αA-and αB-Crystallins Interact With Caspase-3and Bax To Guard Mouse Lens Development

The small heat shock proteins (sHSPs) are members of heat shock protein (HSP) family. In responding to a variety of stress conditions, sHSPs are rapidly increased and take part in the protection against potential injury to the oragn system invovled. The small heat shock protein,α-crystallin, is the lens structural protein, existing in two different isoforms, αA and αB.Lens epithelial cell apoptosis has been revealed as one of common cellular mechanisms mediating non-congential cataract formation. Previous studies have shown that oxidative insult is a common stress in lens epithelial cells and an important initiating factor for cataractogenesis. α-crystallin displays strong ability against induced apoptosis by various stress factors. Regarding the functional mechanism, we and others have demonstrated that α-crystallin can regulate members in both Caspase and Bcl-2families. Our previous studies have also reported that αA-crystallin and αB-crystallin may exert different anti-apoptotic mechanisms under a certain stress condition. While αA-crystallin is able to regulate activation of AKT signaling pathway, α-crystallin negatively regulates the MAPK signaling pathway to suppress apoptosis induced by oxidative and UV irradiation. Although numerous previous studies have demonstrated that αA-and αB-crystallin can regulate the members of Caspase and Bcl-2families, the molecular mechanisms, especially the in vivo functional mechanisms of such regulation remain to be further studied.Our present study amis at determining if αA-crystallin and αB-crystallin bind to Caspase-3and Bax to regulate mouse lens development. Following results were obtained. (1) The surface plasmon resonance assays indicate that both aA-crystallin and aB-crystallin interact with Caspase-3and have different affinities.(2) The Immunohistochemistry assays reveal that all aA-crystallin, aB-crystallin and Caspase-3express in differnent lens developmental stages, and aA-crystallin and aB-crystallin have different colocatizations with Caspase-3.(3) The yeast two-hybrid selection assay demonstrates that aA-crystallin and aB-crystallin can directly bind to Bax with different affinities.(4) Furthermore we use immunohistochemistry to reveal that aA-crystallin and aB-crystallin have colocalization with Bax in different lens developmental stages.(5) At last we confirm the result in co-immunoprecipitation that aA-crystallin and aB-crystallin can form complexes with Caspase-3and Bax in the lens of the newborn mice.Altogether, our results provide first evidence that aA-crystallin and aB-crystallin can directly bind to Caspase-3and Bax to form interacting complexes and guard the mouse lens development both in vitro and in vivo.