Background: Genome-wide association studies (GWAS) have identified hundreds of autoimmune diseases-associated loci so far but much of the heritability of these diseases remains unknown. In an attempt to identify potential causal variants, various studies revealed that the missense variant rs35677470 at DNASE1L3 is associated with the development of systemic lupus erythematosus (SLE), rheumatoid arthritis (RA) and systemic sclerosis (SSc), thus exhibiting a pleiotropic effect. Deoxyribonuclease I-like 3 (DNase1L3) is a member of human DNase I family, representing a nuclease that cleaves double-stranded DNA during apoptosis and is involved in the development of autoimmune diseases [1]. Objectives: To investigate the role of the rs35677470 polymorphism at DNASE1L3 gene leading to the R206C mutation in SLE, RA and SSc [2-3] and the mechanism that may affect the loss of function in the protein structure. Methods: The DNASE1L3 evolution was investigated to define conservation elements in the protein sequence using, BLASTP extended searches [4], TCOFFEE [5] multiple sequence alignments, and MEGAX [6] for phylogenetics analysis. Three-dimensional (3D) homology modeling was used to localize the polymorphism under study. The mutant was constructed by molecular modeling using the structures of homologous DNAses (PDB entries 1atn, 4awn, 3d3w; [7-9]). Molecular mechanics/dynamics studies were applied to validate structural/functional changes caused by the R206C substitution. All figures depicting 3D models were generated using the PyMOL molecular-graphics system V.2.2 (Schrodinger, LLC). Results: The evolutionary analysis shows heavily conserved sequence elements among species indicating structural/functional importance. Structural analysis revealed that the rs35677470 SNP codes for a nonconservative amino acid variation, R206C, disrupts the conserved electrostatic network holding protein secondary structure elements to place. Specifically, the R206 to E170 interaction, part of a salt bridge network stabilizing two a-helices, is being interrupted, thereby affecting the molecular architecture (Fig. 1). Indeed, previous studies on the effect of this SNP in Caucasian populations resulting in a lower level of DNAse1L3 activity are consistent with this observation [10]. Conclusion: This study represents a comprehensive evaluation of the shared autoimmune loci of DNASE1L3 (rs35677470), reported to produce an inactive form of DNaseIL3 [10]. The structural analysis, explains the potential role of the produced mutation by modifying the placement of structural elements and consequently introducing disorder in the protein folding and affecting biological function. Altogether, this study contributes to the delineation of the genetic architecture of SLE, RA and SSc. References: [1]Sisirak V et al (2016). Cell 166:88–101 [2]Westra HJ et al (2018). Nat Genet. 50:1366-74 [3]Acosta-Herrera M et al. (2018). Ann Rheum Dis. 78:311-19 [4]Altschul SF et al (1997) Nucleic Acids Res. 25:3389-402. [5]Notredame et al (2000) JMB, 302:205-17 [6]Kumar S et al (2018) Mol. Biol. Evol. 35:1547-9 [7]Kabsch W et al (1990) Nature 347: 37-44 [8]Parciegla et al (2012) Biochemistry 51: 10250 [9]Sasaki K et al (1993) Acta Cryst., A 49: 111-2 [10]Ueki et al (2009) Clinica Chim. Acta 407:20–4 Disclosure of Interests: None declared