中华病理学杂志
中華病理學雜誌
중화병이학잡지
Chinese Journal of Pathology
2012年
10期
657-661
,共5页
癌,非小细胞肺%细胞因子信号转导蛋白抑制因子%蛋白酪氨酸激酶类
癌,非小細胞肺%細胞因子信號轉導蛋白抑製因子%蛋白酪氨痠激酶類
암,비소세포폐%세포인자신호전도단백억제인자%단백락안산격매류
Carcinoma,non-small-cell lung%Suppressor of cytokine signaling proteins%Proteintyrosin kinases
目的 探讨SOCS3和Pyk2在非小细胞肺癌(NSCLC)中的表达及其相互作用.方法 分别采用免疫组织化学和免疫荧光染色方法检测SOCS3和Pyk2在100例NSCLC组织、人支气管上皮细胞HBE和6个NSCLC细胞系中的表达.采用甲基化特异性PCR方法检测A549细胞中SOCS3基因甲基化状态.A549细胞经蛋白酶体抑制剂β-lactacystin预处理,再加入去甲基化试剂5-aza-2 ’-脱氧胞苷(5-aza),或转染SOCS3突变质粒,检测Pyk2蛋白、Pyk2 Tyr402和ERK1/2磷酸化水平.采用逆转录(RT)-PCR检测Pyk2 mRNA水平.采用Transwell小室测定细胞迁移情况.结果 100例NSCLC组织中,43例(43.0％) SOCS3阳性表达,65例(65.0％) Pyk2高表达.在NSCLC组织和细胞系中均发现SOCS3和Pyk2表达负相关.5-aza能恢复A549细胞中SOCS3的表达.SOCS3依赖于其SH2和KIR功能区与Pyk2相互作用,从而降低Pyk2蛋白、Pyk2 Tyr402和ERK1/2磷酸化水平,抑制A549细胞迁移.结论 NSCLC中,SOCS3可能通过SOCS-box功能区介导的蛋白酶体途径促使Pyk2蛋白降解,阻断A549细胞迁移.
目的 探討SOCS3和Pyk2在非小細胞肺癌(NSCLC)中的錶達及其相互作用.方法 分彆採用免疫組織化學和免疫熒光染色方法檢測SOCS3和Pyk2在100例NSCLC組織、人支氣管上皮細胞HBE和6箇NSCLC細胞繫中的錶達.採用甲基化特異性PCR方法檢測A549細胞中SOCS3基因甲基化狀態.A549細胞經蛋白酶體抑製劑β-lactacystin預處理,再加入去甲基化試劑5-aza-2 ’-脫氧胞苷(5-aza),或轉染SOCS3突變質粒,檢測Pyk2蛋白、Pyk2 Tyr402和ERK1/2燐痠化水平.採用逆轉錄(RT)-PCR檢測Pyk2 mRNA水平.採用Transwell小室測定細胞遷移情況.結果 100例NSCLC組織中,43例(43.0％) SOCS3暘性錶達,65例(65.0％) Pyk2高錶達.在NSCLC組織和細胞繫中均髮現SOCS3和Pyk2錶達負相關.5-aza能恢複A549細胞中SOCS3的錶達.SOCS3依賴于其SH2和KIR功能區與Pyk2相互作用,從而降低Pyk2蛋白、Pyk2 Tyr402和ERK1/2燐痠化水平,抑製A549細胞遷移.結論 NSCLC中,SOCS3可能通過SOCS-box功能區介導的蛋白酶體途徑促使Pyk2蛋白降解,阻斷A549細胞遷移.
목적 탐토SOCS3화Pyk2재비소세포폐암(NSCLC)중적표체급기상호작용.방법 분별채용면역조직화학화면역형광염색방법검측SOCS3화Pyk2재100례NSCLC조직、인지기관상피세포HBE화6개NSCLC세포계중적표체.채용갑기화특이성PCR방법검측A549세포중SOCS3기인갑기화상태.A549세포경단백매체억제제β-lactacystin예처리,재가입거갑기화시제5-aza-2 ’-탈양포감(5-aza),혹전염SOCS3돌변질립,검측Pyk2단백、Pyk2 Tyr402화ERK1/2린산화수평.채용역전록(RT)-PCR검측Pyk2 mRNA수평.채용Transwell소실측정세포천이정황.결과 100례NSCLC조직중,43례(43.0％) SOCS3양성표체,65례(65.0％) Pyk2고표체.재NSCLC조직화세포계중균발현SOCS3화Pyk2표체부상관.5-aza능회복A549세포중SOCS3적표체.SOCS3의뢰우기SH2화KIR공능구여Pyk2상호작용,종이강저Pyk2단백、Pyk2 Tyr402화ERK1/2린산화수평,억제A549세포천이.결론 NSCLC중,SOCS3가능통과SOCS-box공능구개도적단백매체도경촉사Pyk2단백강해,조단A549세포천이.
Objective To investigate the expression of SOCS3 and Pyk2 and their correlations in non-small cell lung cancer (NSCLC).Methods The expression of SOCS3 and Pyk2 was detected in 100 cases of NSCLC,human bronchial epithelial cells (HBE) and 6 lung cancer cell lines by immunohistochemistry and immunofluorescence staining.The methylation status of SOCS3 was investigated in A549 cells by methylation-specific PCR.A549 cells were either treated with a demethylation agent 5-aza-2'-deoxycytidine (5-aza) or transfected with three SOCS3 mutants with various functional domains deleted.Besides,the cells were pretreated with a proteasome inhibitor β-1actacystin where indicated.The effects of SOCS3 on Pyk2 expression,Pyk2 Tyr 402 and ERK1/2 phnsphorylations were assessed by Western blot.RT-PCR was used to estimate Pyk2 mRNA levels.Transwell experiments were performed to evaluate cell migration.Results SOCS3(43.0％,43/100) and Pyk2 (65.0％,65/100) were expressed in NSCLC.A significant negative correlation was found between SOCS3 and Pyk2 in both NSCLC tissues and cell lines.SOCS3 was aberrantly methylated and 5-aza restored SOCS3 expression.Transfection studies indicated that exogenous SOCS3 interacted with Pyk2,and both Src homology 2 (SH2) and kinase inhibitory region (KIR) domains contributed to Pyk2 binding.Furthermore,SOCS3 was found to inhibit Pyk2-associated ERK1/2 activity in A549 cells.SOCS3 possibly promoted degradation of Pyk2 in a SOCS-box-dependent manner and interfered with cell migration.Conclusions The data indicates that SOCS3 definitely plays roles in regulating Pyk2 signaling,and cell motility.Decreased SOCS3 induced by methylation may confer a migration advantage to A549 cells.
