Denaturation of aquaporin 0 (AQP0): Protection by alpha-crystallin via its chaperone function

Aging is the major risk factor for cataracts. Aquaporin 0 (AQP0) plays a crucial role in maintaining lens transparency by transporting water across the lens fiber cell membranes. Earlier we have shown that alpha-crystallin, a member of the small heat shock protein family, protected AQP0 from thermal-denaturation, in a detergent environment. This study is designed to show whether alpha-crystallin exhibits similar protection towards AQP0 when embedded in the lipid bilayer. We also studied the effect of beta- and gamma-crystallins (1:1 ratio by weight). Lens membranes isolated from the outer cortical fibers of bovine lenses were subjected to thermal-stress from 55°C--90°C for 5 mins. Additionally, we studied the cumulative effect of UV-radiation followed by thermal-stress on AQP0 aggregation and protection by alpha-crystallin. Thermal-stress studies showed that AQP0 aggregated when subjected to 80°C or above, by forming protein aggregates that could not enter the gels with a concomitant loss of 26 kda monomeric protein. Inclusion of neither alpha-crystallin (the molecular chaperone) nor other crystallins (beta-, or gamma-) showed any significant change in thermal aggregation of AQP0 in its native environment. However, when lens membranes were subjected to UV-radiation and then subjected to thermal-stress, aggregation of AQP0 was visible as low as 30°C, which increased progressively with a concomitant loss of AQP0 monomers. Presence of alpha-crystallin significantly decreased AQP0 aggregation, suggesting that UV-radiation disrupted lipid bilayer allowing alpha-crystallin to bind to denaturing AQP0. This study provides a physiological relevant model to study AQP0 aggregation and protective function of alpha-crystallin.