Process outgrowth is crucial in oligodendrocyte (OL) development and myelination. It is well accepted that increased levels of proteins affecting the polymerization of cytoskeletal components promote branching. Interestingly, we have suggested that other mechanisms may contribute to oligodendrocyte process outgrowth. We have previously shown that pharmacological inhibitors of histone deacetylation prevent oligodendrocyte branching and we now seek to explore in detail the relationship between these two events. The results presented here indicate that pharmacological inhibitors of histone deacetylation prevent branching, similar to the effect of low doses of cytoskeletal depolymerizing agents. The lack of process outgrowth does not correlate with changes in the levels of tubulin or actin, but correlates with increased levels of microtubule (i.e., stathmin) and microfilaments (i.e., gelsolin) depolymerizing proteins. These data suggest that in OL progenitors, the high levels of depolymerizing proteins maintain a simple morphology, while branching is favored by reduced levels of these cytoskeletal components, consequent to the effect of histone deacetylation on gene expression. We therefore hypothesize that epigenetic regulation of stathmin and gelsolin is a novel regulatory mechanism contributing to OL process outgrowth. In conclusion, our results suggest that process outgrowth in vitro is regulated not only by increased levels of proteins affecting polymerization, but also by decreased levels of proteins affecting depolymerization. The levels of these severing proteins are regulated by chromatin modifiers and therefore suggest that their expression in developing OL is decreased by an epigenetic mechanism. © 2003 Wiley-Liss, Inc.