ADF/cofilin family proteins control formation of oriented actin-filament bundles in the cell body to trigger fibroblast polarization

Abstract

How formation of the front and rear of a cell are coordinated during cell polarization in migrating cells is not well understood. Time-lapse microscopy of live primary chick embryo heart fibroblasts expressing GFP-actin show that, prior to cell polarization, polymerized actin in the cell body reorganizes to form oriented actin-filament bundles spanning the entire cell body. Within an average of 5 minutes of oriented actin bundles forming, localized cell-edge retraction initiates at either the side or at one end of the newly formed bundles and then elaborates around the nearest end of the bundles to form the cell rear, the first visual break in cell symmetry. Localized net protrusion occurs at the opposing end of the bundles to form the cell front and lags formation of the rear of the cell. Consequently, cells acquire full polarity and start to migrate in the direction of the long axis of the bundles, as previously documented for already migrating cells. When ADF/cofilin family protein activity or actin-filament disassembly is specifically blocked during cell polarization, reorganization of polymerized actin to form oriented actin-filament bundles in the cell body fails, and formation of the cell rear and front is inhibited. We conclude that formation of oriented actin-filament bundles in the cell body requires ADF/cofilin family proteins, and is an early event needed to coordinate the spatial location of the cell rear and front during fibroblast polarization.

Cytoskeletal Symmetry Breaking and Chirality: From Reconstituted Systems to Animal Development

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Punctuated actin contractions during convergent extension and their permissive regulation by the non-canonical Wnt-signaling pathway

“…To depolymerize or stabilize F-actin, we treated marginal zone explants with 0.6 M LatB or 5 M jasplakinolide (Jas), respectively, or we overexpressed the Lim kinase mutant LimK T508EE (LimK CA ), which has been reported previously to phosphorylate cofilin and increase the assembly of F-actin (Edwards and Gill, 1999;Mseka et al, 2007). To increase or inhibit myosin-II-induced F-actin contractions, 50 nM calyculin A (CalA), a myosin light chain phosphatase inhibitor, or 50 M Y27632 (a Rho-kinase inhibitor), respectively, were used.…”

Deconstructing the third dimension – how 3D culture microenvironments alter cellular cues

“…One consequence of this is that cells that are cultured on 2D surfaces have a forced apical-basal polarity. This polarity is arguably relevant for some cell types, such as epithelial cells, but is unnatural for most mesenchymal cells, which -when embedded in a 3D ECM -assume a stellate morphology and only polarize from front to rear during migration (Mseka et al, 2007). These changes in cell geometry and organization can directly impact cell function.…”