Patrinostro, XiaobaiO’Rourke, Allison R.Chamberlain, Christopher M.Moriarity, Branden S.Perrin, Benjamin J.Ervasti, James M.2017-08-242017-08-242017-03-15Patrinostro, X., O’Rourke, A. R., Chamberlain, C. M., Moriarity, B. S., Perrin, B. J., & Ervasti, J. M. (2017). Relative importance of βcyto- and γcyto-actin in primary mouse embryonic fibroblasts. Molecular Biology of the Cell, 28(6), 771–782. http://doi.org/10.1091/mbc.E16-07-0503https://hdl.handle.net/1805/13923The highly homologous β (βcyto) and γ (γcyto) cytoplasmic actins are hypothesized to carry out both redundant and unique essential functions, but studies using targeted gene knockout and siRNA-mediated transcript knockdown to examine βcyto- and γcyto-isoform--specific functions in various cell types have yielded conflicting data. Here we quantitatively characterized actin transcript and protein levels, as well as cellular phenotypes, in both gene- and transcript-targeted primary mouse embryonic fibroblasts. We found that the smooth muscle αsm-actin isoform was the dominantly expressed actin isoform in WT primary fibroblasts and was also the most dramatically up-regulated in primary βcyto- or β/γcyto-actin double-knockout fibroblasts. Gene targeting of βcyto-actin, but not γcyto-actin, led to greatly decreased cell proliferation, decreased levels of cellular ATP, and increased serum response factor signaling in primary fibroblasts, whereas immortalization induced by SV40 large T antigen supported fibroblast proliferation in the absence of βcyto-actin. Consistent with in vivo gene-targeting studies in mice, both gene- and transcript-targeting approaches demonstrate that the loss of βcyto-actin protein is more disruptive to primary fibroblast function than is the loss of γcyto-actin.en-USAttribution-NonCommercial-NoDerivs 3.0 United Statesβ (βcyto)γ (γcyto)Cytoplasmic actinsActin transcriptCellular phenotypesSmooth muscle αsm-actin isoformGene targetingCell proliferationβcyto-actin proteinsArticle