Phosphorylation of Cytohesin-1 by Fyn Is Required for Initiation of Myelination and the Extent of Myelination During Development

Abstract

Schwann cells respond to cues from axons by transforming their cellular morphology and forming myelin. We demonstrated that the guanine nucleotide exchange factor (GEF) cytohesin-1 promoted myelination by activating the small guanosine triphosphatase (GTPase) Arf6. In mice, ablating cytohesin-1 delayed myelination and diminished the amount of myelin produced. We determined that the Src-family kinase Fyn phosphorylated tyrosine 382 (Y(382)) of cytohesin-1, and we generated transgenic mice that expressed a Schwann cell-specific phosphorylation mutant of cytohesin-1 (Y382F) that could not be targeted by Fyn. During development, these transgenic mice displayed delayed myelination compared to that of wild-type mice, as well as a decrease in the amount of myelin produced, similar to that observed in cytohesin-1⁻/⁻ mice. These findings demonstrate that phosphorylation of cytohesin-1 by Fyn is required for full myelination and suggest that tyrosine phosphorylation of GEFs may be a mechanism to activate small GTPases engaged in cell morphogenesis.

Arf6 guanine-nucleotide exchange factor cytohesin-2 regulates myelination in nerves

“…S2). It is thus thought that cytohesin-2 has the ability to produce myelin sheaths, although it is also possible that certain Schwann cell cytohesins act collaboratively to produce them (16).…”

“…These steps are important for protein staining in the interiors of myelin segments [16]. Permeabilized cells were blocked with the Blocking One reagent and washed with PBS containing 0.1% Tween-20, then incubated first with primary antibodies and then with fluorescence-labeled secondary antibodies.…”

“…Unless otherwise indicated, all steps were performed at 4 C. The proteins in centrifuged cell supernatants were mixed for 1.5 h with E. coli-produced recombinant GST-tagged GGA3 proteins (20 mg of GGA3 proteins per 800 mg of total proteins from cell lysates) that had been preabsorbed with glutathion-resin (GE Healthcare), collected by centrifugation, and washed with lysis buffer A containing detergents [16]. GGA3 specifically binds to active, GTP-bound Arf1 or Arf6.…”

“…Dissociated explants were established from male or female C57BL/6 mice on embryonic day 12.5 [16]. In brief, dorsal root ganglia (DRGs) were collected and dissociated using 0.25% trypsin and trituration.…”

P2X7 receptors and Fyn kinase mediate ATP-induced oligodendrocyte progenitor cell migration

“…5′-AACTACAACAACTTCCACGCA-3′ was the nucleotide target sequence for rat Fyn [37]. To confirm the interference effect of Fyn shRNA, OPCs were infected with Fyn shRNA or control shRNA separately.…”