嗜熱四膜蟲（T. thermophila）磷酸化修飾位點特征與人類（H. sapiens）及惡性瘧原蟲（P. faciparum）的比較
基于轉(zhuǎn)錄組數(shù)據(jù)（Gene Network）及磷酸化蛋白質(zhì)組數(shù)據(jù)（Kinase Recognition Motif）預(yù)測的嗜熱四膜蟲蛋白質(zhì)磷酸化信號通路范例
11月7日，國際蛋白質(zhì)組學(xué)領(lǐng)域刊物Molecular & Cellular Proteomics在線發(fā)表了題為Phosphoproteomic analysis of protein phosphorylation networks in Tetrahymena thermophila,， a Model Single-Celled Organism的研究論文,，該研究成果由中國科學(xué)院水生生物研究所原生動物功能基因組學(xué)學(xué)科組、水生生物蛋白質(zhì)組學(xué)科組與生物物理研究所蛋白質(zhì)科學(xué)研究平臺合作完成,。
在真核和原核細(xì)胞中,，為了迅速感知并應(yīng)對細(xì)胞內(nèi)外環(huán)境變化，細(xì)胞通常借助可逆的蛋白質(zhì)翻譯后修飾來進(jìn)行信號傳導(dǎo)和對蛋白質(zhì)功能,、亞細(xì)胞定位等的調(diào)節(jié),。作為研究最廣泛的蛋白質(zhì)翻譯后修飾——蛋白質(zhì)磷酸化修飾，其影響了人類細(xì)胞中超過1/3的蛋白質(zhì)的功能,。近來,，隨著磷酸化肽段富集手段和高精度生物質(zhì)譜發(fā)展，蛋白質(zhì)磷酸化修飾位點的鑒定從數(shù)量和精度上得到了長足發(fā)展,；然而,，如何將這些磷酸化蛋白質(zhì)與催化其磷酸化的蛋白質(zhì)激酶一一對應(yīng)成為了近年來該領(lǐng)域的熱點和難點問題。
本研究以具有1069個蛋白質(zhì)激酶（人類僅為518個）的單細(xì)胞真核模式生物——嗜熱四膜蟲為研究對象,，利用TiO2對磷酸化肽段進(jìn)行了富集,，并利2D(SCX/RP)-nanoLC-MS/MS技術(shù)度鑒定得到1008個蛋白質(zhì)的2238個磷酸化修飾位點。通過深入的生物信息學(xué)分析得出：（1）嗜熱四膜蟲磷酸化蛋白質(zhì)參與了細(xì)胞中物質(zhì)運輸,、基因表達(dá)調(diào)控等多個重要生物學(xué)過程,；（2）嗜熱四膜蟲磷酸化位點周圍的氨基酸排布模式（Phosphorylation Motif）與寄生原生生物惡性瘧原蟲及人類存在一些進(jìn)化上保守的模式,，既符合一些蛋白質(zhì)激酶的催化特征，也具有嗜熱四膜蟲所特有的排布模式,；（3）通過結(jié)合基于嗜熱四膜蟲轉(zhuǎn)錄組數(shù)據(jù)構(gòu)建的Gene Network以及基于蛋白質(zhì)磷酸化修飾數(shù)據(jù)集分析得出的Kinase Recognition Motif,，預(yù)測了潛在的“蛋白質(zhì)激酶——磷酸化蛋白質(zhì)”的作用對。
該研究成果首次從蛋白質(zhì)組學(xué)層面對嗜熱四膜蟲的蛋白質(zhì)磷酸化修飾進(jìn)行了分析,，不僅揭示了嗜熱四膜蟲磷酸化修飾位點所具有的進(jìn)化上保守的特征和物種特異性的特征,，還結(jié)合轉(zhuǎn)錄組數(shù)據(jù)對潛在的“蛋白質(zhì)激酶——磷酸化蛋白質(zhì)”作用關(guān)系進(jìn)行了預(yù)測，對后續(xù)嗜熱四膜蟲蛋白質(zhì)磷酸化調(diào)控功能和調(diào)控網(wǎng)絡(luò)的研究奠定了基礎(chǔ),。（生物谷Bioon.com）
生物谷推薦的英文摘要
Molecular & Cellular Proteomics doi: 10.1074/mcp.M112.026575
Phosphoproteomic analysis of protein phosphorylation networks in Tetrahymena thermophila,， a Model Single-Celled Organism
Miao Tian1， Xiulan Chen1,， Qian Xiong1,， Jie Xiong1， Chuanle Xiao2,， Feng Ge1,， Fuquan Yang1 and Wei Miao1，*
Tetrahymena thermophila is a widely used unicellular eukaryotic model organism in biological research and contains more than 1000 protein kinases and phosphatases with specificity for Ser/Thr/Tyr residues. However,， only a few dozen phosphorylation sites in T. thermophila are known,， presenting a major obstacle for further understanding the regulatory roles of reversible phosphorylation in this organism. In this study， we used high accuracy mass spectrometry-based proteomics to conduct global and site-specific phosphoproteome profiling of T. thermophila. In total,， 1384 phosphopeptides and 2238 phosphorylation sites from 1008 T. thermophila proteins were identified through the combined use of peptide prefractionation,， TiO2 enrichment， and 2D-LC-MS/MS analysis. The identified phosphoproteins are implicated in the regulation of various biological processes such as transport,， gene expression,， and mRNA metabolic process. Moreover， integrated analysis of the T. thermophila phosphoproteome and gene network reveals the potential biological functions for many previously unannotated proteins and predicts several putative kinase-substrate pairs. Our data provide the first global survey of phosphorylation in T. thermophila by using a phosphoproteomic approach,， and suggests a wide-ranging regulatory scope of this modification. The provided dataset is a valuable resource for the future understanding of signaling pathways in this important model organism.
