Immunocytochemical localization of polyamines during attachment and spreading of retinal pigment epithelial and intestinal epithelial cells

In order to form and maintain a protective barrier for photoreceptors, the retinal pigment epithelium relies on integrin signaling and related pathways to form adhesion complexes, undergo cell spreading, and establish a confluent cellular monolayer. Polyamines are multifunctional polycations that are essential for cell attachment and spreading, although their exact mechanisms of action are as yet unclear. We report new immunocytochemical evidence suggesting that in the cells of retinal pigment epithelium and also the intestinal epithelium, polyamines are present in a population of intracellular vesicles that appear transiently during initial stages of cell spreading. In newly attached cells with minimal spreading, the vesicles are seen near the nucleus, whereas in more highly spread cells, the vesicles are localized to the plasma membrane, near, but not precisely co-localized with an enzyme marker for adhesion complexes, focal adhesion kinase. We also observe pronounced nuclear staining in newly attached cells that have not spread, whereas this staining is decreased in cells that have spread. Nuclear staining has been previously reported in other cell types and has been attributed to DNA binding of polyamines, which is known to stabilize chromatin structure. We hypothesize that the appearance of polyamine vesicles near focal adhesions of cells undergoing attachment and spreading may reflect the mechanism by which polyamine pools are targeted to appropriate interaction sites necessary for the assembly of adhesion complexes. Alternatively, the vesicles could represent the mechanism by which polyamines are removed from the nucleus and possibly released from the cell.