Shedding of Membrane-associated Mucin MUC16 at Ocular Su

Schepens Eye Research Institute, Boston MA

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Abstract

DESCRIPTION (provided by applicant): The objective of this proposal is to obtain information regarding the physiological role of the membrane associated mucin MUC16 on the ocular surface and the effects that shedding have on normal function. Preliminary studies have shown that MUC16 is expressed at the tips of membrane folds on the apical cell layer (microplicae) on the ocular surface epithelium. Also, soluble forms of MUC16 and several proteases have been detected in samples of human tear fluid. Since a putative protease cleavage site exists on the MUC16 extracellular domain near the cell membrane, it is likely that a mechanism of MUC16 shedding on the ocular surface exists on the ocular surface epithelium. The proposed studies will utilize a truncated, dual-labeled MUC16 construct for expression in cultured cells. Once this protein is expressed on the cell surface, in vitro assays will be developed to quantitate MUC16 ectodomain shedding using both proteases that cleave other membrane-associated mucins and proteases found in the tear film. Finally, the effects of MUC16 ectodomain shedding from the cell surface on several processes will be measured, such as the fate of the remaining intracellular domain, bacterial adherence to the cell surface, and changes in surface architecture. We hypothesize that the ectodomain of MUC16 is shed from the ocular surface epithelium and its shedding increases in response to proteases found in human tear film, which leads to an alteration in normal physiological function.