Timani Khalid Amine, Liu Ying, Suvannasankha Attaya, He Johnny J
Department of Cell Biology and Immunology, University of North Texas Health Science Center, 3500 Camp Bowie Blvd., Fort Worth, TX 76107, USA.
Department of Cell Biology and Immunology, University of North Texas Health Science Center, 3500 Camp Bowie Blvd., Fort Worth, TX 76107, USA.
Int J Biochem Cell Biol. 2014 Sep;54:10-9. doi: 10.1016/j.biocel.2014.06.017. Epub 2014 Jun 28.
Tip110 is a nuclear protein and has been shown to function in tumor antigenicity, regulation of gene transcription, pre-mRNA splicing, stem cell proliferation and differentiation, and embryonic development. To characterize the in vivo functions of Tip110, a transgene cassette expressing human Tip110 protein (hTip110) was used to generate hTip110 transgenic (Tg) mice. Unexpectedly, only Tip110 mRNA but not Tip110 protein was expressed in Tg MEF and tissues. Treatment of Tg MEF with proteasome inhibitors led to detection of hTip110 protein, which prompted us to investigate the regulatory mechanisms of Tip110 degradation in mouse cells. We found that hTip110 was more sensitive to ubiquitin-proteasome system (UPS)-mediated protein degradation than mouse Tip110 (mTip110), likely resulting from more hTip110 ubiquitination. Using affinity chromatography and proteomics, we identified USP15, a deubiquitinating enzyme, to be associated with Tip110. Tip110 expression led to re-distribution of USP15 from the cytoplasm to the nucleus and complete co-localization of Tip110 with USP15 in the nucleus, whereas USP15 expression resulted in hTip110 deubiquitination. Interestingly, USP15 knockdown restored hTip110 protein expression in Tg MEF and USP15 expression had little effects. Taken together, these results provide insights into the regulatory mechanism of human Tip110 degradation by USP15.
Tip110是一种核蛋白，已被证明在肿瘤抗原性、基因转录调控、前体mRNA剪接、干细胞增殖与分化以及胚胎发育中发挥作用。为了表征Tip110的体内功能，使用表达人Tip110蛋白（hTip110）的转基因盒来生成hTip110转基因（Tg）小鼠。出乎意料的是，在Tg小鼠胚胎成纤维细胞（MEF）和组织中仅表达Tip110 mRNA，而不表达Tip110蛋白。用蛋白酶体抑制剂处理Tg MEF导致检测到hTip110蛋白，这促使我们研究小鼠细胞中Tip110降解的调控机制。我们发现hTip110比小鼠Tip110（mTip110）对泛素-蛋白酶体系统（UPS）介导的蛋白质降解更敏感，这可能是由于更多的hTip110泛素化所致。通过亲和层析和蛋白质组学，我们鉴定出一种去泛素化酶USP15与Tip110相关。Tip110的表达导致USP15从细胞质重新分布到细胞核，并且Tip110与USP15在细胞核中完全共定位，而USP15的表达导致hTip110去泛素化。有趣的是，敲低USP15可恢复Tg MEF中hTip110蛋白的表达，而USP15的表达几乎没有影响。综上所述，这些结果为USP15对人Tip110降解的调控机制提供了见解。
