Characterisation of Two Serine Protease Inhibitors Expressed in the Pituitary Gland

Abstract

Serine protease inhibitors (serpins) are a family of structurally related proteins that play key roles in the regulation of proteolytic homeostasis. We have isolated a novel intracellular serpin, termed raPIT5a, from the rat pituitary gland. Northern blot analysis indicated raPIT5a mRNA expression in a range of tissues, including the adrenal gland and the brain. In situ hybridisation histochemistry revealed raPIT5a mRNA expression in specific cell populations in the rat pituitary gland, adrenal gland, and pancreas. Based on sequence similarities to other intracellular serpins, we predicted raPIT5a may inhibit the pro-apoptotic serine protease granzyme B. We confirmed this experimentally by identification of a stable inhibitory complex between granzyme B and raPIT5a. To determine whether granzyme B or granzyme B-related enzymes were expressed in the rat pituitary gland, we performed PCR using primers predicted to amplify granzyme B and two other published granzyme sequences. We identified rat natural killer protease-1 (RNKP-1), the rat homologue of granzyme B, and a novel putative serine protease highly similar to granzyme-like protein III (GLP III), which we termed GLP IIIa. These data suggest raPIT5a may regulate apoptosis in the pituitary by inhibition of granzyme B or GLP IIIa, or members of the caspase enzyme family which have similar substrate specificity. We have also identified expression of a second serpin, called neuroserpin, in pituitary tissue and found that it alters the morphology of the AtT20 corticotrope cell line, presumably through changes in cell adhesion. These results identify new roles for serpins in pituitary cell function.

Neuroserpin: a selective inhibitor of tissue-type plasminogen activator in the central nervous system

“…The release of hormones and neurotransmitters from endocrine cells requires proteolytic processing within their secretory vesicles. The reported presence of neuroserpin transcripts in densecored granules of cells in the anterior, intermediate (9) and posterior lobes of the pituitary gland and in the medullary cells of the adrenal gland (25,31,32), suggest that this serpin may also play a role in the regulation of secretory vesicle function (32), either regulating intragranular proteases or inhibiting enzymes following the regulated secretion. Moreover, over-expression of neuroserpin in an anterior pituitary cell line results in the extension of neurite-like processes, suggesting that this serpin may also play a role in the development of intercelullar communications in endocrine tissue (32).…”

“…The serpins (serine proteinase inhibitors) constitute one of the earliest defined protein superfamilies (6). Serpins are involved in the control of processes such as blood coagulation, fibrinolysis and inflammation, (7)(8)(9) as well as in the regulation of neuronal survival both, during development (10) and following cerebral ischemia (11). There are two structural features of a serpin molecule that play an important role in proteinase inhi-bition (12,13).…”

DrosophilaSerpin 4 Functions as a Neuroserpin-Like Inhibitor of Subtilisin-Like Proprotein Convertases

“…Although the serpins were initially characterized as inhibitors of chymotrypsin-like extracellular serine proteases, in principle they can inhibit any protease that forms a covalent intermediate with the substrate. Examples of serpin-mediated regulation of intracellular processing events include the inhibition of the serine and thiol proteases of neuroendocrine cells (Hwang et al, 1999(Hwang et al, ,2002Fell et al, 2002), as well as the inhibition of SPCs (Dahlen et al, 1998;Dufour et al, 1998;Jean et al, 1998;Tsuji et al, 1999).…”

Molecular collaborations between serpins and trefoil factor promote endodermal cell growth and gastrointestinal differentiation in budding tunicates

“…They bind to serine proteases to induce cell proliferation (Miyata et al . 2002) and to regulate apoptosis (Fell et al . 2002).…”