DIFFERENTIAL DISTRIBUTION OF THE TIGHT-JUNCTION-ASSOCIATED PROTEIN ZO-1 ISOFORM-ALPHA(-ALPHA(-) IN GUINEA-PIG SERTOLI CELLS - A POSSIBLE ASSOCIATION WITH F-ACTIN AND G-ACTIN() AND ISOFORM)

To elucidate the significance of alpha(-) and alpha(+) isoforms of the tight-junction-associated protein ZO-1 in Sertoli cell tight junction regulation, taking into consideration that different isoforms are exp ressed in cells with different junctional morphologies, we investigate d whether alpha(-) and alpha(+) are differentially associated with jun ctions forming the continuous occluding zonules responsible for the bl ood-testis barrier, and/or with junctions forming the focal discontinu ous occluding zonules. In addition, since Sertoli cells contact Sertol i cells and germ cells, we investigated whether each isoform is differ entially associated with distinct classes of germ cells. Our immunoblo t analyses of isolated seminiferous tubules, using affinity-purified p olyclonal antibodies recognizing rat and human alpha(-) and alpha(+), showed that guinea pig testis contained the two ZO-1 isoforms initiall y described in rat and human kidneys, and that alpha(+) and alpha(-) w ere predominantly expressed during puberty and adulthood, respectively , indicating that alpha(+) was predominant during periods of increased junction assembly/disassembly. We used the same antibodies and immuno peroxidase labelling on fetal, neonatal, pubertal, and adult guinea pi g testes sections. Both isoforms were expressed at the site of Sertoli cell-Sertoli cell and Sertoli cell-germ cell junctions in the seminif erous epithelium, before and after birth, and both were localized in c ontinuous and in discontinuous tight junctions. However, the distribut ion of alpha(-) and alpha(+) was not the same in different locations o f the tight junctions. Only alpha(-) was incorporated into junctions j oining the Sertoli cells to all classes of germ cells. The alpha(+) in volved junctions joining Sertoli cells to particular classes of germ c ells, suggesting that Sertoli cell expression of ZO-1 isoforms could b e regulated by unique germ cell-Sertoli cell contacts. Conversely, we found a correspondence between the distribution of F-actin and ZO-1 al pha(+), indicating that the spatial organization of the subsurface act in accompanying cell junctions may affect alpha(+)/alpha(-)-plasma mem brane association.