PBK/TOPK在乳腺癌细胞中介导MEK非依赖性ERK激活中作用机制分析

Mechanism of PBK/TOPK in Mediating MEK-Independent ERK Activation in Breast Cancer Cells

该文探讨了乳腺癌细胞中表皮生长因子(EGF)介导的MEK非依赖性ERK激活通路。Western blot检测EGF刺激下,siRNA抑制MEK1/2后的T47D细胞的p-ERK水平,以验证T47D细胞中存在EGF介导的MEK非依赖性ERK激活的通路。接着使用可能参与MEK非依赖性ERK激活的激酶的小分子抑制剂抑制相关激酶(AC、PKC、Src、PI3K、PDK1和Akt)活性后,检测T47D细胞EGF介导ERK的磷酸化水平。siRNA抑制MEK1/2表达后,T47D细胞在EGF刺激后的仍保留部分p-ERK,即在T47D细胞中,存在EGF介导的MEK非依赖性的ERK磷酸化通路。小分子抑制剂抑制AC、PKC、Src对MEK非依赖性ERK激活途径影响不大。而使用小分子抑制剂抑制PI3K、PDK1和Akt后,ERK的磷酸化水平显著降低,提示PI3K/Akt通路下游的激酶参与T47D中EGF介导的MEK非依赖性ERK激活途径。siRNA干扰PI3K/Akt通路下游PBK/TOPK后并使用U0126抑制MEK功能后,几乎检测不到p-ERK,提示PBK/TOPK参与T47D细胞中EGF介导的MEK非依赖性ERK激活途径。乳腺癌抗雌激素药物耐药株T47D细胞存在EGF介导的MEK非依赖性ERK激活途径,且该途径受PI3K/Akt下游的PBK/TOPK调控。

The aim of this article was to investigate the EGF(epidermal growth factor)-mediated MEKindependent ERK activation pathway in breast cancer cells.Under the stimulation of EGF detected by Western blot,siRNA inhibited the p-ERK level of T47D cells after MEK1/2 treatment.This article was to confirm the presence of EGF-mediated MEK-independent ERK activation in T47D cells.The phosphorylation level of ERK mediated by EGF in T47D cells was then measured after inhibition of the activity of related kinases(AC,PKC,SRC,PI3K,PDK1 and Akt)by small molecular inhibitors of kinases that may be involved in MEK independent ERK activation.After MEK1/2 inhibiton,T47D cells retained partial p-ERK under the stimulation of EGF,EGF-mediated MEKindependent ERK phosphorylation pathway in T47D cells.The AC,and PKC and Src inhibiton by small molecule inhibitors have little effect on MEK-independent ERK activation pathway.The phosphorylation level of ERK was significantly decreased after inhibition of PI3K,PDK1 and Akt by small molecule inhibitors,suggesting that the kinase downstream of the PI3K/Akt pathway was involved in the EGF-mediated MEK-independent ERK activation pathway in T47D cells.After PBK/TOPK downstream of PI3K/Akt pathway interfered by siRNA and MEK1/2 function inhibited treated by U0126,p-ERK was almost undetectable,suggesting that PBK/TOPK was involved in EGF-mediated MEK-independent ERK activation pathway in T47D cells.EGF-mediated MEK-independent ERK activation pathway is present in breast cancer anti-estrogen drug-resistant strain T47D cells,and this pathway is regulated by PBK/TOPK downstream of PI3K/Akt.