αA-Crystallin and αB-Crystallin Reside in Separate Subcellular Compartments in the Developing Ocular Lens

alpha A-Crystallin (alpha A) and alpha B-crystallin (alpha B), the two prominent members of the small heat shock family of proteins are considered to be two subunits of one multimeric protein, alpha-crystallin, within the ocular lens. Outside of the ocular lens, however, alpha A and alpha B are known to be two independent proteins, with mutually exclusive expression in many tissues. This dichotomous view is buoyed by the high expression of alpha A and alpha B in the lens and their co-fractionation from lens extracts as one multimeric entity, alpha-crystallin. To understand the biological function(s) of each of these two proteins, it is important to investigate the biological basis of this perceived dichotomy; in this report, we address the question whether alpha A and alpha B exist as independent proteins in the ocular lens. Discontinuous sucrose density gradient fractionation and immunoconfocal localization reveal that in early developing rat lens alpha A is a membrane-associated small heat shock protein similar to alpha B but with remarkable differences. Employing an established protocol, we demonstrate that alpha B predominantly sediments with rough endoplasmic reticulum, whereas alpha A fractionates with smooth membranes. These biochemical observations were corroborated with immunogold labeling and transmission electron microscopy. Importantly, in the rat heart also, which does not contain alpha A, alpha B fractionates with rough endoplasmic reticulum, suggesting that alpha A has no influence on the distribution of alpha B. These data demonstrate presence of alpha A and alpha B in two separate subcellular membrane compartments, pointing to their independent existence in the developing ocular lens.