Abstract
Abstract: :
Purpose: Investigate the regulation of the water permeabilty of aquaporin 0 Methods: Site directed mutatgenesis, expression in Xenopus oocytes, measurement of osmotic swelling. Results: Aquaporin 0 water permeability is regulated by pH and calcium. External histidines modulate the pH regulation. Conclusions: Regulation of water permeality by ionic composition is a plausible physiological role for aquaporin 0 in the lens. Naturally occurring mutations and knockouts clearly indicate that AQP0 is essential for normal lens development and function. But it is not certain what the functions AQP0 provides the lens are. One possibility is that it facilitates intrinsic circulation of fluid in the lens that maintains lens transparency and homeostasis in the absence of blood vessels. pH and Ca2+ are likely candidates for effecting regulatory control of this circulation, as both of these ions seem to play important roles in the lens. The lens interior is more acidic (pH 6.5) than the surface (pH 7.02), and disturbances in Ca2+ concentration are associated with cataract. Thus Ca2+ and pH are both candidates for ionic regulation of the water permeability of AQP0. We find that two separate sites mediate low pH and low Ca2+ sensitivity of bovine AQP0 expressed in Xenopus oocytes. Low Ca2+ also increases the water permeability of MIPfun, the homolog of AQP0 from Fundulus heteroclitus lens, but alkaline rather than acid pH increases the water permeability of MIPfun. When AQP0 and B1234Q, a Drosophila calmodulin mutant that cannot bind calcium, are co-injected into Xenopus oocytes, the Ca2+ sensitivity is lost but pH sensitivity is not altered. Coinjection of B1234Q cRNA also suppresses the calcium sensitivity of both MIPfun and AQP0/H40C, a pH-insensitive histidine mutation, without changing their pH sensitivities. Additional substitution and translocation mutants probe the effects of external loop histidines on the pH curve.
Keywords: cataract • mutations • calcium