Pannexin–1 Expression in the Lens

Abstract

Abstract: : Purpose: Lens fiber cell metabolism depends on cell–cell communication mediated via gap junctions and cell fusions. Pannexins are a novel family of gap junction–like proteins homologous to the invertebrate innexins that has been characterized recently. We found that gap junction–forming family member pannexin–1 (Panx1) is expressed in lens fiber cells. Here we present the data on protein expression and localization in the mouse lens. Methods: We examined protein expression in neonatal mouse lens using in situ hybridization, real–time PCR, Western blot and immunohistochemistry. We used confocal microscopy and 3–D reconstructions of the lens tissue to visualize the Panx1 cellular distribution in detail. Results:Panx1 transcripts were abundant in equatorial epithelial cells and elongating fiber cells. The real–time PCR supported this data and showed that Panx1 transcripts are much less abundant them Cx50, the major gap junction protein in the lens. Western blot revealed two isoforms of Panx1 in the lens fibers both in soluble and in insoluble fractions of cell lysate. Fiber cells and equatorial epithelium predominantly accumulated the protein, while immunostaining was absent in the central epithelium. Confocal microscopy showed uniform distribution of Panx1–specific staining along fiber cell lateral membranes, with some increased labeling at the posterior and anterior tips. Remarkably, no significant co–localization with gap junction plaques has been observed. According to our data, Panx1 incorporation into plasma membrane of the lens epithelial cells commence at the lens equator, after elongation is initiated. Conclusions:Our data indicate that Panx1 expression in the lens is developmentally regulated and commences as epithelial cells differentiate into fibers. Unlike connexins, this gap junctions–forming protein showed no significant accumulation within regular lens gap–junctions plaques. Relatively high level of Panx1 expression in plasma membranes of fiber cell demonstrates that these cells utilize redundant pathways of cell–cell communication to ensure homeostasis and transparency.

Keywords: gap junctions/coupling • cell-cell communication • protein structure/function