Increased expression and phosphorylation of liver glutamine synthetase in well-differentiated hepatocellular carcinoma tissues from patients infected with hepatitis C virus
Increased expression and phosphorylation of liver glutamine synthetase in well-differentiated hepatocellular carcinoma tissues from patients infected with hepatitis C virus
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DOI：
10.1002/elps.200500718
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发表时间：
2006-04-01
期刊：
影响因子：
2.9
通讯作者：
Nakamura, Kazuyuki
中科院分区：
文献类型：
--
作者：
Kuramitsu, Yasuhiro;Harada, Toshio;Nakamura, Kazuyuki
肝细胞癌（HCC）是最常见的致命性癌症之一，在日本，丙型肝炎病毒（HCV）的慢性感染被认为是主要原因之一。为了确定诊断或治疗HCV相关的HCC（HCV-HCC）的生物标志物，我们试图阐明HCV感染患者癌组织产物的相关因素。从来自HCV-HCC癌组织的肝组织样品和来自患者的相应非癌组织的蛋白质组差异显示分析中，检测到相同分子量（42 kDa）的三个蛋白质点，其在分化良好的癌组织中表达增加。虽然它们的pI不同，但通过PMF和MALDI-TOF MS以及使用抗谷氨酰胺合成酶（GS）特异性mAb的Western blotting，它们被鉴定为GS。免疫组化分析显示肿瘤组织由GS阳性细胞区和GS阴性细胞区两部分组成，提示GS产生细胞在肿瘤组织中呈结节状生长。最酸性的GS同种型的胰蛋白酶肽失去了899.5 Da的信号，对应于SASIRIPR的肽，并获得了1059.5 Da的信号，将其提交给PSD分析。PSD分析表明，通过消除两个磷酸基团的中性损失，应该是在丝氨酸残基的899.5-Da肽，从丝氨酸320精氨酸327 GS。PMF后PSD分析被认为是有用的蛋白质显示分子异质性的磷酸化位点的测定。
Hepatocellular carcinoma (HCC) is one of the most common fatal cancers, and chronic infection with hepatitis C virus (HCV) is thought to be one of the main causes in Japan. To identify diagnostic or therapeutic biomarkers for HCC associated with HCV (HCV-HCC), we tried to elucidate the factors related to the products from cancerous tissues of HCV-infected patients. From proteomic differential display analysis of liver tissue samples from HCV-HCC cancerous tissues and corresponding non-cancerous tissues from patients, three protein spots of the same molecular mass (42 kDa), whose expression increased in well-differentiated cancerous tissues, were detected. Although their pl were different, they were identified as glutamine synthetase (GS) by PMF with MALDI-TOF MS and by Western blotting using anti-GS specific mAb. Immunohistochemical analysis showed that tumor tissue consists of two parts, GS-positive cell and GS-negative cell regions, suggesting that GS-producing cells grew in the tumor tissue as a nodule in nodules. The tryptic peptides of the most acidic GS isoform lost the signal of 899.5 Da, corresponding a peptide of SASIRIPR, and gained a signal of 1059.5 Da, which was submitted to PSD analysis. PSD analysis showed the neutral loss by elimination of two phosphate groups, supposed to be on serine residues of the 899.5-Da peptide, from serine 320 to arginine 327 in GS. PMF followed by PSD analysis is thought to be useful for the determination of phosphorylation sites of proteins showing molecular heterogeneity.