Abstract
Purpose: :
To investigate the molecular and cellular mechanisms for how αA-Y118D mutant proteins with a gain of chaperone-like activity perturb lens transparency and inhibit lens growth.
Methods: :
Mutant lenses were examined by visual observation, histology, immunohistochemistry and confocal microscopy of GFP-positive lens cells. Lens opacity was quantitatively measured by a fiber optic spectrometer. Molecular and biochemical techniques were used to examine the expression, distribution or function of alpha-crystallins and other lens proteins that play roles in lens cell proliferation and differentiation.
Results: :
Mutant αA-Y118D lenses with endogenous aB-crystallin developed mild nuclear cataracts while mutant lenses without endogenous αB-crystallin developed severe whole cataracts with substantial enlarged extracellular spaces between fiber cells. In the epithelial cells of mutant αA-Y118D lenses, αA-Y118D mutant proteins formed intracellular aggregates while endogenous αB-crystallin proteins accumulated near plasma membrane. In the epithelial cells of αA-Y118D mutant lenses without endogenous αB-crystallin, however, both intracellular and plasma membrane-associated αA-Y118D aggregates were observed. Unexpectedly, the growth of αA-Y118D mutant lenses with or without endogenous αB-crystallin was drastically reduced after weaning age when the lens is fully formed. Both delayed macromolecular distribution and fiber elongation were observed in αA-Y118D mutant lenses. BrdU labelling revealed a decrease of epithelial cell mitotic index in mutant lenses.
Conclusions: :
αA-Y118D mutant proteins interact with αB-crystallin to lead to distinct types of cataracts. αB-crystallin plays a crucial role in the regulation of membrane associated αA-Y118D protein aggregation that may perturb plasma membrane channels to cause enlarged extracellular spaces between lens fiber cells. The inhibition of mature lens growth seems to be caused by αA-Y118D mutant protein alone. Thus, αA-Y118D mutation is a useful model for elucidating novel mechanisms that regulate lens growth and for identifying the plasma membrane proteins, recognized by α-crystallins, that maintain lens homeostasis.
Keywords: crystallins • cataract • chaperones