Tissue proteases have been implicated in many reproductive and cyclic processes, some of which are accompanied by a dramatic remodeling of reproductive organs and tissues.
The aims of this proposal seek to characterize the hormone-responsive, collagenolytic enzymes and lysosomal responsible for the breakdown of supporting connective tissues and to identify the regulatory hormones. Our preliminary studies have identified, in hog ovaries, three apparently new proteases with collagenolytic potential that are markedly elevated during pregnancy. This proposal includes studies with all three of these protreases because together they provide the enzymatic basis for a proposed hormone-responsive mechanism of collagen resorption. Such a mechanism may account for the observed complete degradation (within lysosomes) of native collagen internalized by the vacuolar (lysosomal) apparatus. The long term objectives of these studies are designed to elucidate the proteolytic mechanisms by which native collagen (comprised of about 25% proline or hydroxyproline) is totally degraded within lysosomes in the absence of any presently known lysosomal proteases capable of cleaving prolyl bonds. The proteases that we have encountered in hog ovaries appear to possess this potential as a consequence of the ability of one to release N-terminal, collagen related Gly-Pro-X """"""""triplets"""""""" and the ability of the others to cleave prolyl bonds (as well as other linkages) located both internally in polypeptides and at N-terminal, penultimate positions (as in the Gly-Pro-X triplets). these proteases will be extracted from the ovaries of pregnant hogs and purified using methods that take advantage of differential solubilities, molecular size, charge, isoelectric points, hydrophobicity, and special affinities. Fluorometric and cytochemical procedures will be used to quantitate and to localize these proteases in tissues such as the ovary, uterus, and pubic symphysis during ovulation, parturition, uterine involution, and in response to administered hormones. Knowledge of the responsible proteolytic mechanisms will not only contribute to our understanding of the underlying mechanisms that mediate reproductive events ranging from ovulation to the breakdown of the interpubic cartilage, but will also contribute generally to our eventual understanding of the etiology of connective tissue diseases such as arthritis and periodontitis.
