The objective of this proposal is to study the functional role of renal (tissue) kallikrein-binding protein in hypertension. Tissue kallikrein are a group of homologous serine proteinases, encoded by a multigene family and involved in the processing of bioactive peptides and hormones. Kallikrein and its products play important roles in cellular homeostasis and blood pressure regulation. Although the biochemistry and regulation of kallikrein gene expression in normal and disease states have been extensively studied, information regarding factors which modulate kallikrein function is very limited. We have recently identified a novel kallikreinbinding protein in rat and human which forms and irreversible complex with tissue kallikrein. The binding protein in rat and human which form an irreversible complex with tissue kallikrein. The binding protein is defective in spontaneously hypertensive rats (SHR), suggesting that it may be a contributing factor in the hypertensive phenotype of SHR.
The specific aims of this study are: 1) to purify and characterize kallikrein-binding protein; develop monoclonal and polyclonal antibodies for determining distribution, quantities and localization of kallikrein- binding protein by binding assays, radioimmunoassays, Western blotting, immunohistochemistry and for physiological studies; 2) to elucidate the role of kallikrein-binding protein in the regulation of kallikrein bioavailability by in vivo clearance and catabolism studies; 3) to isolate cDna clones encoding the kallikrein-binding protein from a rat liver cDNA library and to use these clones to identify and localize kallikreinbinding protein gene expression by Northern, dot blot and in situ cDNA-mRNA hybridization; 4) to isolate and characterize the gene encoding the kallikreinbinding protein from a rat genomic library and to identify the potential cis- and trans-acting elements involved in tissue-specific expression of the gene by deletion analysis and nuclear protein binding assays; 5) to analyze restriction fragment length polymorphisms associated with the kallikrein-binding protein gene in genetically hypertensive animal models. The information gained form these studies should provide a better understanding of the kallikrein-kinin system's role in hypertension.
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