RT Journal Article
SR Electronic
T1 TRIM9-dependent ubiquitination of DCC constrains kinase signaling, exocytosis, and axon branching
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 154666
DO 10.1101/154666
A1 Melissa Plooster
A1 Shalini Menon
A1 Cortney C. Winkle
A1 Fabio L. Urbina
A1 Caroline Monkiewicz
A1 Kristen D. Phend
A1 Richard J. Weinberg
A1 Stephanie L. Gupton
YR 2017
UL http://biorxiv.org/content/early/2017/06/23/154666.abstract
AB Extracellular netrin-1 and its receptor DCC promote axon branching in developing cortical neurons. Netrin-dependent morphogenesis is preceded by multimerization of DCC, activation of FAK and Src family kinases, and increases in exocytic vesicle fusion, yet how these occurrences are linked is unknown. Here we demonstrate that TRIM9-dependent ubiquitination of DCC blocks the interaction with and phosphorylation of FAK. Upon netrin-1 stimulation TRIM9 promotes DCC multimerization, but TRIM9-dependent ubiquitination of DCC is reduced, which promotes an interaction with FAK and subsequent FAK activation. We found that inhibition of FAK activity blocks elevated frequencies of exocytosis in vitro and elevated axon branching in vitro and in vivo. Although FAK inhibition decreased SNARE-mediated exocytosis, assembled SNARE complexes and vesicles adjacent to the plasma membrane were increased, suggesting a novel role for FAK in the progression from assembled SNARE complexes to vesicle fusion in developing murine neurons.DCCDeleted in Colorectal CancerTRIMTripartite MotifSFKsrc family kinaseDCCKRnon ubiquitinatable DCC mutantVAMPvesicle associated membrane proteinTRIM9ΔRINGTRIM9 lacking the ubiquitin ligase RING domainTRIM9ΔSPRYTRIM9 variant lacking the DCC-binding SPRY domainTIRFTotal Internal Reflection FluorescencepYphosphotyrosineFAKipharmacological FAK inhibitor 14FRNKFAK related non-kinaseSTX-1Asyntaxin 1AIPimmunoprecipitate