Wang J, Xu J, Finnerty J, Furuta M, Steiner D F, Verchere C B
Department of Pathology and Laboratory Medicine and the British Columbia Research Institute for Children's and Women's Health, University of British Columbia, Vancouver, Canada.
Diabetes. 2001 Mar;50(3):534-9. doi: 10.2337/diabetes.50.3.534.
Impaired processing of pro-islet amyloid polypeptide (proIAPP), the precursor of the beta-cell peptide islet amyloid polypeptide (IAPP) (amylin), has been implicated in islet amyloid formation in type 2 diabetes. The prohormone convertase enzymes PC3 (also known as PC1) and PC2 are localized to beta-cell secretory granules with proIAPP and proinsulin and are responsible for proinsulin processing. To determine whether PC2 might be essential for proIAPP processing, we performed Western blot analysis of freshly isolated islets from normal mice and mice lacking active PC2. As expected, the primary species of IAPP immunoreactivity in islets from wild-type mice was fully processed (4-kDa) IAPP, with only small amounts of the 8-kDa precursor (unprocessed proIAPP) present. Islets from heterozygous PC2 null mice were identical to wild-type animals, suggesting that half the normal complement of PC2 is sufficient for normal proIAPP processing. By contrast, in islets from homozygous PC2 null mice, the predominant IAPP-immunoreactive form was of intermediate size (approximately 6 kDa), with no detectable mature IAPP and slightly elevated amounts of the 8-kDa precursor form present. Thus, in the absence of PC2, proIAPP processing appears to be blocked at the level of a proIAPP conversion intermediate. Immunofluorescence of pancreas sections and immunoblotting using antisera raised to the NH2- and COOH-terminal flanking regions of mouse proIAPP demonstrated that the 6-kDa intermediate form was an NH2-terminally extended proIAPP conversion intermediate (processed only at the COOH-terminus). These data indicate that PC2 is essential for processing of proIAPP at the NH2-terminal cleavage site in vivo and that PC3 is likely only capable of processing proIAPP at the COOH-terminal cleavage site.
胰岛β细胞肽胰岛淀粉样多肽（IAPP）（胰淀素）的前体——前胰岛淀粉样多肽（proIAPP）加工受损，与2型糖尿病中的胰岛淀粉样变形成有关。激素原转化酶PC3（也称为PC1）和PC2定位于含有proIAPP和胰岛素原的β细胞分泌颗粒中，并负责胰岛素原的加工。为了确定PC2是否对proIAPP加工至关重要，我们对正常小鼠和缺乏活性PC2的小鼠新鲜分离的胰岛进行了蛋白质印迹分析。正如预期的那样，野生型小鼠胰岛中IAPP免疫反应性的主要形式是完全加工的（4 kDa）IAPP，仅存在少量8 kDa前体（未加工的proIAPP）。杂合PC2基因敲除小鼠的胰岛与野生型动物相同，这表明正常PC2量的一半足以进行正常的proIAPP加工。相比之下，在纯合PC2基因敲除小鼠的胰岛中，主要的IAPP免疫反应形式是中等大小（约6 kDa），没有可检测到的成熟IAPP，并且8 kDa前体形式的量略有增加。因此，在没有PC2的情况下，proIAPP加工似乎在proIAPP转化中间体水平受阻。胰腺切片的免疫荧光和使用针对小鼠proIAPP的NH2和COOH末端侧翼区域产生的抗血清进行的免疫印迹表明，6 kDa中间形式是NH2末端延伸的proIAPP转化中间体（仅在COOH末端加工）。这些数据表明，PC2对于体内proIAPP在NH2末端切割位点的加工至关重要，并且PC3可能仅能够在COOH末端切割位点加工proIAPP。
