摘要
目的探讨肿瘤坏死因子α诱导蛋白1(TNFAIP1)介导的RhoA泛素化在黑色素瘤增殖和侵袭中的作用。方法进行生物信息学分析以检测黑色素瘤中TNFAIP1和RhoA的表达,然后测定黑色素瘤患者的临床组织的表达。分别通过RT-qPCR和蛋白质印迹测定评估基因或蛋白质的表达。体外构建TNFAIP1和或RhoA过表达的人皮肤黑色素瘤细胞系WM2664和A2058模型。细胞增殖和侵袭分别通过集落形成、CCK-8和Transwell测定进行研究。将表达TNFAIP1和Ub-K48的质粒共转染到处理过的黑色素瘤细胞中,同时进行免疫沉淀测定以确定TNFAIP1和RhoA之间的相互作用。建立体内模型以确认TNFAIP1和RhoA对肿瘤生长和转移的影响。结果与癌旁正常组织相比,肿瘤组织中TNFAIP1表达显著降低(P<0.01)和RhoA表达显著升高(P<0.01)。TNFAIP1高表达的黑色素瘤患者预后良好,而具有RhoA高表达的黑色素瘤患者预后不良。在体外,TNFAIP1的过表达显著抑制黑色素瘤细胞的增殖和侵袭。TNFAIP1介导K48连接的RhoA泛素化以促进其降解。进一步的救援实验发现,RhoA过表达在体内和体外显著抑制了TNFAIP1过度表达在黑色素瘤增殖和侵袭中的调节作用。结论TNFAIP1在黑色素瘤中发挥肿瘤抑制作用,其通过诱导RhoA泛素化抑制肿瘤细胞的增殖和侵袭。
Objective To investigate the role of tumor necrosis factorα-induced protein 1(TNFAIP1)-mediated RhoA ubiquitination in melanoma cell proliferation and invasion.Methods Bioinformatics analysis was performed to assess TNFAIP1 and RhoA expression in melanoma cell,followed by analyzing its expression in clinical tissues from melanoma patients.Gene and protein expression was assessed by RT-qPCR and Western blot assays,respectively.Human skin melanoma cell lines WM2664 and A2058 overexpressing TNFAIP1 and/or RhoA were established in vitro.Cell proliferation and invasion were investigated by colony formation,CCK-8 and Transwell assays.Plasmids expressing TNFAIP1 and RhoA were cotransfected into treated melanoma cells and immunoprecipitation assays were performed to determine the interaction between TNFAIP1 and RhoA.An in vivo model was established to confirm the effects of TNFAIP1 and RhoA on tumor growth and metastasis.Results Compared with adjacent normal tissues,TNFAIP1 expression was significantly decreased(P<0.01)and RhoA expression was significantly increased(P<0.01)in tumor tissues.Melanoma patients with high TNFAIP1 expression had a good prognosis,whereas melanoma patients with high RhoA expression had a poor prognosis.In vitro,TNFAIP1 overexpression significantly inhibited melanoma cell proliferation and invasion.TNFAIP1 mediated K48-linked ubiquitination of RhoA to promote its degradation.Rescue experiments showed that RhoA overexpression significantly inhibited the regulatory role of TNFAIP1 overexpression in melanin cell proliferation and invasion in vivo and in vitro.Conclusions TNFAIP1 has a tumor suppressor role in melanoma,which inhibits tumor cell proliferation and invasion by inducing RhoA ubiquitination.
作者
肖潇
冯浩
唐桦
李可
李蓝
XIAO Xiao;FENG Hao;TANG Hua;LI Ke;LI Lan(Department of Dermatology,Hunan Provincial People’s Hospital,the First Affiliated Hospital of Hunan Normal University,Changsha 410000,China)
出处
《中国比较医学杂志》
CAS
北大核心
2022年第10期59-68,共10页
Chinese Journal of Comparative Medicine
基金
湖南省中医药科研计划项目(201977)。
