Analysis of neuronal migration defects in src‐1 mutant and Fyn transgenic C. elegans

Abstract

Fyn is a Src Family Kinase (SFK) important for cerebral cortical development. Its SH2 domain tightly regulates catalytic activity via interaction with an inhibitory phosphotyrosine (Y531) in the C‐terminus. Recent mass spectrometry studies identified novel phosphotyrosines (pY185, pY213, pY214) in Fyn's SH2 domain which may regulate Fyn's activity. The goal of this project is to analyze whether newly identified phosphotyrosines in Fyn's SH2 domain alter neuronal migration in vivo. Transgenic worms expressing wild type human Fyn or a Fyn SH2 binding mutant (R176E) in mechanosensory neurons were engineered to determine if Fyn expression rescues the neuronal migration phenotype in an SFK (src‐1) null mutant. A transgenic construct was also engineered containing phosphomimetic mutations at Y185, Y213, and Y214. Fyn expression in transgenic lines was validated by Western blot, and preliminary analysis of neuronal migration in L4 stage C. elegans showed no alterations from non‐transgenic worms. Transgenic lines will be crossed with src‐1 mutants and analyzed for rescue or altered rescue of the src‐1 migration phenotype to determine whether phosphotyrosines within Fyn's SH2 domain are important in the regulation of neuronal development. This project was supported by the Vermont Genetics Network through grant #8P20GM103449 from the INBRE Program of the NIGMS and NCRR, components of the NIH.