Autoregulation of actin synthesis responds to monomeric actin levels

Abstract

Regulation of the assembly and expression of actin is of major importance in diverse cellular functions such as motility and adhesion and in defining cellular and tissue architecture. These biological processes are controlled by changing the balance between polymerized (F) and soluble (G) actin. Previous studies have indicated the existence of an autoregulatory pathway that links the state of assembly and expression of actin, resulting in the reduction of actin synthesis after actin filaments are depolymerized. We have employed the marine toxins swinholide A and latrunculin A, both disrupting the organization of the actin-cytoskeleton, to determine whether this autoregulatory response is activated by a decrease in the level of polymerized actin or by an increase in monomeric actin concentrations in the cell. We showed that in cells treated with swinholide A the level of filamentous actin is decreased, and using a reversible cross-linking reagent, we found that actin dimers are formed. Latrunculin A also disassembled actin filaments, but produced monomeric actin, followed by a reduction in actin and vinculin expression, while swinholide A treatment elevated the synthesis of these proteins. In cells treated with both latrunculin A and swinholide A, dimeric actin was formed, and actin and vinculin synthesis were higher than in control cells. These results suggest that the substrate that confers an autoregulated reduction in actin expression is monomeric actin, and when its level is decreased by dimeric actin formation, actin synthesis is increased.

Cascade pathway of filopodia formation downstream of SCAR

“…Levels of total actin as shown by western blotting also decreased but only by 1.6-3-fold (n=3) (blot not shown). Because G-actin has been shown to provide autoinhibitory regulation of actin expression (Lyubimova et al, 1997), the lower total actin levels are probably a secondary effect of SCAR depletion because of net actin depolymerization and transient increase of G-actin levels.…”

Regulation of MKL1 via actin cytoskeleton dynamics drives adipocyte differentiation

“…2b). We next tested whether actin-depolymerizing agents latrunculin A (LatA), which increases monomeric G-actin 27,30 , or swinholide A (SwinA), which increases dimeric actin that does not interact with MKL1 (refs 27,30,31), might mimic the effects of inducers of adipogenesis on dynamics of the actin cytoskeleton and MKL1 localization in DFAT cells expressing mCherry-MKL1. Indeed, LatA or SwinA induced disruption of actin stress fibres and increased the cellular abundance of G-actin monomers or dimers, respectively ( Fig.…”

Geldanamycin Anisimycins Activate Rho and Stimulate Rho- and ROCK-Dependent Actin Stress Fiber Formation

“…Western blotting for actin and tubulin (Sigma) was used at a concentration of 1:1,500 and 1:1,000, respectively. For detergent extraction of F-actin, a subconfluent 100-mm plate of HeLa cells was fractionated as previously described (29). Twenty microliters of each extraction were loaded and blotted for actin content as described above.…”

“…Tubulin levels are similarly unaffected. To determine whether Hsp90 inactivation altered the cellular ratio of F-actin to G-actin, we used a qualitative assay to measure relative F-actin and G-actin levels based on the differential detergent extractability of F-actin and G-actin (29). As shown in Fig.…”