Wang Lei, Gulappa Thippeswamy, Menon K M J
Department of Obstetrics and Gynecology, University of Michigan Medical School, Ann Arbor, MI-48109, USA.
Biochim Biophys Acta. 2010 May;1803(5):591-7. doi: 10.1016/j.bbamcr.2010.02.003. Epub 2010 Feb 16.
Luteinizing hormone receptor (LHR) mRNA binding protein (LRBP), identified as mevalonate kinase, has been shown to be a trans factor mediating the post-transcriptional regulation of LHR mRNA expression in ovaries. LRBP binds to the coding region of LHR mRNA and accelerates its degradation. Our previous studies in an in vitro system showed that LRBP represses the translation of LHR mRNA by forming an untranslatable ribonucleoprotein (mRNP) complex, further suggesting that the untranslatable mRNP complex is directed to the mRNA repression/decay machinery for subsequent mRNA turnover. In the present studies, we used yeast two-hybrid system to screen a cDNA library which was constructed from LHR down-regulated ovaries. Two proteins were identified interacting with LRBP: ribosomal protein S20 (RP S20) and ubiquitin conjugating enzyme 2i (UBCE2i). Their interactions with LRBP were confirmed by the mating assay, co-immunoprecipitation analyses and in vitro sumoylation assays. Furthermore, we show that LRBP is a target for modification by SUMO2/3 but not by SUMO1, at K256 and/or K345. Mutation of both lysine residues is sufficient to abrogate the sumoylation of LRBP. These findings suggest that the direct interaction of LRBP with the translation machinery, through RP S20, may be responsible for the transition of LHR mRNA to an untranslatable complex, and that sumoylation of LRBP may play a role in targeting the untranslatable mRNP complex to the mRNA decay machinery in specific cytoplasmic foci.
黄体生成素受体（LHR）mRNA结合蛋白（LRBP），被鉴定为甲羟戊酸激酶，已被证明是一种反式因子，介导卵巢中LHR mRNA表达的转录后调控。LRBP与LHR mRNA的编码区结合并加速其降解。我们之前在体外系统中的研究表明，LRBP通过形成不可翻译的核糖核蛋白（mRNP）复合物来抑制LHR mRNA的翻译，进一步表明不可翻译的mRNP复合物被导向mRNA抑制/降解机制以进行后续的mRNA周转。在本研究中，我们使用酵母双杂交系统筛选了一个由LHR下调的卵巢构建的cDNA文库。鉴定出两种与LRBP相互作用的蛋白质：核糖体蛋白S20（RP S20）和泛素结合酶2i（UBCE2i）。它们与LRBP的相互作用通过交配试验、免疫共沉淀分析和体外SUMO化试验得到证实。此外，我们表明LRBP是SUMO2/3而非SUMO1在K256和/或K345处修饰的靶点。两个赖氨酸残基的突变足以消除LRBP的SUMO化。这些发现表明，LRBP通过RP S20与翻译机制的直接相互作用可能负责LHR mRNA向不可翻译复合物的转变，并且LRBP的SUMO化可能在将不可翻译的mRNP复合物靶向特定细胞质灶中的mRNA降解机制中发挥作用。
