Fig. 4. Actin filament disassembly does not play a major role in the formation of either thin bundles or junctional actin. (A) Recently disassembled filaments do not contribute monomers for incorporation at cell-cell junctions. Cell-cell contacts were induced (Std. Ca²⁺) in the presence of 0.5 µM jasplakinolide (Jasp.) or vehicle control (methanol, MeOH). Cells were stained with anti-E-cadherin and anti-actin antibodies, because jasplakinolide competes with phalloidin for F-actin interaction. (B) Actin disassembly is not required for bundle reorganization. Keratinocytes expressing GFP-actin were treated with 0.02 µM jasplakinolide (Jasp.) or methanol (MeOH) during junction assembly. After 60 minutes, thin bundles were visible either as flanking filaments to junctional actin (middle images Jasp.) or coincident to junctions (bottom images, Jasp.). Arrows indicate thin bundles; arrowheads point to junctional actin. (C) Keratinocytes were classified into four different categories according to the spatial localization of thin bundles as described in Fig. 1 legend. The same proportion of cells was found in category 2 and 3 for both the Jasplakinolide-treated (Jasp.) and untreated (control) cells. Bar, 50 µm.