mechanisms of enantiodiscriminations in PBLG phases. Besides the efficiency of intermolecular polymer-solute NOE, the magnitude of STD signals also depends on accumulation of molecules labeled by the magnetization transfer in the bulk during the saturation process. This is driven by the duration of polymer saturation, 𝑡 !"# , as well as the exchange features between "bound" and "free" states. [START_REF] Gairí | Saturation transfer difference spectroscopy[END_REF]29,37] This latter corresponds to the time-residence, 𝑡 !"# , of the transient polymer-solute complex and fractions of solutes in both states as well (see Figure 1a).

Contrarily to STD-NMR in liquids, anisotropic STD-NMR experiments investigate solute molecules that are oriented on average in the magnetic field, and hence it could be argued that site-specific 1 H STD amplification factors measured in liquid crystals is biased by possible anisotropic dipolar relaxation mechanisms ('intermolecular NOE'). In the case of (weakly aligning) PBLG-based lyotropic systems, we may safely assume that this bias does not exist (at least significantly). Indeed whether the main relaxation processes of solute (dipolar mechanisms for 1 H and quadrupolar mechanisms for 2 H) are sensitive to solute molecular ordering, S, the spectral density J(ω) shows a dependence in S 2 in a first approximation. [38] Thus, when the solute ordering is small (S < 10 -3 ), the S 2 dependency on « anisotropic » 1 H dipolar relaxation mechanism is negligible, and so the classical isotropic relaxation models (used to explain the 1 H STD-NMR) remain valid. [START_REF] Vold | Nuclear Magnetic Resonances of Liquid Crystals[END_REF] In the PBLG mesophases, solute ordering is ranging between 10 -3 and 10 -5 . These values clearly indicates that the contribution of "anisotropic" dipolar relaxation mechanisms to 1 Qualitative STD-NMR study on three model chiral alcohols. In this exploratory work, anisotropic 1 H STD-NMR experiments are carried out on three chiral secondary alcohols (PL, BL and HL), sharing a common hydroxyl, methyl and methine group on the stereogenic center, but three different fourth substituents (rigid or flexible moiety) (see Figure 2a). All of them are characterized by a marked molecular shape anisotropy around the stereogenic center (see Figure 2b), while PL and BL have been abundantly exploited as model analytes to evaluate the enantiodiscriminating capability of various chiral mesophases by 13 C, 2 H or NAD NMR. [START_REF] Lesot | [END_REF][START_REF] Meddour | [END_REF]41,42] With respect to molecular interactions, the donor proton of the OH group can a priori easily establish an H-bonding (-OH) with the carbonyl site of the PBLG side-chains, while the aromatic and acetylenic substituents can provide π-π interactions (or even π-stacking) with the terminal aromatic ring of the polymer. [START_REF] Arunan | Definition of the hydrogen bond (IUPAC Recommendations[END_REF]44,[START_REF] Lewis | Modern Computational Approaches to Understanding Interactions of Aromatics[END_REF] To qualitatively evaluate their respective interactions with the polymer side-chain, STD-NMR experiments are performed using identical experimental conditions for the sample preparation.

Thus, the polarity of the co-solvent (CD 2 Cl 2 ), ratios of PBLG/co-solvent (w PBLG /w co-solv. ) or PBLG/total weight (w PBLG /w Tot. ) as well as molar quantity in analyte have been kept constant (see Table SI-1). Each enantiomer of the three chiral analytes (HL, PL, BL) are analyzed separately, with the exception of HL for which only samples of (S)-HL and (S/R)-HL have been considered. Interestingly, these further STD signals (only observed at long saturation times (𝑡 !"# > 5 s)) are not associated to the same site in each enantiomer (the methine site for the (S)-isomer and methyl site for the (R)-isomer) (see Figure 2c). These differences in STD spectra clearly evidence a difference of enantiomeric orientation with respect to the external field B 0 in their "bound" state, namely when they strongly interact with the polymer side-chains. The highest STD amplification factors (%) are observed in the case of the acetylenic alcohol BL (see Table SI-3). For both enantiomers, all 1 H sites (see Figure 2c) exhibit a significant amplification factor on the 1 H STD spectrum (from 5.1 to 16.7%), including for the ethynyl group (see Figure 3 andTable SI-5). These results clearly suggest an adequate/optimal spatial organization of the ethynyl moiety (and the whole molecule) towards the terminal aromatic group of the glutamic chain. This example appears to be the first direct proof of possible π-π interactions between unsaturated bounds of the solute and the aromatic ring of PBLG, and their important role in the enantiodiscrimination mechanism. The lack of noticeable STD effect visible on the aromatic ring of PL, for which π-π interactions could also be expected, could simply be understood as a consequence of higher steric hindrance around the asymmetric carbon that prevents an easy interaction with the glutamic phenyl group (in the inner space between to side-chains) (see Figure SI-1). Finally, relative STD values from sites-to-sites vary from the (S)-isomer to the (R)-isomer of BL, suggesting once again that enantiomers are oriented differently at the vicinity of the polymer sidechains, and confirming the first observation with PL. This difference of epitope mapping between the two enantiomers towards PBLG during the binding process are further investigated in a quantitative manner in the following section.

Quantitative STD-NMR analysis of (R)-and (S)-BL in PBLG.

Beyond the first qualitative study described above, a quantitative approach is then carried out to finely evaluate the variation of epitope mapping without possible experimental bias. We focus here on the case of BL as it delivers intense STD signals and well resolved spectral patterns for both isomers making it a suitable model for such an investigation. STD-NMR experiments are first performed on two separated samples of enantiopur (S)-and (R)-BL with the same composition, and prepared with the same protocol (see Table SI-1). The temperature is fixed at 275 K in order to circumvent the peak-overlap between the hydroxyl and the acetylenic signals.

As seen in Figure 3a, the comparison of anisotropic 1 H NMR spectra shows clear spectral enantiodiscriminations with large changes in the spectral patterns, especially for the methyl group.

An even better spectral discrimination is obtained with other nuclei such as 13 To assess this feature in a reliable way, a quantitative approach of STD-NMR in PBLG is considered. Indeed, in the first approach explored previously, there are several bias that may prevent a direct and reliable interpretation of STD signals in terms of internuclei distances and thus of relevant geometrical insights on the binding process. Possible biases are: i) a fast polymersolute rebinding process during t sat , [START_REF] Gairí | Saturation transfer difference spectroscopy[END_REF][START_REF] Angulo | [END_REF]47,48] and ii) a strong variation of longitudinal relaxation time, T 1 ( 1 H), from site-to-site. [START_REF] Angulo | [END_REF]47,48] While the former is here negligible owing to the high solute-topolymer ratio used in this work, the latter has to be considered, given the dispersion of T 1 ( 1 H) at the atomic level (see Section VII in SI) and because t sat is here higher than T 1 ( 1 H) values.

Although bias measurements arising from relaxation are, in principle, assumed to be identical for both enantiomers, small variations in viscosity due to imperfect reproducibility of the mesophase preparation cannot be excluded. To overcome such pitfalls, a build-up curve approach is performed where anisotropic STD-NMR experiments are repeated with an incrementing t sat delay (see Figure 4). The amplification factor STD of a site i evolves as an asymptotic mono-exponential function of the saturation time, t sat , according to Eq. 4. [START_REF] Angulo | [END_REF]47,48] 𝑆𝑇𝐷

! !,! 𝑡 !"# = 𝑆𝑇𝐷 !,!"# !,! 𝑡 !"# 1 -e !! !"# !,! × ! !"# (4)
where k sat is the rate of saturation (equivalent to the slope of the curve) while the term

𝑆𝑇𝐷 !,!"# !,!
𝑡 !"# corresponds to the maximum amplification factor achieved by a given nuclei 𝑖 (asymptote of the curve). After fitting the experimental values according to Eq. 4, it is possible to estimate the value of 𝑆𝑇𝐷 ! !,! for a t sat tending towards zero, denoted 𝑆𝑇𝐷 ! !,! , by calculating the product 𝑆𝑇𝐷 !"# !,! × 𝑘 !"# !,! . [START_REF] Angulo | [END_REF]47] Thus, the 𝑆𝑇𝐷 ! !,! value is a quantity unbiased by the T 1 value of the site considered. This approach is a well-established procedure to extract quantitative data from STD 1D NMR experiments, and can be extended to anisotropic 1D NMR experiments. This build-up curve protocol is performed on the two enantiopure oriented samples and the very good agreement between the experimental data and the analytical expression of Eq. 4 is visible on Figure 4. Details about the goodness of the fit are available in Section VII.3 of the SI.

The 𝑆𝑇𝐷 ! !,! values for each site of (R/S)-BL are then determined and listed in Table 1, along with the associated fitting parameters. All the values are given with a 95% confidence interval. The 𝑆𝑇𝐷 ! values for both enantiomers are in the same order of magnitude, with comparable total 𝑆𝑇𝐷 ! (24.0% and 22.1% for (S) and (R)-isomers, respectively) reflecting a similar affinity towards the chiral polymer. Another common feature is the maximal 𝑆𝑇𝐷 ! reached for the hydroxyl group (H-2),

showing that H-bonding plays a central role in the polymer-solute interaction. Yet, the variation of 𝑆𝑇𝐷 ! values from site-to-site is quite different between the two enantiomers. This can be assessed by calculating the relative 𝑆𝑇𝐷 ! factors where the proton with the highest 𝑆𝑇𝐷 ! amplification factor is arbitrarily fixed at 100% and the values from the other hydrogens are then determined relative to this site. As shown in Table 1 and Figure 5b, the relative 𝑆𝑇𝐷 ! values are significantly different from one enantiomer to the other, in particular regarding the methine and methyl sites. This variation of epitope mapping confirmed by a quantitative STD approach highlights different orientations of the two enantiomers during the binding process. In other words, the enantiodiscrimination phenomenon is here mostly driven by diastereomorphous interactions between solutes in the bound state and the polymer -chains, i.e. 𝑆 !" ! ! ≠ 𝑆 !" ! ! , rather than a difference in affinity with respect to PBLG.

STD-NMR analysis of (R/S)-BL in PBLG.

While the STD-NMR data collected on separated samples have established differences of interactions between the enantiomers of BL with PBLG, we have investigated whether this difference still exists in racemic series. Indeed, a competitive binding process between both enantiomers may lead to different STD features than the ones observed in the enantiopure samples.

In this perspective, a STD experiment is performed on a racemic series of (R/S)-BL dissolved in PBLG/CD 2 Cl 2 and the subsequent STD spectrum shows significant signals for each hydrogen site. To screen STD differences between the two enantiomers, it is necessary to assign signals to

both. BL provides relevant spectral features for such a study as its enantiomers lead to significantly different ( 1 H-1 H)-RDCs and 1 H-RCSAs, so that some signals can be assigned unambiguously on 1D 1 H spectrum (see Figure 6). This assignment of each line has been supported by comparison with the 1 H signals measured on the enantiopure samples. Note, however, that the hydroxyl groups (sites H-2) of the enantiomers resonate at the same frequency due to fast chemical exchange. Thus, this signal cannot be here exploited to track STD differences for the two enantiomers (see Figure SI-10).

Comparison of anisotropic STD-NMR and reference spectra of (R/S)-BL points out variations of 1 H STD signals for each enantiomer. This is clearly observed in the methyl region (H-4) on Figure 6c where lines assigned to (S)-BL are less intense compared to the ones from the (R)-isomer. A similar observation (but less significant) can be done for the alcynic proton H-3 (see Figure 6b), while the STD signals at the site H-1 are identical for the (R)-and (S)-enantiomers (see Figure 6a). As in the case of enantiopure samples, these results highlight enantioselective interactions during the binding process with the PBLG side-chains in a racemic mixture. The enantiospecific solute-polymer complex shows geometrical differences reminiscent of those observed previously, but with some differences.

In one hand, both enantiomers in the bound state are oriented so that the methyl group is the farthest site from the polymer, and once again, this feature is more pronounced for the (S)-isomer.

In another hand, the identical STD signals for both enantiomers at the site H-1 suggest methine at a same distance from PBLG, which is not the case in enantiopure sample where different 𝑆𝑇𝐷 ! values have been determined (7.5 vs. 5.6 %) as seen in Table 1. Therefore, the presence of both enantiomers dissolved in a same chiral mesophase still leads to enantioselective interactions with respect to PBLG, but with slight changes in the orientation of enantiomers at the vicinity of the polymer.

Unsurprisingly, this affects the magnitude of RDC( 1 H-1 H)s in both oriented samples. The absolute values of the total couplings constants,

T( 1 H-1 H) = J( 1 H-1 H)+ 2D( 1 H-1 H),

STD-NMR analysis of (R/S)-BL in the achiral PBG phase.

First STD results collected with an enantiopure or racemic mixture of BL clearly reveal again interactions the polypeptide chain with a noticeable difference of epitope mapping between the complex (R)-BL/PBLG and (S)-BL/PBLG. Combined with the molecular shape anisotropy of BL, these of intermolecular interactions in the "bound" complex participate to the enantiodiscrimination mechanisms involving the chirality of the solute and polymer.

An interesting situation exists when the enantiomers of (R/S)-BL are dissolved in a mesophase made of a racemic mixture of PBLG and PBDG, its enantiomers (right and left handness helix). In the fast exchange regime for the solute, this orienting system becomes achiral by compensation, preventing all enantiodiscrimination phenomena to be observed in average. [49] Under these conditions, one might expect similar STD spectra for both enantiomers in such an achiral medium.

To answer to this intriguing question, From practical point of view, the routine 1 H STD pulse sequence could be applied with no peculiar modification (see Figure SI-2). In the "on-experiment", polymer spins were saturated by a train of 50 ms Gaussian selective pulses -applied during the preparation step -with a total time corresponding to the saturation time t sat . The selective pulse frequency offset was fixed to selectively saturate the polymer nuclei while leaving solutes undisturbed. The choice of this frequency offset was optimized to yield maximal STD amplification factors (see Figure SI-3). The off-experiment was carried out in exact similar conditions but in the absence of saturation. In order to limit the impact of instrumental instabilities of the spectrometer, the acquisition of "on/offresonance" spectra can be alternated.

To deal with the 1 H NMR contribution from polymer, integration was performed after a tailored baseline correction, i.e. Whittaker Smother algorithm (provided by MestreNova). This procedure enabled more repeatable integration values than the one obtained after a CPMG (Carr-Purcell-Meiboom-Gill) filter. [50] The latter option suffered from phase distortions due to the difficulty of cancelling the total spin-spin coupling modulation (T HH ) even for short echo times.

Conclusion

Establishing and then prioritizing the key molecular features governing the molecular shape recognition mechanisms involved in enantiodiscrimination phenomena in lyotropic CLCs is essential for two reasons: [51,52,53] i) optimize computational predictions (such as molecular dynamics simulation) of existing enantiodiscriminating helically-chiral polymers with ultimate goal of solving the thorny challenge of absolute configuration determination, ii) design novel polymerbased effective enantiodiscriminating systems.

So far the majority of experimental studies dedicated to this purpose are based on the evaluation of molecular order parameters for each enantiomer (Saupe's matrix parameters) and the analysis of their orientational difference (through the comparison of alignment tensors: variation of principal axis direction, 9D angle, …). In this work, we propose a paradigm shift by directly characterizing the transient complex formed by the enantiomers and the orienting chiral polymer with the help of 1 H STD-NMR experiments.

Interestingly, we demonstrate that this approach allows to experimentally determine the active interaction sites between enantiomers and the PBLG side-chains and how enantiomers can spatially interact differently with them. Furthermore, this method supported by a build-up curve approach, enables a direct and unbiased comparison of the orientation of enantiomers at the vicinity of the polymer side-chains.

This first-ever experimental STD study offers great hope and paves the way to the screening of series of model molecules in order to evaluate their interactions with accessible fragments of polymer as PBLG, but also with other polypeptide chiral polymers (such as PELG or PCBLL [54,55] or new polymers more recently designed such as PSMBLG, PBPMLG, …), [56,57] as well as other enantiodiscriminating chiral polymer families such as polyacetylenes. [19,22,58] Finally, to further study pairs of enantiomers dissolved in a single oriented sample, including STD scheme into bi-dimensional NMR experiments should be valuable in case of complex an overlapped 1 H STD spectra. An alternative strategy involving proton-decoupled 13 C-NMR detection of STD spectra could also be very effective, not least because the 13 C-{ 1 H} signals of enantiomers in the PBLG phases can be discriminated on the basis of differences of residual 13 C chemical shift

Figure 1 .

 1 Figure 1. (a) Schematic principle of anisotropic STD-NMR using PBLG polymer as CLC and (S)-PL as solute. K D and t res correspond to the dissociation constant and the time-residence of the transient polymer-solute complex, respectively, while LC stands for liquid crystal. (b) One example of anisotropic 1 H NMR (top) and anisotropic 1 H STD-NMR spectrum (bottom) recorded in PBLG/CD 2 Cl 2 (295 K) at 600 MHz. Here the (S)-PL is used as chiral solute. Note the very broad 1 H signal (centered around 7.3 ppm) corresponding to the aromatic 1 H sites of the PBLG side-chain in 1b (top). A baseline correction is applied on the STD spectrum (bottom). Weak negative signals for phenyl and methyl groups are artefacts arising from imperfect on/off-spectra subtraction.

  H STD signals can be neglected, and hence the 1 H STD signals are not biased by molecule orientation in PBLG-based lyotropic systems.

Anisotropic 1 H

 1 Figure 2a). In particular, no STD signals are detected on the spectrum of HL except for the OH group (see Figure2c), showing a poor affinity of this saturated alcohol with PBLG (see FigureSI-1). In the case of the aromatic alcohol, PL, only relatively small STD signals arise from the hydrogen sites bonded to the stereogenic center or very closed (methine and methyl groups, respectively), while no STD signals is detected from the phenyl group, even increasing the number of scans.

Figure 2 .

 2 Figure 2. (a) Generic structure of model chiral molecules studied here where the R 4 substituent is a butyl, phenyl and ethynyl group. The color code used for each substituent (-OH (black), -H (red), -CH 3 (blue), -R 4 (green)) is applied throughout the article and in SI. (b) Crude schematic representation of the molecular shapes of four substituents around the stereogenic center. (c) Colorized view of hydrogen sites showing a significant 1 H STD effects in enantiomers of HL, PL and BL, depending on the intensity of their associated STD signal.

Figure 3 .

 3 Figure 3. (a) Reference anisotropic 600 MHz 1 H NMR spectra of the (S)-BL (a) and (R)-BL dissolved in PBLG/CD 2 Cl 2 (275 K), respectively. (b) 1 H STD-NMR spectrum of BL obtained after subtraction of spectra recorded with/without saturation of PBLG. The 1 H STD experiments were carried out here with a t sat of 6 s. Baseline correction was applied on both reference and STD spectra to remove the PBLG signal contribution. The 1 H STD spectra are displayed with a ×5 vertical zoom.

Figure 4 .

 4 Figure 4. (a, b) Build-up curves providing STD % of (S)-BL and (R)-BL as a function of the saturation time (t sat ).

Figure 5 .

 5 Figure 5. (a) Plot of variation of STD 0 (%) versus the molecular sites (H-1 to H-4.) for the (R)-and (S)-isomer of BL in PBLG, recorded separately. Error bars correspond to 95% confidence intervals. (b) Epitope mapping of the (S)-and (R)-BL binding the polymer side-chains for sites H-1 to H-4. Values correspond to the relative STD 0 % after build-up curve fitting.

  within the methyl pattern (A 3 MX spin system) range from 𝑇 A 3 𝑅 (𝑒𝑒 = 100%) = 219 Hz, 𝑇 AM 𝑅 (𝑒𝑒 = 100%) = 92.9 Hz and 𝑇 AX 𝑅 (𝑒𝑒 = 100%) = 19.7 Hz to 𝑇 A 3 𝑅 (𝑒𝑒 = 0%) = 172.8 Hz, 𝑇 AM 𝑅 (𝑒𝑒 = 0%) = 76.8 Hz and 𝑇 AX 𝑅 (𝑒𝑒 = 0%) = 15.0 Hz for the (R)-isomer, and from 𝑇 A 3 𝑆 (𝑒𝑒 = 100%) = 83.4 Hz, 𝑇 AM 𝑆 (𝑒𝑒 = 100%) = 54.9 Hz and 𝑇 AX 𝑆 (𝑒𝑒 = 100%) = 20.5 Hz to ( 𝑇 A 3 𝑆 (𝑒𝑒 = 0%) = 64.9 Hz, 𝑇 AM 𝑆 (𝑒𝑒 = 0%) = 50.2 Hz, 𝑇 AX𝑆 (𝑒𝑒 = 0%) = 15.9 Hz for the (S)-isomer. Thus, the (R)-and (S)-isomers show a decrease of about 20% in average in all three coupling constants, with a variation dropping to 10% for the second coupling constant T AM for the (S)-isomer.

Figure 6 .

 6 Figure 6. Comparison between anisotropic reference (top row) and STD spectra (bottom row) recorded on a racemic sample of (R/S)-BL dissolved in a PBLG/CD 2 Cl 2 mesophase. The STD experiment is performed in 32 scans with a 𝑡 !"# of 6 s. Zooms on the (a) methyl, (b) methine and (c) ethynyl group regions are given. To ease the comparison, STD spectra are displayed at different scales as specified in brackets. Lines from (S)-and (R)-isomers are tagged by blue solid circles and orange stars, respectively. The assignment shown is obtained by comparison with the signals observed on the enantiopure samples.

  Each enantiomer shows significant STD signals with amplification factors higher than 5% for all the sites at a saturation time t sat of 6 s, with however,

	variations of relative STD signals that may reflect different geometries of "enantiomer-polymer
	complex".

C (see Figures SI-14 and SI-15) or 2 H (see Figure SI-16).

Table 1 .

 1 Site

		STDmax	ksat (s	-1 )	STD0 (%)	Relative	STDmax	ksat (s	-1 )	STD0 (%)	Relative
						STD0 (%)					STD0 (%)
	H-1	17.0 ± 0.8	0.44 ± 0.07	7.5 ± 1.2	80.0	12.3 ± 0.8	0.46 ± 0.10	5.6 ± 1.3	67.3
	H-2	18.2 ± 0.1	0.52 ± 0.02	9.4 ± 0.3	100	12.7 ± 0.6	0.66 ± 0.16	8.4 ± 2.1	100
	H-3	16.6 ± 2.0	0.32 ± 0.09	5.3 ± 1.7	56.0	12.6 ± 0.7	0.42 ± 0.07	5.3 ± 0.9	64.3
	H-4	7.8 ± 1.6	0.24 ± 0.10	1.8 ± 0.9	19.7	9.4 ± 0.8	0.31 ± 0.06	2.9 ± 0.6	34.8

-specific STD 0 (%) obtained for the two enantiopure oriented samples.

(S)-BL (R)-BL

Sites

a STD0 (%) values are calculated after confidence interval.

  1 H STD-NMR experiments are carried on (R/S)-BL oriented in the PBG achiral mesophase (PBG/CD 2 Cl 2 ) (see Table SI-1). Only the hydroxyl groups give rise to a STD signal, unlike to the other hydrogen sites. This result is very different from those obtained with a chiral mesophase. What that suggests remains a matter of hypothesis at this level. Samples are approximately prepared with ~15 mg of enantiomer each, ~100 mg of PBLG (DP = 800) and ~600 mg of dry CD 2 Cl 2 with m PBLG /m tot of ~14 %. Details for the sample

	One potential explanation relies on the three-sites exchanging system whereby each enantiomer
	(R or S) is in fast exchange with the L-and D-polymer. From this dynamic point of view, the time-
	residences of the different enantiomer-polymer arrays might significantly decrease, preventing any
	efficient intermolecular magnetization transfer between solutes and polymer, and hence the
	absence of detectable STD signals.
	Experimental section
	NMR sample.

compositions and their preparation (fire-sealed tubes) are reported in the Supp. Info (see Table

SI-

1). NMR experiments. All anisotropic 1 H NMR and 1 H STD-NMR spectra were recorded on a 14.1 T Bruker (Avance II) NMR spectrometer equipped by a 5-mm 2 H selective cryogenic probe operating at 600.13 MHz for 1 H. Specific experimental details are given in the figure captions or in the SI. If not otherwise specified, the sample temperature was set to 295 K.

Acknowledgments

P.L. and B.G. acknowledges the CNRS for its recurrent funding of fundamental research and the Université Paris-Saclay for its support, as well as Juliette Delay for her participation to this project in the frame of her first year of Master student's internship.

Data Availability Statement

The data that support the findings of this study are available in ESI and from the corresponding author upon reasonable request.

_______________________________________________________________________________________________

anisotropies ( 13 C-RCSA) (see 13 C-{ 1 H} 1D NMR spectra in SI). [1,2,[START_REF] Meddour | [END_REF]59] New investigations exploring these two options are currently in progress.

Conflict of Interest

The authors declare no conflict of interest.


Georges Gagneré
email: georges.gagnere@univ-paris8.fr

Cédric Plessiet
email: cedric.plessiet@univ-paris8.fr

Experiencing avatar direction in low cost theatrical mixed reality setup

Keywords: Computing methodologies → Computer Augmented acting, avatar direction, mixed reality, motion capture, performing arts

We introduce 1 the setup and programming framework of AvatarStaging theatrical mixed reality experiment. We focus on a configuration addressing movement issues between physical and 3D digital spaces from performers and directors' points of view. We propose 3 practical exercises.

CCS CONCEPTS

• Human-centered-computing → Interaction design → Empirical studies in interaction design; • Computer systems organization → Real-time systems → Real time system architecture; • Applied computing→ Arts and humanities→ performing arts •

We are sharing the results of a theatrical experiment based on relationships between performers and avatars in an environment mixing 3D digital space and physical « traditional » stage space. We organize the way in which two states of reality (the virtual world of 3D digital assets and the performer's physical world) work together for the pleasure of an audience. We focus in this paper on avatar acting issues and technological solutions we experienced to conduct theatrical artistic processes all along 2017, that resulted in public performances in December 2017 at Le Cube, digital creative center in Issy-les-Moulineaux, near Paris [START_REF] Gagneré | Challenges of movement quality using motion capture in theater[END_REF]. And we propose 3 practical exercises to enact the theoretical issues.

On the physical side of the mixed reality setup

Our research is rooted in the confrontation of a theatrical acting process and an avatar real-time motion capture control process on the same physical stage. We started by using a Kinect device, and we went on using one of the low-cost geo-spatial motion capture suits that appeared on the market around 2015 (fig. 1) [11]. The performer wearing the mocap suit is called the 'mocaptor' (fig. 2) -and we simply called the physical actor without a mocap suit in front of the avatar 3D screen the 'performer'. We'll therefore have onstage mocaptors, avatars and performers. At its inception in 2014, our research explored two hypotheses: on one hand, we would offer possibilities for the stage director and the audience to access the digital nature of 3D scenery and acting avatars by wearing HMD helmets; on the other hand, we would provide access to the digital space by projecting it onto a fixed screen at the rear of the stage [3] [4]. Because of the theatrical context of the 2017 experience, we adopted the second hypothesis. We allowed all the collaborators (performers, mocaptors, stage director, digital artist), but also the physical spectators, who are the fundamental part of the theatre live process, to share together the physical side of the mixed reality set up.

Keeping a visual feedback to control the avatar acting process

Our choice to work without HMD consequently prevents the mocaptor from accessing her digital body from an immersed point of view. She only can see her digital body through the 2D screen on the stage, or through « feedback » monitors placed downstage, which allow for rather more distant conditions of body awareness. Nevertheless, the acting issues from the immersive point of view [START_REF] Amato | Les avatars jouables des mondes numériques. Théories, terrains et témoignages de pratiques interactives[END_REF] [6], raised by the fact of inhabiting an avatar 'second body' and making her present to others (performer or audience), remains valuable. This situation is familiar in theatre through the figure of a puppeteer manipulating a puppet and maintaining a distant point of view on it. Wearing a mocap suit puts the mocaptor in the same situation, overseeing a 'natural' transfer of body movements. And we decided to keep her intimately involved in the control of her avatar by giving her a permanent visual feedback. • Space E: audience and stage director space The mixed reality stage results from interrelations between spaces A and B. The avatar's movements in B and the relationships between A and B depend upon the action of the mocaptor in C and the realtime play-out and operation in D. The resulting image of space B (on the video projection screen) has the same appearance as a 2D movie image shot with a camera. Nevertheless, it is fundamental to keep in mind that B is a (virtual) 3D space, similar to the physical stage in front of the video screen.

From the mocaptor to the avatar

We take from computer graphics conventions a way of describing the movement of a character [START_REF] Foley | Computer graphics. Principles and practices[END_REF]. The mocaptor performs relative movements with her body parts (legs, arms, head, etc.), in relation to a point called her 'reference' T (a starting point within the digital spatial grid). As we work in a digital space B, we need to independently set the reference T of the digital character, built by the software associated with the motion capture suit, in the coordinates system of B. This operation can completely change the position of the avatar in relation to the position of the mocaptor. Firstly, this fundamental separation between digital space B and physical space C is bodily difficult to deal with for a performer used to acting on a traditional stable stage. Secondly, we can control in real time the parameters of the avatar's T, that is deeply disorienting from the perspective of the mocaptor.

Thinking about the 3D nature of avatar space B, we first imagine an approximate analogical reproduction of the digital scenery in mocaptor space C in order to make the avatar movement easier for the mocaptor to perform. But it quickly became too difficult to implement because of multiplication of feedback constraints.

Reducing the mocaptor space C

Relying on the separation between spaces B and C, we reduce the scope of acting space C inside an absolute volume, that would be projected in space B relative to the digital scenery. We considered this smaller absolute volume in facilitating the establishment of the mocaptor's visual contact with the avatar through feedback monitors (fig. 4). Figure 5 shows one basic configuration we used during the 2017 artistic and pedagogical experiments. This setup will also be used for the 3 exercises. It consists in two corridors on each lateral side of the stage A with a feedback monitor on the downstage extremities. The mocaptors use the big video projection screen as an alternative feedback when they are walking upstage. This set-up optimizes the visual feedback arrangement, given that the mocaptors largely kept their head forward facing along the corridor. In this configuration the mocaptors are very close to the performers and keep an indirect but strong connection with the physical stage. It helps in sharing the same acting energy. The director can compare the mocaptors' movements and the results of their transposition on the avatars' bodies, so as to focus on efficient directorial responses with a view to enhancing expressivity.

MANIPULACTOR ASSISTANCE FOR CONTROLLING THE AVATAR

The manipulactor, a puppeteer of the mocaptor

Regarding the scenic actions on the physical stage A, we know that performers often act without keeping an absolute reference to the space. When they are in dialogue with each other, they often take care to offer their face to the audience. In doing so, they often must cheat on the effective line of sight of their partner, because of lighting positions or the orientation of scenery. For opera singers, the imperative need to project the voice towards the audience asks them to deal with 'artificial' spatial relations. They also must permanently keep in touch with the conductor through multiple video feedbacks all around the stage. Playing with the parameters of the avatar reference transform T (see 2.2) allows one to orient the reduced acting volume of the mocaptor in relation to scenic needs. We can use the metaphor of a puppeteer guiding the puppet through the stage, the difference being that the extent of the manipulation is not limited by physical constraints while we are in a digital and real-time-rendered space. We give the responsibility to guide the avatar to an assistant (of both the digital artist and the stage director), who should both understand acting rules and computing possibilities: we called him the 'manipulactor'.

AKeNe programming framework

The programming framework allowing the AvatarStaging experiments uses the AKeNe library [START_REF] Plessiet | Avatar Staging: an evolution of a real time framework for theater based on an on-set previz technology[END_REF]. The development is focusing on two main directions. On one side, we use and improve modules for combining movement data and solving motion retargeting issues. For instance, we use a puppeteering system that allows extraction of the mocap data to mix them with inputs coming from a gamepad, or even from AI videogame modules as pathfinder (fig. 6). On the other side, we use an event manager for cueing positions and actions in the digital scenery.

Actions and tool of the manipulactor

The manipulactor is finally able to guide the avatar by:

• moving in a forward/backward direction • moving laterally to the right or the left • moving up and down • rotating the yaw for changing her forward direction • rotating the pitch to put her on a horizontal position and make her float in the air All these actions can be done in addition to the mocaptor movements. We use a gamepad controller for applying them to the avatar, taking advantage of the powerful design of the device for combining and fine-tuning the appropriate transformations. The manipulactor focuses on two main goals:

• adjusting the scenic address of the avatar towards its performing partner in the A space, from the point of view of the audience; and respecting perspectival constraints [START_REF] Gagneré | Espace virtuel interconnecté et Théâtre. Une recherche-création sur l'espace de jeu théâtral à l'ère du réseau[END_REF] • accompanying the mocaptor in augmented movements that are usually not accessible to a performer, as, for instance, floating in the air.

PRACTICE: 3 EXERCICES

We finally describe three practical situations aimed both for demonstrating AvatarStaging setup and programming framework AKeNe, and for exercising as a mocaptor or manipulactor to achieve expressive avatar movements.

Situation 1: Walking

The first situation (fig. 7) consists in having avatars naturally walking and pausing in the digital space. Mocaptors and manipulactors shall collaborate to expand the constrained walking space in C to the larger space B, trying to keep a natural pacing. They will also be asked to propose basic acting interactions between the avatars, as interlacing trajectories or looking at each other.

Situation 2: Walking and watching

From the previous situation, we add a performer in front of the screen as a third partner (fig. 8). We'll explore the complexity of changing watching goal from the other avatar to the performer, and vice versa.

Situation 3: The crowd

We finally propose to improvise with a group of 6 avatars in motion (fig. 9). Each mocaptor controls 3 avatars (identical or not), and each avatar is guided by one manipulactor. In our experiments, we felt that the mocaptor is a kind of puppeteer inside the avatar, where the avatar has a "physiology" close to that of a wooden puppet skeleton (or a stylized robot). When the avatar is a 3D scan with imperfect skinning, the mocaptor has the feeling of living inside a kind of mascot -that's to say, a body mask (or human-sized doll). The action of the manipulactor is like that of the traditional puppeteer who is external to the puppet. One could then compare the relationship between mocaptor and manipulactor to that (extraordinarily) of a living puppet who would suddenly see some of her wires taken back by an outside hand. We therefore need to establish avatar control-sharing protocols for setting up specific actions (scenic address, large space routes, stage entries).

When the mocaptor (both puppeteer and puppet), walks under the manipulactor's control, she finds herself in the position of a golem, a mythological creature capable of movement but devoid of free will. We forecast to extend the mocaptor/manipulactor relationships by exploring other movement combinations and plausible movement qualities. Hybridisation with autonomous virtual actors is one direction [START_REF] Plessiet | Autonomous and Interactive Virtual Actor, Cooperative Virtual Environment for Immersive Previsualisation Tool Oriented to Movies[END_REF]. Addressing facial expression and splitting control in between several mocaptors is another (fig. 10).

Figure 1 :Figure 2 :

 12 Figure 1: Perception Neuron motion capture suit

Figure 3 :Figure 4 :

 34 Figure 3: AvatarStaging mixed reality setup

Figure 5 .Figure 6 .

 56 Figure 5. Configuration of AvatarStaging

Figure 7 .Figure 8 .

 78 Figure 7. Situation 1 -Walking

Figure 9 .

 9 Figure 9. Situation 3 -The Crowd


Georges Gagneré
email: georges.gagnere@univ-paris8.fr

Cédric Plessiet
email: cedric.plessiet@univ-paris8.fr

Andy Lavender
email: a.lavender@warwick.ac.uk

Tim White
email: t.white@warwick.ac.uk

Challenges of movement quality using motion capture in theatre

We describe 1 two case studies of AvatarStaging theatrical mixed reality framework combining avatars and performers acting in an artistic context. We outline a qualitative approach toward the condition for stage presence for the avatars. We describe the motion control solutions we experimented with from the perspective of building a protocol of avatar direction in a mixed reality appropriate to live performance.

CCS CONCEPTS

• Human-centered-computing → Interaction design → Empirical studies in interaction design; • Computer systems organization → Real-time systems → Real time system architecture • Applied computing→ Arts and humanities→ performing arts • Computing methodologies → Computer graphics → Animation → Motion capture

In this paper we focus on the generation and performance of avatars, who exist within a conventional screen space (featuring 3D in-world scenography) that must also be understood in co-relation to a three-dimensional theatre space and co-present performers, in a real-time moment of creation and interrelation. This raises questions concerning the 'avatarization' of the theatrical physical body and the emergence of new acting processes, from the point of view of the performing arts and the physical stage. Acting issues are indeed often explored in an autonomous perspective [START_REF] Batras | A virtual reality agent-based platform for improvisation between real and virtual actors using gestures[END_REF] [START_REF] Bevacqua | INGREDIBLE: A platform for full body interaction between human and virtual agent that improves co-presence[END_REF]. But it appears that even if the avatar is under human control, the plausibility of her presence remains an issue [10] and requires new approaches to tune interactions [START_REF] Gillies | What is Movement Interaction in Virtual Reality for?[END_REF].

AvatarStaging mixed reality setup

A physical actor, called the 'mocaptor', wears a motion capture geo-spatial low-cost system (fig. 1). She acts in space C and controls an avatar in digital scenery B. The avatar interacts with a performer through a 3D image projected in the rear of space A, in front of the stage director and audience in space E. Transformations in real time of the position of the avatar are performed both by the digital artist in space D, and the 'manipulactor' in space E, who uses a gamepad to modify the avatar reference transform T (the avatar's position) in space B. The manipulactor moves the avatar in a forward/backward direction, laterally to the right or the left, and up and down. She rotates the yaw for controlling the movement direction, and the pitch to put the avatar on a horizontal position. She focuses on two main functions:

• adjusting the scenic address of the avatar towards the performer in the A space, from the point of view of the audience; and respecting perspectival constraints [START_REF] Gagneré | Espace virtuel interconnecté et Théâtre. Une recherche-création sur l'espace de jeu théâtral à l'ère du réseau[END_REF] • accompanying the mocaptor in augmented movements that are usually not accessible to a performer, as, for instance, floating in the air. We have called this hybrid setup AvatarStaging and achieve it using the AKeNe library implemented in a videogame engine for controlling the avatars [START_REF] Gagneré | Experiencing avatar direction in low cost theatrical mixed reality setup[END_REF].

Pedagogical and professional use cases

We introduce below the relationships between mocaptors, manipulactors, avatars and performers in the context of two case studies. We consider the outcomes of practice-based research conducted by way of a series of interdisciplinary workshops in 2017 as part of a Paris 8 university research project entitled The Augmented Stage, focusing on the development and presentation of theatre and performance in and through digital technologies [START_REF]Masques technologiques: altérités hybrides de la scène contemporaine[END_REF]. The project involved collaboration between software programmers, digital designers and theatre makers and explored the experimental setup with students (fig. 2) and professional artists [START_REF]Agamemnon Redux » after Eschyle[END_REF]. Acting issues raised by inhabiting the second body of an avatar are central [START_REF] Ayiter | Avatar actors[END_REF] [9].

CASE STUDY N°1: DIALOGUE BETWEEN AVATARS AND PEFORMERS

The first case study is based on configuration 1 of the AvatarStaging set up, that shows two corridors C on each lateral side of the stage A with a feedback monitor on the downstage extremities (fig. 3), allowing the mocaptors to see the avatars created in-screen behind them. The mocaptors use the big video projection screen as an alternative feedback when they are walking upstage. This set-up optimizes the visual feedback arrangement.

From a small corridor to a large 3D scenery

The 3D space is larger than the length of the corridors. When an avatar crosses it from one door to the other, she follows this routine: at the extremity of the corridor, she stops her walk, turns, and during this rotation the manipulactor changes the yaw parameter of his avatar in the opposite direction, in order to counterbalance the rotation. The avatar seems to wait in situ. As soon as the U-turn is completed, the mocaptor goes on to her goal (the end-point of this movement trajectory) (fig. 4). To achieve a smooth and imperceptible counterbalancing, the mocaptor rotation must be precise at the outset, so that the manipulactor does not have to hesitate. It should be slow, allowing time for the manipulactor carefully to control the gamepad thumb stick. The relationship between the two partners was founded on an empathic collaboration geared towards a symbiosis of the avatar movement.

An empathetic collaboration for augmented acting

We know that the mocaptor prefers an empathetic relationship with the image of her avatar on the feedback monitors [START_REF] Gagneré | Traversées des frontières[END_REF]. That means that she looks at the image and 'translates' the point of view of the avatar: if the avatar faces her on the screen, she will go to her right to have the avatar moving to the left in the image, but to her own right in the relative 3D space. Performing a mental rotation symmetry, the mocaptor is adopting a disembodied self-location to take the avatar visuo-spatial perspective [START_REF] Thirioux | Walking on a line: A motor paradigm using rotation and reflection symmetry to study mental body transformations[END_REF]. The manipulactor should develop the same relationship with the avatar through the screen to understand where the avatar must go. When the link is broken for any reasons, it is hard to change the control rules and return to the sympathetic and reflective/symmetrical point of view.

Constraints in combining interactions

In this use case, the mocaptor faces three other acting partners: the other avatar inside the 3D scenery and the two physical performers on stage A. An important consideration in conventional theatre is the circulation of attention: who is at the centre of the respective actor's attention, and to whom should other partner actors pay attention? This is decided in common agreement between actors and director, whilst the means of presenting this focus remains a powerful tool of expressivity for the actor. The manipulactor organizes the system of attention so that the corridor axis provides the address to the performers in space A [START_REF] Gagneré | Experiencing avatar direction in low cost theatrical mixed reality setup[END_REF]. The mocaptor ascertains the direction of her avatar partner in space B with the help of the feedback monitor. Indeed, with the audience watching the same image, the mocaptor consequently only needs to imagine around her the digital scenery, respecting the feedback image, and find the position of her partner situated in the same scenery (fig. 5 and6). The mocaptor has three choices at any moment: looking in the direction of the corridor axis to have a contact with the onstage performer; looking in the direction of the eyeline that matches the imagined avatar partner; or looking at another point to express a personal feeling deliberately disconnected from her partners, which offers a rich expressive variety. This requires a deep concentration in the continuous collaboration between mocaptor and manipulactor to maintain the integrity and consistency of the movement intentionality of the avatar.

Who is finally controlling the avatar?

Walking within a small corridor necessarily gives responsibility for the movement orientation to the manipulactor -that's to say, an operator who is offstage. We made the same conclusion about the necessity of keeping the scenic address of the avatar in relation to the performers: only the manipulactor can achieve this because of her distance from the stage. However, guiding the mocaptor in this way (at least, by intervening in the movement of the avatar) does not deprive her of initiative. On the contrary, close attention to the avatar's body language helps the manipulactor to understand the intentionality of the movement. Sharpening her avatar body analysis, the manipulactor succeeds in catching the mocaptor's desires that increase the plausibility of the avatar's presence [10]. From the mocaptor's point of view, the difficulty was to maintain a degree of liberty from the defining linear axis of the corridor: when was it pertinent to look aside and break the scenic address built by the manipulactor? Contradictory moves can easily break the spatial continuity of the avatar's acting in relation to her partners and the audience. This remains a live issue that must continually be kept in mind to present a seamless, spatially coherent performance. We are facing a symbiotic but unpredictable improvisation, balanced by a common understanding of the objectives of the scenic actions, driven by the stage director. The dialogue between both human controllers seems to be deeper when it is formulated in term of movement quality instead of specific goals [START_REF] Fdili Alaoui | From dance to touch: movement qualities for interaction design[END_REF]. However, the grammar of these new interactions needs to be consolidated.

CASE STUDY N°2: FUSION OF PERFORMER AND MOCAPTOR

Configuration 2 illustrates the case when spaces A and C are juxtaposed, transforming performers into mocaptors (fig. 7). This is the AvatarStaging configuration set-up for case study N°2 [START_REF]Agamemnon Redux » after Eschyle[END_REF], with no side corridors dedicated to the control of the avatar. The performers have one feedback monitor downstage and use the video projection screen upstage for visual feedback when facing away from the audience. This required manipulactors and mocaptors to sharpen their skills to maintain a plausible effect of liveness in the avatars in-screen.

Movement contextualization

Case study N°2 entailed a parallel presentation at any moment. The performers presented their characterization directly to the audience, rather than to each other -that's to say, they did not interact in terms of the normal spatial relationships you would expect between performers onstage; meanwhile they created the movements of the avatars in the virtual scenography, and a plausible (realist-oriented) spatial relationship between the avatars in-screen. This was made possible by experienced mocaptors being able to play two levels of scenic reality at once, and manipulactors able to adapt, on the fly, the avatar movements to the 3D situation. We presented two dialogue situations in very different contexts: a banquet with two robot figures in a cold white room (fig. 8), and few minutes later a bath with a naked warrior caressed by a woman (fig. 9), played by the same mocaptors. In each situation, we notice how the body of the avatar deeply impacts the way the audience receives the performed actions. All these different nuances of movement, coming from the actions of two performer/mocaptors who were required to operate within a restricted physical space, offer a rich field of body-movement research concerning relations between live action, characterization, real-time avatar generation and dramatic/narrative development.

Opening dramaturgical possibilities

An interesting consequence of this set-up was to open for the spectator the larger meaning of interrelated body movements, which are distinct but co-created across different kinds of space. This offers new dramaturgical possibilities both for the stage director and the performer/mocaptor. It led us to the idea of changing the point of view within any particular scene, using the powerful real-time programming features currently offered by the video game engine. Fig. 10 shows four different shots we built to run one scene of the show. This second way of contextualizing avatar movements is also challenging for the mocaptors, who must play virtual eyelines and relationships from an isolated position. In future, we aim to formalize the protocols between the stage director, the mocaptor, the manipulactor and the performer that are needed to establish good practices for avatar direction.

Figure 1 :

 1 Figure 1: AvatarStaging mixed reality setup

Figure 2 :

 2 Figure 2: Masterclass in Conservatoire National Supérieur d'Art Dramatique (Paris)

Figure 3 .Figure 4 :

 34 Figure 3. Configuration of case study N°1

Figure 5 .Figure 6 .

 56 Figure 5. Case study N°1 -scenic eyelines

Figure 7 .Figure 8 .

 78 Figure 7. Configuration of case study N°2

Figure 9 .

 9 Figure 9. Case study N°2 -the bath

ACKNOWLEDGMENTS

This research was made within the framework of "Scène Augmentée" research project, supported by Labex Arts H2H, Investissements d'Avenir French Program, with its partners: Paris 8 University laboratories INRéV and Scènes du monde, création, savoirs critiques, the Conservatoire National Supérieur d'Art Dramatique de Paris, compagnie Gente Gente!, Le Cube -Centre de création numérique, Issy-Les-Moulineaux, Warwick University, and didascalie.net.


Hyojun Lee

Amélie Jacquot

Dominique Makowski

Margherita Arcangeli

Jérôme Dokic

Pascale Piolino

Marco Sperduti
email: marco.sperduti@u-paris.fr

Beauty is in the eye of the beholder: evidence from a common mnemonic advantage between aesthetics judgement and self-reference

Keywords: aesthetics judgement, beauty, self, memory

A long-lasting and unresolved debate in the field of aesthetics is whether beauty is inherent to the object of appreciation or to the subject contemplating it. Several studies suggest that physical features (e.g., symmetry, contrast) of an artwork influence aesthetic rating.

Nevertheless, this objectivist approach fails to explain the idiosyncratic nature of aesthetic experiences (AE). Recent models propose a multi-process account of AE, integrating a subjective evaluation based on self-referential processing. This proposition seems coherent with neuroimaging studies showing activation of a common neural network during AE and self-reference. Nevertheless, behavioural data supporting this hypothesis is missing. We took advantage of the self-reference effect (SRE) in memorythe mnemonic advantage for material encoded in a self-related mode -to test the hypothesis that aesthetic judgement is based on self-related processes. We predicted that if aesthetic judgement recruits selfreferential processing, incidentally encoding artworks in this condition should produce a similar mnemonic advantage as the SRE. To test this hypothesis, 30 participants incidentally encoded 60 painting in three conditions: self-reference, judgement of beauty and judgement of symmetry (control condition). We found that items encoded in the aesthetic judgment condition were as well recognized as those encoded in self-reference condition when participants gave extreme judgements on the beauty scale during encoding. These findings suggest that at least intense AEs ' f f.

Introduction

A long-lasting and unresolved debate in the field of aesthetics is whether beauty is inherent to the object of appreciation or to the subject contemplating it. In other words: is beauty in the eyes of the beholder? In recent years, the scientific interest for the foundation of aesthetic experiences (AE) has been renewed by the emerging field of neuroaesthetics. Mirroring this philosophical and historical debate, there have been two main approaches explaining aesthetic appreciation in modern neuroaesthetic research. The dominant research endeavour in this field, that we can call the objectivist approach, has tried to determine the physical features of an artwork influencing aesthetic judgement. Another, more neglected, line of research has investigated the subjective factors modulating aesthetic judgement.

The objectivist approach has shown that various physical properties of an object are reliable predictors of AE. The symmetry of an artwork is considered a stable and robust predictor of aesthetic preference. Indeed, symmetry positively influenced aesthetic preference for geometric shapes, and this effect was additionally resistant to familiarisation (Tinio & Leder, 2009). In another study, a mild disruption in symmetry resulted in a significant decline in aesthetic preference for geometric shapes (Gartus & Leder, 2013). Complexity also appears as influencing aesthetic preference: its effect on aesthetic judgement has been found for abstract and representational artwork [START_REF] Osborne | The relationship between aesthetic preference and visual complexity in absract art[END_REF][START_REF] Roberts | Complexity and aesthetic preference for diverse visual stimuli[END_REF], or for geometric shapes (Tinio & Leder, 2009). In the same vein, aesthetic preference was greater for photographs with higher level of fractal dimensions (Spehar et al., 2003). Regarding visual contrast, participants' preference for abstract and representational paintings was greater when the contrast was adjusted higher than the original level, compared to a lowerthan-original contrast, independently f j ' and social status (Van Dongen & Zijlmans, 2017). Another study, providing further understanding of this effect, suggested the existence of an 'optimal level of contrast in paintings' most preferred by viewers (Dijkstra & van Dongen, 2017). Abstract paintings were most appreciated when the contrast was moderately higher than the original, but not excessively. Curvature and angularity are also properties that influence aesthetic judgement. Higher preference for curved shapes and polygons over angular ones has been reported [START_REF] Bertamini | Do observers like curvature or do they dislike angularity[END_REF][START_REF] Silvia | Do People Prefer Curved Objects? Angularity, Expertise, and Aesthetic Preference[END_REF].

Aesthetic judgement is also affected by the content of the artwork. Some studies observed a higher appreciation among the general population for representational compared to abstract art [START_REF] Roberts | Complexity and aesthetic preference for diverse visual stimuli[END_REF][START_REF] Sidhu | Prediction of beauty and liking ratings for abstract and representational paintings using subjective and objective measures[END_REF], and more generally for real-world scenes compared to abstract images (Vessel & Rubin, 2010). Taken together, these findings show that the physical features of visual stimuli j ' j .

These studies are grounded in cognitive models trying to isolate single key factors determining AE (e.g., [START_REF] Berlyne | Aesthetics and Psychobiology[END_REF][START_REF] Reber | Processing Fluency and Aesthetic : B ' Ex ? Personality and[END_REF], and they give precious information on the physical characteristics of an artwork influencing AE. Nevertheless, they probably fail to capture the complexity of the phenomenon, neither they explain the widespread intuition that AE is somewhat subjective. An intuition that seem confirmed by experimental studies

showing that there is low inter-individual agreement on aesthetic response to visual artistic stimuli, suggesting that aesthetic experience is highly subjective (Vessel et al., 2012). To account for this subjective variability, more recent models propose a multi-process account of AE [START_REF] Leder | A model of aesthetic appreciation and aesthetic judgments[END_REF][START_REF] Leder | Ten years of a model of aesthetic appreciation and aesthetic j : T -Developments and challenges in empirical aesthetics[END_REF]. Leder and collaborato ' model proposes five main processing stages leading to aesthetic judgement: perception, implicit memory integration, explicit classification, cognitive mastering, evaluation, and continuous emotion evaluation. By assessing dimensions, such as the evaluative one, pertaining to elements unrelated to the object, these models account for AE beyond the simple elaboration of physical properties. Critically for the present work, the last stages of this model, cognitive mastering and evaluation, account for the subjective component of aesthetics judgement. In particular, the authors propose that self-related cognitive information could be a gateway in understanding and evaluating an artwork. Anecdota " (Leder et al., 2004, p.499). Thus, they explicitly link AE with self-related processing associated to autobiographical information.

[…] f w f ' G z ' k f j '"
The link between AE and the self seems to be sustained by the fact that many people consider their artistic taste to be an important part of their identity, their sense of who they are (Vessel et al., 2013).

These observations echo neuroimaging findings showing that key regions of the default mode network (DMN) are activated during aesthetic judgement [START_REF] Jacobsen | Brain correlates of aesthetic judgment of beauty[END_REF][START_REF] Kawabata | Neural Correlates of Beauty[END_REF]Martín-Loeches et al., 2014;Vessel et al., 2012Vessel et al., , 2019)). In particular, the medial prefrontal cortex (mPFC) is of utmost relevance. Importantly, the DMN and the mPFC are known to underpin self-representation at different levels of abstraction (for a meta-analysis, see Martinelli et al., 2013). Nevertheless, it is necessary to avoid haphazardly associating two cognitive processes only on the basis of shared cortical activations. Behavioral data suggesting a possible link between the self and AE also exist. For example, a large corpus of literature demonstrates the importance of familiarity on the aesthetic judgement of proverbs, human faces, and music [START_REF] Bohrn | When we like what we know -A parametric fMRI analysis of beauty and familiarity[END_REF]Bornstein, 1989;[START_REF] Park | Roles of familiarity and novelty in visual preference judgments are segregated across object categories[END_REF][START_REF] Schubert | The influence of emotion, locus of emotion and familiarity upon preference in music[END_REF]Verhaeghen, 2018). Moreover, it is interesting to note that some studies reported that aesthetic judgement during incidental encoding lead to increased memory performance for representational and abstract art pictures [START_REF] Nadal | The Search for Objective Measures of Aesthetic : T f T[END_REF], for photographs of real-world scenes (Choe et al., 2017), and for paintings [START_REF] Ishai | Perception, memory and aesthetics of indeterminate art[END_REF]. Some authors proposed that this effect could be due to the fact that aesthetic judgment may have increased self-related processing (Choe et al., 2017). Indeed, it is well known that items requiring a self-related processing gain a robust mnemonic advantage, in comparison to other types of treatment (e.g., semantic processing), an effect known as self-reference effect (SRE) in memory (Conway, 2005;Cunningham et al., 2008;[START_REF] Kalenzaga | Episodic memory and self-f : Insights from an fMRI study in younger and older adults[END_REF]Leshikar et al., 2015;Sui & Humphreys, 2015;Symons & Johnson, 1997). Nevertheless, no study to date directly compared the mnemonic advantage produced by aesthetic judgement and self-reference in order to investigate the potential existence of a common mechanism organizing AE and the self-representation.

To test this hypothesis, we asked participants to incidentally encode artworks in three conditions: an aesthetic judgement condition, a self-referential judgement condition, and a control condition (judgement of symmetry). Our main hypothesis was that if AE is linked to self-referential processing, we should find a comparable mnemonic advantage for the selfreferential and the aesthetic judgement condition, compared to a control condition requiring judgement of low-level visual features. An exploratory and complementary hypothesis was that the mnemonic advantage for item encoded in the aesthetic judgement condition should ' .

, accordingly to a recent study showing better memory performance for the location of paintings that elicited extreme aesthetic experiences, whether positive or negative [START_REF] Babo-Rebelo | Looking, liking and locating : An experimental aesthetics of orienting[END_REF], we made the hypothesis that paintings receiving judgments at the two poles would receive the greatest mnemonic advantage.

Material and Methods

Participants 30 university students (27 women; mean age 20.7  2.76 years) were recruited for this study.

The participants were undergraduate students in psychology at the University of Paris. All participants had a normal or corrected vision. No participant showed expertise in art, based on the Aesthetic Fluency Scale [START_REF] Silvia | Knowledge-f x : Ex fluency[END_REF][START_REF] Smith | The Nature and Growth of Aesthetic Fluency[END_REF]; mean score 11.63  4.69 out of 40). All participants were informed of the academic nature of the study and accepted that their responses would be processed anonymously. After the nature of the procedure had been fully explained, all participants gave written informed consent before carrying out the study. The protocol was carried out in accordance with the local ethical standards.

Material Visual stimuli

One hundred pieces of visual art were selected from the Wiki Art data base, across nine different artistic styles representing some of the most important styles between the 16 th and 20 th century: Nordic renaissance art, Baroque art, Rococo art, Romanticism, Realism, Symbolism, Expressionism, Impressionism, and Post-impressionism. We only selected color and representational paintings with a landscape width-height ratio. We excluded painting including easily recognizable elements (e.g., ' signature, writings). A complete list of the painting is presented in the Supplementary Material 1. Among the selected 100 pieces, 60

were used as target stimuli during the encoding phase. The remaining 40 were used as distractors in the recognition phase. The distractors were visually paired with some of the target stimuli in terms of content (people, animals, landscape, style etc.) and color schemes, in order to make sure they were not dissimilar. The two groups of stimuli did not show any

Encoding phase

There were three within-subject experimental conditions (encoding conditions): an aesthetic judgement condition, a self-referential judgement condition, and a symmetry judgement condition (control). In the aesthetic judgement condition, the subjects were asked to judge their appreciation of the (" w f ") f 0 0.

In the self-referential judgement condition, the subjects were asked to judge to what degree

(" w f ") f 0 0. j on, the j w k j ' f (" w ") f 0 0. w f f f w test (incidental encoding).
Prior to the beginning of the encoding phase, a painting not employed in the experimental

k ( . . V k V ' "T B B " w )
has been used to instruct participants. The self-reference condition was explained to the subjects with the example f w w : "A have experienced the event represented in this painting, it is possible that the painting reminds you personal memories such as a friendly hillside picnic, a museum visit where you may have seen similar paintings, a scene of a movie, or even a visual representation of a story ". T w x j w asking them to simply judge how subjectively beautiful they found the image. The symmetry condition was explained with this image by explaining that although some elements of the painting are quite symmetrical (e.g., the shape and colors of the hill and the sky), some other elements are not (e.g., the left-heavy way people are disp ' appearances, the diagonally placed central weapon, etc.), and that they could judge the global symmetry in their own way.

Each condition contained 20 target images presented in a block. The distribution of the 60 target stimuli on the encoding conditions was counterbalanced across participants, so that each item was presented in each experimental condition. The order of block was randomized across subjects. In each block, a trial started with a fixation cross for 500 ms that was followed by the presentation of a stimulus for 3 seconds. Once the stimulus presentation ended, the rating scale appeared on the screen, where the subjects had to enter their score according to the experimental condition. There was no time limit for the evaluation. Once the participants responded, the next trial started. The end of the block was signaled by the presentation of written instructions for the next block.

Recognition phase

During the recognition phase, all target stimuli were presented intermixed with 40 distractor stimuli in a random order. After the presentation of the fixation cross (500ms), each stimulus was presented for 3 seconds. Once the stimulus presentation was over, participants had to indicate if they had seen the picture before. They could choose between 3 different responses : "Y " " " " ". If the w w "Y " " " participants were asked to indicate in which encoding condition they had seen the image. There was no time limit to answer. Once the recognition response was entered, the fixation cross appeared, followed by the next trial.

Procedure

The experiment was conducted at the MC²Lab, located at the Paris University Psychology

Institute. Participants were invited to an experimental room, where they were seated at approximately 40cm in front of a 14-inch computer screen (1920x1080, 60Hz). The screen was adjusted to maximum brightness for all participants. The experiment, implemented in Psychopy v3.1.1 [START_REF] Peirce | PsychoPy-Psychophysics software in Python[END_REF], was conducted in three parts for all participants in this order:

the encoding phase, the retention interval, the recognition phase. The duration of the retention interval was about 30 minutes (30.32±8.13 minutes). During this phase, participants filled in four questionnaires1 , and watched an 8-minute short film (Alike, Lara & Cano Méndez, 2015).

All the above listed material except for the Aesthetic Fluency Scale was used solely as a way to guarantee a sufficiently long retention interval. The results for these questionnaires were thus not analyzed.

Data analysis

The first two sets of analyses model the item (Yes responses) and source recognition (a binary variable) as a function of the condition (3 levels; Beauty, Self-reference and Symmetry) using mixed logistic models (participants and items were entered as random factors).

Marginal means-based contrasts were then estimated to allow us exploring the pairwise differences between the levels. In the second part, we additionally modelled the effect of the rating during encoding (a continuous variable ranging from 0 to 10), in each condition, for item and source recognition, allowing to investigate possible non-linearity using second order polynomials.

Data processing was carried out with R (https://www.r-project.org/) and the easystats suite (Lüdecke et al., 2019;Makowski, Ben-Shachar, & Lüdecke, 2019). The whole analysis was performed under the Bayesian framework using MCMC sampling with the rstanarm package (Goodrich et al., 2018; http://mc-stan.org/). To assert effect significance, we used the Probability of Direction (effects wer " f " w pd > 97%), a Bayesian equivalent of the p-value (Makowski, Ben-Shachar, Chen, et al., 2019). For clarity, only the relevant effects will be described in the text, but the full reproducible analysis script (containing the full description of all models along with complementary results and figures as well complete descriptive statistics) is available in Supplementary Materials 2.

Results

Effect of Condition

The mixed logistic model predicting the item recognition had a total explanatory power (Bayes R2; Gelman et al., 2019) of 22%, from which 3% (marginal R2) were related to the effect of the condition alone. Within this model, the Self-reference condition led a significantly higher item recognition probability than the Beauty (difference = 0.91, 95% CI [0.61, 1.23], pd = 100%) and the Symmetry (difference = 1.10, 95% CI [0.80, 1.40], pd = 100%) conditions. The difference between the latter two was not significant (difference = 0.18, 95% CI [-0.08, 0.46], pd = 90.75%). See Figure A1.

The mixed logistic model predicting the source recognition had a total explanatory power of 10%, from which 3% was related to the effect of the condition alone. Within this model, there were no significant differences between any of the conditions (Beauty -Self-reference, difference = -0.28, 95% CI [-0.58, 0.03], pd = 96.35%; Beauty -Symmetry, difference = -0.

22, 95% CI [-0.50, 0.13], pd = 91%; Self-reference -Symmetry, difference = 0.06, 95% CI [-0.25, 0.41], pd = 65%). See Figure B1.

Effect of Rating

The mixed logistic model predicting the item recognition had a total explanatory power of 24%, from which 5% were related to the condition and the rating. Within this model, only the rating of Symmetry displayed a significant linear positive relationship with the probability of item recognition (median = 12.50, 95% CI [2.79,23.14], pd = 99%). However, the rating of Beauty had a significant quadratic relationship (median = 24.29, 95% CI [14.02,36.62], pd = 100%), with middle ratings leading to a lower probability of item recognition. Additionally, contrast analysis confirmed that at the rating extremities (0 and 10), the difference between the Beauty and the Self-reference was not significant (difference = 0.55, 95% CI [-0.34, 1.52], pd = 87.92%; difference = 0.55, 95% CI [-0.60, 1.69], pd = 82.85%, respectively). See

Discussion

In this study, we investigated the relationship between AE and the self at the behavioral level.

We tested the hypothesis that AE is grounded on self-reference by examining the common mnemonic advantage produced by incidentally encoding aesthetic visual stimuli under aesthetic judgement, self-reference and a control condition. Given that self-referential encoding produces robust mnemonic advantage (Conway, 2005;Cunningham et al., 2008;[START_REF] Kalenzaga | Episodic memory and self-f : Insights from an fMRI study in younger and older adults[END_REF]Leshikar et al., 2015;Sui & Humphreys, 2015;Symons & Johnson, 1997), we expected that comparable results would be observed for aesthetic encoding, owing to the potential shared mechanism. The main result, confirming our hypothesis, was that items in the aesthetic judgment condition were as well recognized as those encoded in selfreference condition when participants gave extreme judgements on the beauty scale during encoding.

First of all, we replicated the self-reference effect (SRE) in memory. Indeed, items encoded in this condition were generally better recognized than the other two conditions (aesthetics and symmetry judgement). This result confirms the effectiveness of our experimental manipulation. Our main result was that items encoded in the aesthetic judgement condition, although they were not generally better recognized, showed the same recognition probability of items encoded in the self-reference condition, when participants had given an extreme judgement (very high or very low) during encoding. Several studies reported a positive link between aesthetic evaluation and memory. For example, [START_REF] Nadal | The Search for Objective Measures of Aesthetic : T f T[END_REF] reported that aesthetic preference was higher for artworks that have left stronger memory traces. Similar results have been observed by [START_REF] Ishai | Perception, memory and aesthetics of indeterminate art[END_REF] reporting that the higher the appreciation of the stimuli during encoding, the more probable their recognition was. In the same vein, Choe et al. (2017) showed that rating aesthetic value during an incidental encoding task boosted memory performances, compared to an intentional encoding condition or to a search task. Finally, a recent study reported better memory performance for the location of paintings that elicited extreme aesthetic experiences, whether positive or negative [START_REF] Babo-Rebelo | Looking, liking and locating : An experimental aesthetics of orienting[END_REF]. Although these results suggest that in general aesthetic judgement enhances memory performance, at first glance, this effect seems to vary between studies. Indeed, memory can be facilitated independently of the extent of the rating during encoding (Choe et al., 2017), can linearly vary with the rating [START_REF] Ishai | Perception, memory and aesthetics of indeterminate art[END_REF], or can be associated with extreme (positive or negative) judgements. This heterogeneity could be linked to the type of aesthetic judgement required. Indeed, Ishai and colleagues (2007) asked participants how strongly the paintings affected them. This measure can capture both positive and negative aspect of the AE. Babo-Rebelo et al. ( 2020) employed liking and intensity rating (the squared liking rating), the latter being a more robust predictor of subsequent memory. In this light, one hypothesis would be that the absolute intensity of AE, more than the valence, determines the subsequent memory enhancement. Our findings seem to be coherent with this hypothesis. Moreover, we showed, for the first time, that the memory enhancement for items receiving extreme aesthetic rating is comparable to that produced by self-reference. Our results directly support the proposal that the memory advantage produced by AE is linked to the recruitment of self-referential processing (Choe et al., 2017).

Interestingly, some neuroimaging studies seem to confirm that intense AE recruits brain regions involved in self-referential processing. For example, Vessel et al. increased activity only for the most moving stimuli. This was particularly true for the medial prefrontal cortex (MPFC). Interestingly, in another study the MPFC showed a nonlinear pattern of activity when participants were asked to judge their appreciation of a human face and body stimuli. Indeed, this region showed increased activation for both ugly and beautiful stimuli, compared to neutral ones (Martín-Loeches et al., 2014). Again, these results suggest that strong AE, whatever their valence, recruit brain regions underpinning self-representation.

Our behavioral data seems to corroborate the suggestion that at least strong AE can activate ' f f (Vessel et al., 2013), involving either an apprehension of the j ' ( f ) (in the case of negative valence).

One alternative explanation for our results could be that the reported effect is not due to aesthetic judgement per se, but would be linked to the emotional reaction associated to highly aesthetically moving stimuli. Indeed, emotional evaluation is central to AE (Chatterjee & Vartanian, 2014), and comes into play at almost every processing level during AE [START_REF] Leder | A model of aesthetic appreciation and aesthetic judgments[END_REF][START_REF] Leder | Ten years of a model of aesthetic appreciation and aesthetic j : T -Developments and challenges in empirical aesthetics[END_REF]. In addition, emotional content is known to facilitate memory (Adelman & Estes, 2013;Leppänen et al., 2007;[START_REF] Meng | The impact of : V[END_REF][START_REF] Schaefer | The effects of emotional intensity on ERP correlates of recognition memory[END_REF]Sharot & Phelps, 2004). Further studies should explore the relation between AE and the self, disentangling the potential effects of emotional content not controlled in the present work.

We also reported that judgement of symmetry showed a linear relationship with subsequent recognition probability. This can be due to the well-known link between symmetry and aesthetic preference as mentioned in the introduction (Gartus & Leder, 2013;Tinio & Leder, 2009). Probably, stimuli judged as highly symmetric were also implicitly judged as beautiful, and produced the same mnemonic advantage. Further studies should use different control conditions not pertaining to judgement of features that are known to be associated with aesthetic evaluation. Concerning source memory, the results were less clear. Indeed, for this measure we did not report any effect of the encoding condition. The blocked presentation of conditions could have facilitated the source memory task leading to a ceiling effect as shown by the high rate of correct answers (see Supplementary Material 2). Thus, although the probability of source memory for items encoded in the aesthetic judgement condition followed the same u-shaped pattern than recognition probability, these data are less easily interpretable.

In conclusion, we presented here behavioral results corroborating previous neuroimaging findings suggesting that intense AEs are strictly linked to self-referential processing. These data support the idea that beauty, but also ugliness, is (at least partly) in the eye of the beholder, and give a cognitive explanation to intersubjective variability in aesthetic appreciation. Beyond the fundamental theoretical interest in the field of neuroaesthetics, our results could have some implications for clinical research. Indeed, some studies reported that ff f A z ' showed a preserved stability of aesthetic preferences, even if they not have explicit memory for the artworks (Halpern et al., 2008;[START_REF] Silveri | T f B " A z ' D ( )[END_REF]. These findings suggest that AE could be a window to preserved portions of the self in these patients.

Author contributions

  significant differences concerning their physical features: luminance (Targets mean = 93.28  40.53; Lure mean = 101.5  40.79; t(98) = -1.00, p = .321); contrast (Targets mean = 50.60  11.11; Lure mean = 48.8  9.90; t(98) = 0.82, p = .415).

Figure A2 .Figure 1 .

 A21 Figure A2.

  (2012) asked participants to rate how strongly paintings move them while recording their brain activity with fMRI. They identified two brain networks showing different pattern of activity. The activity in the first network, composed by sensory regions, increased linearly with

Figure 1 .

 1 Figure 1. The estimated probability of item (A) and source (B) recognition averaged by conditions (1) and its modulation by the rating (2). The error bars represent the 95% Credible Intervals (CI). Thin lines represent individual posterior draws (i.e., the possible effects) and the thick line shows the median effect.

  HL, AJ and MS wrote the article. HL and MS conceptualized the experiment. HL and DM did the data analyses. All the authors contributed to the final draft of the article. PsychologicalBulletin, 106, 265-89. Chatterjee, A., & Vartanian, O. (2014). Neuroaesthetics. Trends in Cognitive Sciences, 18(7), 370-375. https://doi.org/10.1016/j.tics.2014.03.003 Choe, K. W., Kardan, O., Kotabe, H. P., Henderson, J. M., & Berman, M. G. (2017). To Journal of Vision, 17(12), 8. https://doi.org/10.1167/17.12.8 Conway, M. A. (2005). Memory and the self. Journal of Memory and Language, 53(4), 594-628. https://doi.org/10.1016/j.jml.2005.08.005 Cunningham, S. J., Turk, D. J., Macdonald, L. M., & Neil Macrae, C. (2008). Yours or mine? Ownership and memory. Consciousness and Cognition, 17(1), 312-318. https://doi.org/10.1016/j.concog.2007.04.003 Dijkstra, K., & van Dongen, N. N. N. (2017). Moderate Contrast in the Evaluation of Paintings Is Liked More but Remembered Less than High Contrast. Frontiers in Psychology, 8. https://doi.org/10.3389/fpsyg.2017.01507 Gartus, A., & Leder, H. (2013). The Small Step toward Asy : A f Broken Symmetries. I-Perception, 4(5), 361-364. https://doi.org/10.1068/i0588sas Gelman, A., Goodrich, B., Gabry, J., & Vehtari, A. (2019). R-squared for Bayesian Regression Models. The American Statistician, 73(3), 307-309.

	B	. F. (	). Ex		Aff : O	w	-Analysis of Research
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regression modeling via Stan. R package version 2.17. 4. Online< http://mc-stan. org.

The Desire for Aesthetics Scale(Lundy et al.,

2010), the Aesthetic Fluency Scale[START_REF] Silvia | Knowledge-f x : Ex fluency[END_REF][START_REF] Smith | The Nature and Growth of Aesthetic Fluency[END_REF], the Tellegen Absorption Scale(Tellegen & Atkinson, 1974), and the Autism Quotient[START_REF] Auyeung | The Autism Spectrum Q : ' V (AQ-Child)[END_REF].

Acknowledgement

This study was funded by the SublimAE project (ANR-18-CE27-0023), and for MA and JD, also supported by the ANR-17-EURE-0017 FrontCog and the ANR-10-IDEX-0001-02 PSL.

MS would like to thank Dario, Zeno Sperduti for the deep inspiration about beauty.

Leppänen, J. M., Kauppinen, P., Peltola, M. J., & Hietanen, J. K. (2007)