摘要
背景与目的:近年来,核基质蛋白(nuclearmatrixproteins,NMPs)成分、结构与功能改变在肿瘤发生发展过程中的作用已成为人们研究的热点问题。分离并鉴定肿瘤相关的NMPs,是寻找肿瘤特异性标志物并进一步研究肿瘤发病机制的一个新途径。本研究通过检测类视网膜母细胞瘤结合蛋白8(similartoretinoblastomabindingprotein8,STRBP8)在人永生化食管上皮细胞癌变过程中的差异表达,为寻找食管癌特异性标志物和研究食管癌的发病机制提供新线索。方法:硫酸铵沉淀法提取人胚永生化食管上皮细胞系SHEE和由SHEE转化而来的食管癌细胞系SHEEC的NMPs,利用双向电泳、基质辅助激光解吸电离飞行时间质谱(MALDI鄄TOF鄄MS)和RT鄄PCR等技术检测STRBP8在SHEE和SHEEC中的差异表达。将STRBP8cDNA序列连接到pGEM鄄Teasy载体后,转化TOP10F蒺大肠杆菌感受态细胞,经筛选获得含STRBP8的阳性克隆,DNA序列测定后进行BLAST比对。结果:通过双向电泳和MALDI鄄TOF鄄MS技术分离并鉴定出核基质蛋白STRBP8只在SHEEC中表达,RT鄄PCR分析发现STRBP8mRNA也只出现在SHEEC细胞中。将其cDNA序列插入pGEM鄄Teasy载体后,经转化和筛选获得含STRBP8的阳性克隆。测序结果显示克隆得到的STRBP8cDNA片段与Genbank数据库上的序列完全一致。结论:STRBP8作为一种新?
BACKGROUND & OBJECTIVE: Recently, changes in composition, structu re, and function of nuclear matrix proteins (NMPs) in generation and development of tumors evoked more and more attention. Separation and identification of tumo r-related NMPs is a new way to search for tumor specific biomarkers, and to stud y tumor pathogenesis. This study was to analyze differential expression of STRBP 8, one of esophageal carcinoma specific NMPs, in cancerization of immortalized h uman esophageal epithelial cells. METHODS: NMPs were extrcted from immortalized human esophageal epithelial cell line SHEE and malignantly transformed esophagea l carcinoma cell line SHEEC. Differential expression of STRBP8 was detected by t wo-dimensional electrophoresis (2-DE), matrix-assisted laser desorption/ionizati on time of flying mass spectrometry (MALDI-TOF-MS), and reverse transcription-po lymerase chain reaction (RT-PCR). STRBP8 cDNA obtained by RT-PCR was linked to p GEM-T easy vector, and introduced into TOP10F' E.coli competent cells. Positive clones were sequenced and analyzed with BLAST. RESULTS: STRBP8 was only detected in SHEEC cells by 2-DE, MALDI-TOF-MS, and RT-PCR. The sequence of positive clon es contained STRBP8 cDNA was identical to that in GenBank database. CONCLUSION: STRBP8, as a candidate oncogene, might relate to cancerization of esophageal epi thelial cells, which might be a specific biomarker of esophageal carcinoma, and probe into the pathogenesis of esophageal carcinoma.
出处
《癌症》
SCIE
CAS
CSCD
北大核心
2005年第4期385-390,共6页
Chinese Journal of Cancer
基金
国家自然科学基金项目(No.30370641)
广东省自然科学基金重点项目(No.37788)~~
关键词
食管肿瘤
核基质蛋白
STRBP8
双向电泳
MALDI-TOF-MS
Esophageal neoplasms
Nuclear matrix proteins
STRBP8
Two-dimensional electrophoresis
Matrix-assisted laser desorption/ionization time of flying mass spectrometry
