Phosphorylation of M6a at Serine-267 induces filopodia formation in rat hippocampal neurons

GARCIA, M; FORMOSO, K; FRASCH, AC; CAMILA SCORTICATI

Huerta Grande, Cordoba

Congreso; XXIX Annual Meeting and SAN-ISN small Conference and Course; 2014

Sociedad Argentina de Investigación en Neurociencias

The neuronal membrane glycoprotein M6a is a member of the proteolipid protein family. M6a has 278 amino acids that form four transmembrane domains, two external loops, and the N- and C-terminal regions in the cell cytoplasm. M6a induces neurite outgrowth, increases filopodium/spine density and participates in synaptogenesis, but the mechanism of action remains unknown. We previously demonstrated that the lack of both PKC and CK phosphorylation sites of M6a at the C-terminal domain impaired filopodium mobility in neurons. Recently, a phosphorylated form of M6a at S-267 has been found in postnatal and in 21-days old mice brains. In silico analysis of residue 267 showed that it could be a target of PKC. To analyze the importance of the phosphorylation of M6a we overexpressed mutants S267A-M6a (non-phosphorylatable) or S267D-M6a (constitutively phosphorylated) in hippocampal neurons and quantified filopodium formation. The results showed that both M6a wild type and S267D mutant significantly increased the number of processes in 20 um of neurite length compared with control group. However, in S267A expressing neurons the number remained at control levels. By treating M6a expressing neurons with different doses of Gö6976, a PKC inhibitor, we determined that the calcium dependent PKCs are involved in M6a-induced filopodia. In summary, we concluded that phosphorylation of S267 is implicated in M6a filopodia formation pathway and might involve PKC calcium dependent isoforms.