## 3 Discussion
We showed that the affinity constants of LAIR-1 Ig-like domain interaction with C1 qstem_nc2 and the LAIR-1 mutations effects are similar to those obtained with full length C1 qCLR_nc2 (Figure 4B,C and Figure 7 and Table 1). This unambiguously excludes putative LAIR-1 binding sites in the C1q N-terminal collagen bundle even though recently reported results suggested the opposite. Indeed, [PERSON] and colleagues showed that a molecular mimic combining short sequences of HMGB and the N-terminal region of C1q A chain was able to interact simultaneously with RAGE and LAIR-1 and efficiently polarize monocytes to an anti-inflammatory phenotype [36]. We suggest that this C1q A chain sequence, containing one PGXP motif, can be recognized by LAIR-1 in the context of the minimal structure of this mimic but is not accessible in the complex C1q architecture, probably hidden by the other C1q collagen stems forming the bundle. Furthermore, the generation of a short and functional heterotrimeric C1q collagen stem opens the way for various applications. For example, C1 qstem_nc2 could be used for structural studies of C1 qCLR ligands. This approach could be enlarged to other collagen containing proteins such as the soluble defense collagens family. Moreover, a directed mutational analysis of LAIR-1 Ig-like domain based on the literature knowledge on LAIR-1 interaction with collagens, allowed a precise mapping of LAIR-1 residues involved in the interaction with C1 qCLR. Indeed, we identified a C1q binding groove in LAIR-1 where the central residues (R59 and E61) directly interact with the collagen backbone and more distal residues (I102, E72, E111 and P107) are likely involved in the recognition specificity and the correct positioning of the C1 qCLR in front of the LAIR-1 binding region. Our results are in agreement with a previous study identifying LAIR-1 key residues for collagen interaction [22]. The crystal structure of the GPVI in complex with a collagen peptide recently deposited in the PDB (code 5 OU8) reveals that GPVI recognition of the collagen GPO motifs relies on two central arginine and glutamate residues homologous to the R59 and E61 of LAIR-1. Therefore, LAIR-1 and GPVI may share a common interaction mechanism with collagens, based on the direct recognition of the collagen through their central arginine and glutamate residues. LAIR-2 is a soluble collagen receptor that interacts with the same ligands as LAIR-1 and is suggested to act as a competitor to regulate LAIR-1 collagen-induced inhibitory activity. However, the comparison of the affinity values of LAIR-1 and LAIR-2 interaction with C1q still deserves further investigation. Indeed, [PERSON] and colleagues previously reported that LAIR-1 exhibits more affinity for C1q than LAIR-2 while [PERSON] et al. showed the opposite [14; 26]. One contribution for the difference could be that the results of these two studies were obtained using LAIR proteins in different oligomeric states. Indeed, the first team used monomeric Ig-like domains whereas the latter used Fc-fused dimeric Ig-like domains. Since we observed that in the collagen binding groove I102 is substituted by a leucine in LAIR-2, we wanted to test the effect of this substitution on C1q binding in a LAIR-1 context, and generated the LAIR-1 I102L variant. We showed that C1q interaction with the LAIR-1 I102L variant is decreased, a result in line with the study of [PERSON] et al., also obtained using monomeric LAIR-1. It is noteworthy that within the GPVI, the corresponding residue of I102 is a serine, which is quite smaller. This suggests that the specific steric binding constraint for the residue facing I102 in LAIR-1 is likely missing for GPVI binding (Figure 5A), even if LAIR-1 and GPVI partly share a common collagen recognition mechanism. The likely similar mechanism of collagen backbone recognition by LAIR-1 and GPVI allowed us to identify three putative LAIR-1 binding motifs on one C1q collagen stem (underlined in Figure 8A).