De Leon Gabriel, Sherry Tara C, Krucher Nancy A
Department of Biology and Health Science, Pace University, Pleasantville, New York 10570, USA.
Cancer Biol Ther. 2008 Jun;7(6):833-41. doi: 10.4161/cbt.7.6.5839. Epub 2008 Mar 5.
There is abundant evidence that Retinoblastoma (Rb) activity is important in the control of cell proliferation and apoptosis. Reversible phosphorylation of the Rb protein that is carried out by cyclin dependent kinases and Protein phosphatase 1 (PP1) regulates its functions. A PP1 interacting protein, PNUTS (Phosphatase Nuclear Targeting Subunit) is proposed to be a regulator of Rb phosphorylation. In this study, PNUTS knockdown in MCF7, SKA and HCT116 cancer cells causes a reduction in viability due to increased apoptosis. However, normal cells (MCF10A breast and CCD-18Co colon) do not exhibit reduced viability when PNUTS expression is diminished. PNUTS knockdown has no effect in Rb-null Saos-2 cells. However, when Rb is stably expressed in Saos-2 cells, PNUTS knockdown reduces cell number. Knockdown of PNUTS in p53-/- HCT116 cells indicates that p53 is dispensable for the induction of apoptosis. Loss of PNUTS expression results in increased Rb-phosphatase activity and Rb dephosphorylation. E2F1 dissociates from Rb in cells depleted of PNUTS and the resulting apoptosis is dependent on caspase-8. These results indicate that Rb phosphorylation state can be manipulated by targeting Rb phosphatase activity and suggest that PNUTS may be a potential target for therapeutic pro-apoptotic strategies.
有充分证据表明，视网膜母细胞瘤（Rb）活性在细胞增殖和凋亡的控制中起着重要作用。由细胞周期蛋白依赖性激酶和蛋白磷酸酶1（PP1）进行的Rb蛋白可逆磷酸化调节其功能。一种PP1相互作用蛋白，PNUTS（磷酸酶核靶向亚基）被认为是Rb磷酸化的调节剂。在本研究中，MCF7、SKA和HCT116癌细胞中的PNUTS敲低由于凋亡增加导致活力降低。然而，当PNUTS表达降低时，正常细胞（MCF10A乳腺细胞和CCD - 18Co结肠细胞）并未表现出活力降低。PNUTS敲低对Rb缺失的Saos - 2细胞没有影响。然而，当Rb在Saos - 2细胞中稳定表达时，PNUTS敲低会减少细胞数量。p53基因敲除的HCT116细胞中PNUTS的敲低表明p53对于凋亡的诱导是可有可无的。PNUTS表达的丧失导致Rb磷酸酶活性增加和Rb去磷酸化。在PNUTS缺失的细胞中，E2F1与Rb解离，并且由此产生的凋亡依赖于caspase - 8。这些结果表明，可以通过靶向Rb磷酸酶活性来操纵Rb磷酸化状态，并表明PNUTS可能是治疗促凋亡策略的潜在靶点。
