Cells Lacking β-Actin are Genetically Reprogrammed and Maintain Conditional Migratory Capacity*

Abstract

Vertebrate nonmuscle cells express two actin isoforms: cytoplasmic ␤-and ␥-actin. Because of the presence and localized translation of ␤-actin at the leading edge, this isoform is generally accepted to specifically generate protrusive forces for cell migration. Recent evidence also implicates ␤-actin in gene regulation. Cell migration without ␤-actin has remained unstudied until recently and it is unclear whether other actin isoforms can compensate for this cytoplasmic function and/or for its nuclear role. Primary mouse embryonic fibroblasts lacking ␤-actin display compensatory expression of other actin isoforms. Consistent with this preservation of polymerization capacity, ␤-actin knockout cells have unchanged lamellipodial protrusion rates despite a severe migration defect. To solve this paradox we applied quantitative proteomics revealing a broad genetic reprogramming of ␤-actin knockout cells. This also explains why reintroducing ␤-actin in knockout cells does not restore the affected cell migration. Pathway analysis suggested increased Rho-ROCK signaling, consistent with observed phenotypic changes. We therefore developed and tested a model explaining the phenotypes in ␤-actin knockout cells based on increased Rho-ROCK signaling and increased TGF␤ production resulting in increased adhesion and contractility in the knockout cells. Inhibiting ROCK or myosin restores migration of ␤-actin knockout cells indicating that other actins compensate for ␤-actin in this process. Consequently, isoactins act redundantly in providing propulsive forces for cell migration, but ␤-actin has a unique nuclear function, regulating expression on transcriptional and post-translational levels, thereby preventing myogenic differentiation.

Assembly of non-contractile dorsal stress fibers requires α-actinin-1 and Rac1 in migrating and spreading cells

“…The observed increase in b-actin at the leading edge of migrating and early spreading cells in the absence of dorsal stress fibers is consistent with this. Interestingly, b-actin deficient fibroblasts display a migration defect apparently associating with increased ventral stress fibers (Bunnell et al, 2011;Tondeleir et al, 2012) suggesting that bactin is involved in dorsal stress fiber assembly. A possibility is also that dorsal stress fiber driven actin polymerization promotes cell migration and spreading by acting as 'mother filaments' for other lamellar actin network structures as suggested for lamellipodial filopodia as a way to increase nucleation points for the Arp2/3 complex (Yang et al, 2007).…”

An actomyosin-like cytoskeleton in the cyanobiont (Nosctoc sp.) of Peltigera canina

“…Also animals express multiple actins and the different isoforms of actin can coexist in the same cell. However, they are differentially regulated and in mice they are not functionally substitutable (Kumar et al, 1997;Chaponnier et al, 1995;Tondeleir et al, 2012). We obtained in a cyanobacterium a positive antigen-antibody response to the anti-b-actin (Sigma A2228) and anti-a-actin (Abcam, ab 40863) antibodies and this revealed the presence of a protein with a molecular mass of 50-55 kDa (Fig.…”

“…Eukaryotic cell motility requires the involvement of actin and associated proteins (Tondeleir et al, 2012;Uehara et al, 2010). Two types of motility occur either polymerization-or contractile-based and both types can occur simultaneously in cells.…”

β-Actin regulates interleukin 6-induced p21 transcription by interacting with the Rpb5 and Rpb7 subunits of RNA polymerase II

“…Loss of actin inhibited Pol II transcription and the recruitment of Pol II into PICs ). There are six highly conserved actin isoforms in vertebrate cells, and β-and γ-actin are the major actin isoforms expressed in non-muscle cells (Tondeleir et al 2012). Whether β-actin directly regulates Pol II transcription remains unclear.…”