Oligodendrocyte lineage cells undergo an apparent step-wise progression from multipotent precursors to a post-mitotic, myelinating phenotype (Baumann and PhamDinh, 2001
). These steps entail changes in cell cycle, motility, morphology and gene expression, implicating the action of a coordinated network of regulatory mechanisms. One of the principal mechanisms engages molecular signals transduced by receptor tyrosine kinases (RTKs). For example, Fgf receptors Fgfr1 and Fgfr2 are required for formation of OPCs in mouse embryonic forebrain (Furusho et al., 2011
) and Fgf receptor signaling is necessary for development of zebrafish hindbrain oligodendrocytes (Esain et al., 2010
). Once OPCs are formed, signaling mediated by the growth factor PDGF and its receptor PDGFRa promotes their proliferation (Calver et al., 1998
). Fgfr signaling in oligodendrocyte formation appears to be conveyed by a Ras/MAPK pathway (Furusho et al., 2011
; Kessaris, 2004
) but the details of the downstream signal transduction mechanisms for RTK function in oligodendrocyte development remain largely unknown.
, which we identified in a screen for genes expressed by oligodendrocyte lineage cells, encodes a protein that potentially mediates RTK signaling. The Swap70 protein has within it a pleckstrin homology domain, which binds PtdIns(3,4,5)P3 (PIP3
), a molecule produced from PIP2
by PI3 kinase following activation by RTK signaling. Studies using cultured fibroblasts showed that mouse Swap70 can function as a PIP3
-dependent guanine nucleotide exchange factor (GEF) for Rac1 GTPase (Shinohara et al., 2002
), which regulates various cell processes such as the cell cycle and motility. In fibroblasts Swap70 promoted membrane ruffling via Rac1 regulation of the actin cytoskeleton (Shinohara et al., 2002
) and Swap70
mutant B cells were less polarized, had unstable lamellipodia and inefficiently migrated into lymph nodes (Pearce et al., 2006
). Therefore, we thought that Swap70 might be important for regulating OPC motility, membrane process dynamics and axon wrapping. Consistent with this possibility, we found that transgenically expressed Swap70 fusion proteins localized to OPC processes during migration and to oligodendrocyte membrane that wrapped axons to form internodes and that this localization required the pleckstrin homology domain. However, neither loss nor gain of Swap70 function affected OPC migration, membrane extension and retraction or axon wrapping. Although we did not find evidence for additional genes with similarity to swap70
in the zebrafish genome, we cannot rule out the possibility that loss of Swap70 function is compensated in our experiments. Nevertheless, our data do not support the hypothesis that Swap70 mediates signal transduction necessary for OPC migration and axon wrapping.
Instead, we found that loss of Swap70 function resulted in a deficit of OPCs. Our data indicate that the deficit results not from reduced OPC proliferation but from decreased production of OPCs. The olig2+ precursors that give rise to OPCs remained intact in swap70 deficient larvae, indicating that the OPC deficit did not result from failure to form or maintain neural precursors. In fact, to our surprise, swap70MO-injected larvae had more neural precursors than normal, as indicated by sox2 expression, and they had elevated numbers of neural cells in the S and M phases of the cell cycle. Therefore, loss of Swap70 function results in maintenance of excessive numbers of dividing neural precursors raising the possibility that Swap70 helps promote the transition from neural precursor to specified OPC.
Loss of Fgfr function in mice and zebrafish (Furusho et al., 2011
; Esain et al., 2010
) and loss of Swap70 function in zebrafish result in a deficit of OPCs, raising the possibility that Swap70 mediates Fgfr signaling necessary for OPC specification. The MAPK pathway also mediates Fgfr signaling and, in culture MAPK function appears necessary for Fgfr-dependent formation of OPCs (Kessaris, 2004
). Immunocytochemistry using a phospho-specific ERK antibody revealed no differences in swap70
morpholino injected larvae (data not shown), indicating that signaling through both MAPK and PIP3
-mediated pathways downstream of Fgf receptors may be required for OPC formation.
One possible role for Swap70 in regulating neural precursor division is through regulation of Rac1, which can promote cell cycle progression via c-Jun kinase (Olson et al., 1995
). However, this is difficult to reconcile with the increase of dividing neural precursors in swap70
deficient larvae because loss of Swap70 GEF activity should result in less Rac1 stimulation of cell cycle progression. Another possibility is that Swap70 plays an unknown role in regulating the cell cycle within the nucleus. Consistent with this possibility, Swap70 contains nuclear localization sequences and we found nuclear localization of Swap70 protein in some cells, particularly in those that were associated with the proliferative ventricular zone of the spinal cord.
A nuclear function for Swap70 has been shown previously but of a very different sort than we imagine for neural precursors. Swap70 was first identified as part of a protein complex in mouse B cells that promotes heavy-chain immunoglobulin class switching by DNA recombination (Borggrefe et al., 1998
). Swap70 localizes to cytoplasm of mast cells and B cells and moves to the nucleus of the latter following B cell activation (Borggrefe et al., 1999
; Gross et al., 2002
; Masat et al., 2000
) and B cells of Swap70
mutant mice are hypersensitive to γ-irradiation and impaired in their ability to switch to the IgE class of immunoglobulins (Borggrefe et al., 2001
). The molecular mechanisms of Swap70 nuclear function remain unknown.
Apart from their altered IgE response, Swap70
mutant mice appear phenotypically normal (Borggrefe et al., 2001
), suggesting that Swap70
function is limited to cells of the hematopoietic lineage. However, other cell and tissue types in mice, including lung and uterus (Borggrefe et al., 1999
), embryonic fibroblasts (Shinohara et al., 2002
) and the nervous system (Lein et al., 2007
) express Swap70. Our data showing that oligodendrocyte lineage cells of zebrafish express swap70
and that Swap70 function promotes transition of neural precursors to specified OPCs but not OPC migration or axon wrapping raises the possibility that Swap70 proteins, in addition to their role in immunoglobulin class switching and B cell motility, regulate transduction of signals that influence neural cell cycle and specification.