PURPOSE: This study was undertaken to improve understanding of the defective lens developmental changes induced by the transgenic overexpression of the Rho GDP dissociation inhibitor RhoGDIalpha. The study was focused on a single differentially expressed gene encoding ponsin, a cell adhesion interacting signaling adaptor protein. METHODS: Total RNA extracted from the P7 lenses of Rho GDIalpha transgenic mice was subjected to cDNA microarray analysis. Ponsin distribution in the mouse lenses was determined by immunofluorescence and immunoblot analyses. Interactions among ponsin, actin, and Rho GTPase signaling pathways were explored in lens epithelial cells. RESULTS: The RhoGDIalpha transgenic mouse lenses revealed a marked downregulation of expression of multiple splice variants of ponsin. Expression of one of the ponsins (U58883) was found to be abundant in normal mouse lenses. Although ponsin was localized predominantly to the focal adhesions in lens epithelial cells, it was distributed to both the epithelium and fibers, with some isoforms being enriched primarily in the Triton X-100-insoluble fraction in lens tissue. Further, whereas constitutively active RhoA induced ponsin clustering at the leading edges, inhibition of Rho kinase and latrunculin treatment were noted to lead to decreases in ponsin protein levels in lens epithelial cells. CONCLUSIONS: Abundant expression of ponsin, a focal adhesion protein in the lens tissue indicates a potential role for this protein in lens fiber cell migration and adhesion. Ponsin expression appears to be closely dependent on Rho GTPase-regulated integrity of actin cytoskeletal organization.