Fell B, Smith A M, Hill R M, Parmar P K, Coates L C, Mezey E, Birch N P
Molecular Neuroendocrinology Laboratory, School of Biological Sciences, The University of Auckland, New Zealand.
Arch Physiol Biochem. 2002 Apr;110(1-2):26-33. doi: 10.1076/apab.110.1.26.909.
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.
丝氨酸蛋白酶抑制剂（丝氨酸蛋白酶抑制剂家族）是一类结构相关的蛋白质，在蛋白水解稳态调节中起关键作用。我们从大鼠垂体中分离出一种新型细胞内丝氨酸蛋白酶抑制剂，称为raPIT5a。Northern印迹分析表明raPIT5a mRNA在一系列组织中表达，包括肾上腺和脑。原位杂交组织化学显示raPIT5a mRNA在大鼠垂体、肾上腺和胰腺的特定细胞群中表达。基于与其他细胞内丝氨酸蛋白酶抑制剂的序列相似性，我们预测raPIT5a可能抑制促凋亡丝氨酸蛋白酶颗粒酶B。我们通过鉴定颗粒酶B和raPIT5a之间稳定的抑制复合物，通过实验证实了这一点。为了确定颗粒酶B或颗粒酶B相关酶是否在大鼠垂体中表达，我们使用预测可扩增颗粒酶B和其他两个已发表的颗粒酶序列的引物进行了PCR。我们鉴定出大鼠自然杀伤蛋白酶-1（RNKP-1），颗粒酶B的大鼠同源物，以及一种与颗粒酶样蛋白III（GLP III）高度相似的新型假定丝氨酸蛋白酶，我们将其称为GLP IIIa。这些数据表明raPIT5a可能通过抑制颗粒酶B或GLP IIIa或具有相似底物特异性的半胱天冬酶酶家族成员来调节垂体中的细胞凋亡。我们还在垂体组织中鉴定出第二种丝氨酸蛋白酶抑制剂神经丝氨酸蛋白酶的表达，并发现它改变了AtT20促肾上腺皮质激素细胞系的形态，可能是通过细胞粘附的变化。这些结果确定了丝氨酸蛋白酶抑制剂在垂体细胞功能中的新作用。
