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酪氨酸磷酸酶家族小鼠IA-2和PTP35基因的鉴定及其在神经内分泌组织中的表达。

Identity of mouse IA-2 and PTP35 genes of the tyrosine phosphatase family, and their expression in neuroendocrine tissues.

作者信息

Suk K, Hwang D Y, Kim S, Kim Y H, Kim K A, Seo Y S, Suh Y, Lee S B, Namkung Y, Shin H S, Lee M S

机构信息

Clinical Research Center, Samsung Biomedical Research Institute, Seoul, South Korea.

出版信息

Diabetes Res Clin Pract. 2000 Sep;50(1):17-26. doi: 10.1016/s0168-8227(00)00162-5.

Abstract

Recently, IA-2, one of the major diabetic autoantigens, and PTP35 cDNA were independently isolated by subtraction cloning using insulinoma cells and a polymerase chain reaction (PCR)-based search for conserved sequences using NIH3T3 fibroblast cell line, respectively. By Southern blot analysis and nucleotide sequence determination of reverse transcription PCR products, we showed that IA-2 and PTP35 are identical and exist as a single gene in a mouse genome. The expression of IA-2/PTP35 messages was detected by northern blot analysis in MIN6N8 cells, an insulinoma cell line derived from non-obese diabetic mice, but its expression level was not affected by the ambient glucose level, phorbol-12-myristate 13-acetate or tumour necrosis factor-alpha. We also generated polyclonal antibodies to murine IA-2/PTP35 by immunization with recombinant proteins. Subsequent immunohistochemical analysis using these polyclonal antibodies disclosed that IA-2/PTP35 is strongly expressed in mouse neuroendocrine tissues such as pancreatic islets and the hypothalamus-pituitary gland. These results suggest that IA-2/PTP35 functions primarily in neuroendocrine tissues.

摘要

最近,主要的糖尿病自身抗原之一IA-2和PTP35 cDNA分别通过使用胰岛素瘤细胞的消减克隆以及使用NIH3T3成纤维细胞系基于聚合酶链反应(PCR)搜索保守序列而被独立分离出来。通过Southern印迹分析和逆转录PCR产物的核苷酸序列测定,我们表明IA-2和PTP35是相同的,并且在小鼠基因组中作为单个基因存在。通过Northern印迹分析在MIN6N8细胞(一种源自非肥胖糖尿病小鼠的胰岛素瘤细胞系)中检测到IA-2/PTP35信使的表达,但其表达水平不受环境葡萄糖水平、佛波醇-12-肉豆蔻酸酯13-乙酸酯或肿瘤坏死因子-α的影响。我们还通过用重组蛋白免疫产生了针对小鼠IA-2/PTP35的多克隆抗体。随后使用这些多克隆抗体进行的免疫组织化学分析表明,IA-2/PTP35在小鼠神经内分泌组织如胰岛和下丘脑 - 垂体中强烈表达。这些结果表明IA-2/PTP35主要在神经内分泌组织中发挥作用。

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