Davoodi-Semiromi Abdoreza, Laloraya Malini, Kumar G Pradeep, Purohit Sharad, Jha Rajesh Kumar, She Jin-Xiong
Department of Pathology, Immunology, and Laboratory Medicine, College of Medicine, University of Florida, Gainesville, Florida 32610, USA.
J Biol Chem. 2004 Mar 19;279(12):11553-61. doi: 10.1074/jbc.M312110200. Epub 2003 Dec 29.
A number of cytokines that finely regulate immune response have been implicated in the pathogenesis or protection of type 1 diabetes and other autoimmune diseases. It is, therefore, of pivotal importance to examine a family of proteins that serve as signal transducers and activators of transcription (STATs), which regulate the transcription of a variety of cytokines. We report here a defective gene (Stat5b) located on chromosome 11 within a previously mapped T1D susceptibility interval (Idd4) in the nonobese diabetic (NOD) mice. Our sequencing analysis revealed a unique mutation C1462A that results in a leucine to methionine (L327M) in Stat5b of NOD mice. Leu(327), the first residue in the DNA binding domain of STAT proteins, is conserved in all identified mammalian STAT proteins. Homology modeling predicted that the mutant Stat5b has a weaker DNA binding, which was confirmed by DNA-protein binding assays. The inapt transcriptional regulation ability of the mutated Stat5b is proved by decreased levels of RNA of Stat5b-regulated genes (IL-2Rbeta and Pim1). Consequently, IL-2Rbeta and Pim1 proteins were shown by Western blotting to have lower levels in NOD compared with normal B6 mice. These proteins have been implicated in immune regulation, apoptosis, activation-induced cell death, and control of autoimmunity. Therefore, the Stat5b pathway is a key molecular defect in NOD mice.
许多精细调节免疫反应的细胞因子与1型糖尿病及其他自身免疫性疾病的发病机制或保护作用有关。因此，研究作为信号转导子和转录激活子（STATs）的一类蛋白质至关重要，这类蛋白质可调节多种细胞因子的转录。我们在此报告，在非肥胖糖尿病（NOD）小鼠先前定位的1型糖尿病易感性区间（Idd4）内，位于11号染色体上的一个缺陷基因（Stat5b）。我们的测序分析揭示了一个独特的突变C1462A，该突变导致NOD小鼠Stat5b中亮氨酸突变为甲硫氨酸（L327M）。亮氨酸（Leu327）是STAT蛋白DNA结合结构域的首个残基，在所有已鉴定的哺乳动物STAT蛋白中均保守。同源建模预测突变的Stat5b与DNA的结合较弱，这一点通过DNA-蛋白质结合试验得到证实。Stat5b调节基因（IL-2Rβ和Pim1）的RNA水平降低，证明了突变的Stat5b转录调节能力不足。因此，蛋白质印迹法显示，与正常B6小鼠相比，NOD小鼠中IL-2Rβ和Pim1蛋白水平较低。这些蛋白质与免疫调节、细胞凋亡、激活诱导的细胞死亡及自身免疫控制有关。因此，Stat5b通路是NOD小鼠的关键分子缺陷。
