FIG. 2.

Altered cellular morphologies and actin expression levels of Srf^−/− ES cells and their colonies. (A) Light microscopy of individual ES cells. (B) Light microscopy of ES cell colonies. (C) SEM of ES cell colonies. (A to C) Cell and colony morphologies of Srf^−/+ (left panels) or Srf^−/− (right panels) ES cells were determined after initial cultivation for 48 h in complete growth medium, trypsinization, and subsequent plating for either 3 h (A) or 72 h (B and C). Cell protrusions (marked by arrows in panel A) were observed very frequently in the homozygous mutant ES cells but were found rarely in Srf^−/+ cells. (D) Western blot analysis of total cellular actin protein in ES cells. Whole cell extracts from ES cells of the indicated Srf genotypes were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, blotted, and probed with a pan-actin antiserum (upper panel). Coomassie staining of the gel after blotting confirms equal loading (middle panel). Densitometric quantification of the actin signals is shown in the lower panel, where the signal from the E14.1 Srf^+/+ wild-type cells was arbitrarily set at one. (E) Histological distributions of F-actin and E-cadherin in cell colonies derived from ES cells of different Srf genotypes. Wild-type cells (left panel) and the two Srf^−/− cell lines 226-81 (middle panel) and 226-100 (right panel) were grown on coverslips for 48 h. Expression of F-actin (upper row) and E-cadherin (lower row) was detected with Texas red phalloidin and an E-cadherin-specific antiserum, respectively.