Department of Radiation Oncology, The Ohio State University Medical School, Columbus, Ohio, USA.
Prostate. 2010 Jun 1;70(8):807-16. doi: 10.1002/pros.21114.
p21-activated kinase 6 (PAK6) is a serine/threonine kinase belonging to the p21-activated kinase (PAK) family. We investigated the role of PAK6 in radiation-induced cell death in human prostate cancer cells.
We used a short hairpin RNA (shRNA) strategy to stably knock down PAK6 in PC3 and DU145 cells. Radiation sensitivities were compared in PAK6 stably knockdown cells versus the scrambled shRNA-expressing control cells.
PAK6 mRNA and protein levels in PC3 and DU145 cells were upregulated upon exposure to 6 Gy of radiation. After irradiation, an increased percentage of apoptotic cells and cleaved caspase-3 levels were demonstrated in combination with a decrease in cell viability and a reduction in clonogenic survival in PAK6-knockdown cells. In addition, transfection with PAK6 shRNA blocked cells in a more radiosensitive G2-M phase and increased levels of DNA double-strand breaks. We further explored the potential mechanisms by which PAK6 mediates resistance to radiation-induced apoptosis. Inhibition of PAK6 caused a decrease in Ser(112) phosphorylation of BAD, a proapoptotic member of the Bcl-2 family, which led to enhanced binding of BAD to Bcl-2 and Bcl-X(L) and release of cytochrome c culminating into caspase activation and cell apoptosis.
The combination of PAK6 inhibition and irradiation resulted in significantly decreased survival of prostate cancer cells. The underlying mechanisms by which targeting PAK6 may improve radiation response seem to be multifaceted, and involve alterations in cell cycle distribution and impaired DNA double-strand break repair as well as relieved BAD phosphorylation.
p21 激活激酶 6（PAK6）是丝氨酸/苏氨酸激酶，属于 p21 激活激酶（PAK）家族。我们研究了 PAK6 在人前列腺癌细胞辐射诱导细胞死亡中的作用。
我们使用短发夹 RNA（shRNA）策略在 PC3 和 DU145 细胞中稳定敲低 PAK6。比较 PAK6 稳定敲低细胞与乱序 shRNA 表达对照细胞的辐射敏感性。
PC3 和 DU145 细胞暴露于 6 Gy 辐射后，PAK6 mRNA 和蛋白水平上调。照射后，PAK6 敲低细胞中凋亡细胞的百分比增加，cleaved caspase-3 水平升高，细胞活力降低，克隆形成存活减少。此外，PAK6 shRNA 转染可使细胞阻滞在更敏感的 G2-M 期，并增加 DNA 双链断裂水平。我们进一步探讨了 PAK6 介导辐射诱导细胞凋亡抵抗的潜在机制。PAK6 抑制导致促凋亡 Bcl-2 家族成员 BAD 的 Ser(112)磷酸化减少，导致 BAD 与 Bcl-2 和 Bcl-X(L)结合增强，细胞色素 c 释放，最终导致 caspase 激活和细胞凋亡。
PAK6 抑制联合照射可显著降低前列腺癌细胞的存活率。靶向 PAK6 可能改善辐射反应的潜在机制似乎是多方面的，包括细胞周期分布改变和 DNA 双链断裂修复受损以及 BAD 磷酸化缓解。
