链脲佐菌素诱导糖尿病大鼠模型脑内肌球蛋白-IIB 的过表达。

Overexpression of myosin-IIB in the brain of a rat model of streptozotocin-induced diabetes.

机构信息

Institute of Genetics and Biochemistry, Federal University of Uberlândia, Campus Umuarama, 38400-902, Uberlândia-MG, Brazil.

出版信息

J Neurol Sci. 2011 Apr 15;303(1-2):43-9. doi: 10.1016/j.jns.2011.01.017.

DOI:10.1016/j.jns.2011.01.017

PMID:21306737

Abstract

The Ca(2+)/calmodulin complex interacts with and regulates various enzymes and target proteins known as calmodulin-binding proteins (CaMBPs). This group of proteins includes molecular motors such as myosins. In this study, we show that non-muscle myosin-IIB is overexpressed in the brains of diabetic rats. We isolated CaMBPs from the brains of non-diabetic rats and rats with streptozotocin-induced diabetes and purified them by immobilized-calmodulin affinity chromatography. The proteins were eluted with EGTA and urea, separated by SDS-PAGE, digested and submitted to peptide mass fingerprinting analysis. Thirteen intense bands were found in both types of brains, two were found exclusively in non-diabetic brains and four were found exclusively in diabetic brains. A large fraction of the eluted proteins contained putative IQ motifs or calmodulin-binding sites. The results of the myosin-IIB affinity chromatography elution, western blot and RT-PCR analyses suggest that myosin-IIB protein and mRNA are expressed at high levels in diabetic brains. This is the first study that has demonstrated differential expression of CaMBPs in diabetic and non-diabetic brain tissue through a comparative proteomic analysis, and it opens up a new approach to studying the relationship between the expression of myosins in the brain, hyperglycemia and intracellular calcium regulation.

摘要

钙调蛋白/钙复合物与各种酶和靶蛋白相互作用并调节这些酶和靶蛋白，这些靶蛋白被称为钙调蛋白结合蛋白（CaMBPs）。这组蛋白包括肌球蛋白等分子马达。在这项研究中，我们显示非肌肉肌球蛋白-IIB 在糖尿病大鼠的大脑中过度表达。我们从非糖尿病大鼠和链脲佐菌素诱导的糖尿病大鼠的大脑中分离出 CaMBPs，并通过固定化钙调蛋白亲和层析对其进行纯化。用 EGTA 和尿素洗脱蛋白，通过 SDS-PAGE 分离，消化后进行肽质量指纹图谱分析。在两种类型的大脑中都发现了 13 个强烈的条带，其中 2 个仅存在于非糖尿病大脑中，4 个仅存在于糖尿病大脑中。洗脱的蛋白中有很大一部分含有推定的 IQ 基序或钙调蛋白结合位点。肌球蛋白-IIB 亲和层析洗脱、western blot 和 RT-PCR 分析的结果表明，肌球蛋白-IIB 蛋白和 mRNA 在糖尿病大脑中表达水平较高。这是第一项通过比较蛋白质组学分析证明糖尿病和非糖尿病脑组织中 CaMBPs 差异表达的研究，为研究大脑中肌球蛋白的表达、高血糖和细胞内钙调节之间的关系开辟了新途径。