Váradi A, Molnár E, Ostenson C G, Ashcroft S J
University of Oxford, Nuffield Department of Clinical Biochemistry, John Radcliffe Hospital Headington, UK.
Biochem J. 1996 Oct 15;319 ( Pt 2)(Pt 2):521-7. doi: 10.1042/bj3190521.
Glucose-dependent sequestration of Ca2+ into endoplasmic reticulum and its subsequent release play an important role in the control of intracellular Ca2+ concentration, which regulates insulin secretion in pancreatic beta-cells. The active uptake of cytosolic Ca2+ into endoplasmic reticulum is mediated by sarco-(endo)plasmic reticulum Ca(2+)-ATPases (SERCAs). We found, using RT-PCR with isoform-specific primers, that SERCA 2 and SERCA 3 mRNAs are co-expressed in human and rat islets of Langerhans and in the RINm5F beta-cell line. Immunochemical analysis also revealed the existence of two SERCA proteins with molecular masses of 110 and 115 kDa in beta-cell membranes. The 115 kDa protein was identified as SERCA 2b by its reaction with an isoform-specific antibody and the 110 kDa protein most probably corresponds to SERCA 3. The presence of two functionally different SERCA isoforms raises the possibility that they are located in distinct Ca2+ stores. There is evidence that altered Ca2+ handling in the beta-cell may contribute to the decreased insulin secretion seen in non-insulin dependent diabetes mellitus (NIDDM). We therefore investigated SERCA 2 and SERCA 3 mRNA expression by quantitative RT-PCR in islets prepared from Goto-Kakizaki (GK) rats, a non-obese spontaneous model of NIDDM. We found a significant reduction (about 68%) in SERCA 3 isoform expression. Since SERCA 2 expression was not significantly reduced, these genes are independently regulated and probably play distinct roles in islets of Langerhans. The marked decrease of SERCA 3 expression may constitute a defect in Ca2+ signalling in GK rat islets which could be a component of NIDDM.
葡萄糖依赖的钙离子隔离于内质网及其随后的释放，在控制细胞内钙离子浓度中起重要作用，而细胞内钙离子浓度调节胰腺β细胞中的胰岛素分泌。胞质钙离子向内质网的主动摄取由肌浆（内质）网钙离子 - ATP酶（SERCAs）介导。我们使用同工型特异性引物进行逆转录聚合酶链反应（RT-PCR）发现，SERCAs 2和SERCAs 3 mRNA在人和大鼠胰岛以及RINm5F β细胞系中共同表达。免疫化学分析还揭示了β细胞膜中存在两种分子量分别为110 kDa和115 kDa的SERCA蛋白。通过与同工型特异性抗体反应，115 kDa蛋白被鉴定为SERCA 2b，而110 kDa蛋白很可能对应于SERCA 3。两种功能不同的SERCA同工型的存在增加了它们位于不同钙离子储存部位的可能性。有证据表明，β细胞中钙离子处理的改变可能导致非胰岛素依赖型糖尿病（NIDDM）中胰岛素分泌减少。因此，我们通过定量RT-PCR研究了从NIDDM的非肥胖自发模型Goto-Kakizaki（GK）大鼠制备的胰岛中SERCAs 2和SERCAs 3 mRNA的表达。我们发现SERCAs 3同工型表达显著降低（约68%）。由于SERCAs 2的表达没有显著降低，这些基因是独立调节的，并且可能在胰岛中发挥不同的作用。SERCAs 3表达的显著降低可能构成GK大鼠胰岛中钙离子信号传导的缺陷，这可能是NIDDM的一个组成部分。
