摘要
背景与目的:全长型脾脏酪氨酸激酶[full-length form of spleen tyrosine kinase,Syk(L)]在乳腺癌中具有抑癌功能,可移位到细胞核内,作为转录抑制因子调节基因的转录。本研究拟探讨Syk(L)行使转录抑制功能的具体分子生物学机制。方法:将pFLAG-CMV-Syk(L)转染入人胚肾细胞HEK293中,采用免疫沉淀方法检测组蛋白去乙酰化酶(histone deacetylases,HDACs)与外源性Syk(L)的结合。采用免疫沉淀方法检测乳腺癌细胞MB468中内源性Syk(L)与HDAC1的结合。构建带有FLAG标志的Syk(L)各个功能域质粒,并将其转染入HEK293细胞中,采用免疫沉淀方法检测HDAC1与Syk(L)各个功能域之间的结合。采用HDAC活力检测系统检测Syk(L)免疫沉淀复合物中的HDAC活性。结果:HEK293细胞中外源性Syk(L)可与HDAC1、3、6、7结合,Syk(L)通过SH2功能域和KD功能域与HDAC1相互结合。乳腺癌细胞MB468中内源性Syk(L)与HDAC1可相互结合。Syk(L)免疫沉淀复合物有明显的HDAC活性。结论:在乳腺癌中,抑癌基因Syk(L)可能通过与HDAC形成复合物调节基因的转录。
BACKGROUND & OBJECTIVE: Full-length form of spleen tyrosine kinase [Syk(L)] is a tumor suppressor gene in breast cancer, which may translocate into cell nucleus and act as a transcriptional repressor. This study was to explore the mechanism of Syk(L) regulated gene transcription in breast cancer cells. METHODS: Flag-tagged Syk(L) was transfected into human embryonic kidney (HEK) 293 cells, and the interaction of Syk(L) and histone deacetylases (HDACs) was detected by immunoprecipitation. The binding of endogenous HDAC1 and Syk(L) in breast cancer cell line MB468 was also detected by immunoprecipitation. Flag-tagged Syk(L) domains SH2, KD, and IDB(L) were transfected into HEK293 cells, respectively, and their interaction with HDAC1 was detected. The activity of HDACs in the immunoprecipited complexes of Syk(L) was tested by HDAC activity detecting system. RESULTS: Exogenous Syk(L) was bound with HDAC1, 3, 6, and 7 in HEK293 cells, and endogenous Syk(L) was bound with HDAC1 in MB468 cells. SH2 and KD, but not IDB(L), were coimmunoprecipitated with HDACI. The activity of HDACs was detected in the immunoprecipited complexes of Syk(L). CONCLUSIONS: Syk(L) regulates gene transcription in breast cancer by binding with HDACs to form transcription repressive complexes.
出处
《癌症》
SCIE
CAS
CSCD
北大核心
2007年第5期469-472,共4页
Chinese Journal of Cancer
基金
广东省自然科学基金(No.06300853)
教育部博士点基金(No.20020558050)~~
关键词
乳腺肿瘤
脾脏酪氨酸激酶
组蛋白去乙酰化酶
转录抑制
Breast neoplasm
Spleen tyrosine kinase
Histone deacetylase
Transcriptional repressor