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

• Published in: The Journal of biological chemistry 2012-12, Vol.287 (50), p.42407-42416
• Main Authors: Gangalum, Rajendra K., Horwitz, Joseph, Kohan, Sirus A., Bhat, Suraj P.
• Format: Article
• Language: English
• Subjects: alpha-Crystallin A Chain - chemistry; alpha-Crystallin A Chain - metabolism; alpha-Crystallin B Chain - chemistry; alpha-Crystallin B Chain - metabolism; Animals; Cell Biology; Crystallin; Endoplasmic Reticulum (ER); Endoplasmic Reticulum - metabolism; Endoplasmic Reticulum - ultrastructure; Heat Shock Protein; Lens; Lens, Crystalline - embryology; Lens, Crystalline - ultrastructure; Membrane; Rats; Rats, Sprague-Dawley; Rough ER; Smooth ER; Ultracentrifugation; αA-Crystallin; αB-Crystallin
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M112.414854

αA-Crystallin (αA) and αB-crystallin (αB), the two prominent members of the small heat shock family of proteins are considered to be two subunits of one multimeric protein, α-crystallin, within the ocular lens. Outside of the ocular lens, however, αA and αB are known to be two independent proteins, with mutually exclusive expression in many tissues. This dichotomous view is buoyed by the high expression of αA and αB in the lens and their co-fractionation from lens extracts as one multimeric entity, α-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 αA and αB exist as independent proteins in the ocular lens. Discontinuous sucrose density gradient fractionation and immunoconfocal localization reveal that in early developing rat lens αA is a membrane-associated small heat shock protein similar to αB but with remarkable differences. Employing an established protocol, we demonstrate that αB predominantly sediments with rough endoplasmic reticulum, whereas αA 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 αA, αB fractionates with rough endoplasmic reticulum, suggesting that αA has no influence on the distribution of αB. These data demonstrate presence of αA and αB in two separate subcellular membrane compartments, pointing to their independent existence in the developing ocular lens. Background: The small heat shock proteins, αA-crystallin and αB-crystallin are considered to be two subunits of one single monolithic lens protein, α-crystallin. Results: αA-Crystallin and αB-crystallin fractionate independent of each other and in two separate membrane compartments. Conclusion: αA-Crystallin and αB-crystallin are two independent proteins in the lens. Significance: These data provide functional insight into why αA-crystallin and αB-crystallin null mice have disparate phenotypes.