Cole Eric T, Zhan Yu, Abi Saab Widian F, Korchnak Amanda C, Ashburner Brian P, Chadee Deborah N
Department of Biological Sciences, University of Toledo, 2801 West Bancroft Street, Toledo, OH 43606, USA.
Biochim Biophys Acta. 2009 Dec;1793(12):1811-8. doi: 10.1016/j.bbamcr.2009.09.014. Epub 2009 Sep 25.
Mixed lineage kinase 3 (MLK3) is a mitogen activated protein kinase kinase kinase (MAP3K) that activates multiple MAPK signaling pathways. Nuclear factor kappa B (NF-kappaB) is a transcription factor that has important functions in inflammation, immunity and cell survival. We found that silencing mlk3 expression with RNA interference (RNAi) in SKOV3 human ovarian cancer epithelial cells and NIH-3T3 murine fibroblasts led to a reduction in the level of the inhibitor of kappa B alpha (IkappaBalpha) protein. In addition, we observed enhanced basal IkappaB kinase (IKK) activity in HEK293 cells transiently transfected with MLK3 siRNA and in NIH3T3 cells stably expressing MLK3 shRNA (shMLK3). Furthermore, the basal level of NF-kappaB-dependent gene transcription was elevated in shMLK3 cells. Silencing mlk3 expression conferred resistance of cells to etoposide-induced apoptotic cell death and overexpression of wild type MLK3 (MLK3-WT) or kinase-dead MLK3 (MLK3-KD) promoted apoptotic cell death and cleavage of poly (ADP-ribose) polymerase (PARP). Overexpression of MLK3-WT or MLK3-KD enhanced etoposide-induced apoptotic cell death and cleavage of PARP. These data suggest that MLK3 functions to limit IKK activity, and depleting MLK3 helps protect cells from etoposide-induced cell death through activation of IKK-dependent signaling.
混合谱系激酶3（MLK3）是一种丝裂原活化蛋白激酶激酶激酶（MAP3K），可激活多种丝裂原活化蛋白激酶（MAPK）信号通路。核因子κB（NF-κB）是一种转录因子，在炎症、免疫和细胞存活中具有重要功能。我们发现，在SKOV3人卵巢癌上皮细胞和NIH-3T3小鼠成纤维细胞中，用RNA干扰（RNAi）沉默mlk3表达会导致κBα抑制蛋白（IkappaBalpha）水平降低。此外，我们观察到，在瞬时转染MLK3 siRNA的HEK293细胞和稳定表达MLK3短发夹RNA（shMLK3）的NIH3T3细胞中，基础IkappaB激酶（IKK）活性增强。此外，shMLK3细胞中NF-κB依赖性基因转录的基础水平升高。沉默mlk3表达赋予细胞对依托泊苷诱导的凋亡性细胞死亡的抗性，野生型MLK3（MLK3-WT）或激酶失活型MLK3（MLK3-KD）的过表达促进凋亡性细胞死亡和聚（ADP-核糖）聚合酶（PARP）的裂解。MLK3-WT或MLK3-KD的过表达增强了依托泊苷诱导的凋亡性细胞死亡和PARP的裂解。这些数据表明，MLK3的功能是限制IKK活性，耗尽MLK3有助于通过激活IKK依赖性信号传导保护细胞免受依托泊苷诱导的细胞死亡。
