Posttranscriptional control of gene expression

我们的研究集中于基因表达的调控，特别是控制细胞和病毒mRNA表达的机制。由HIV-1研究开创的逆转录病毒模型系统的使用，导致了信使核糖核酸代谢领域的重大发现。这些研究的结果是阐明了mRNAs和蛋白质都利用的来自细胞核的主要运输途径。我们的工作导致发现了两个关键逆转录病毒系统的转录后调控机制：人类免疫缺陷病毒1型(HIV-1)和猿猴D型逆转录病毒(SRV)。HIV-1的转录后表达是通过与顺式作用的Rev反应元件(RRE)相互作用的基本病毒Rev蛋白介导的，而SRV的表达是通过与顺式作用的构成运输元件(CTE)相互作用的细胞因子介导的。促进病毒mRNAs表达的机制还涉及细胞因素，因此，对它们的鉴定和表征对于我们理解细胞基因表达具有重要意义。我的实验室使用生物化学和功能基因组学和蛋白质组学工具相结合的方法，研究了介导HIV-1和SRV细胞mRNA运输和表达的分子步骤。对大分子转录后调控和核质运输机制的剖析对于理解癌症发生的过程是相关的。 在以前的研究中，我们已经鉴定了几种D型逆转录病毒和一些脑池内A粒子(IAP)逆转录病毒的CTE RNA转运元件。这些研究证实，两个内部环路的整体结构和顺序对功能至关重要。通过体外RNA选择，我们最近发现了一个具有原始CTE性质的基因组元件，它能够介导非洲爪哇卵母细胞核的RNA输出(Zolotukhin，A.S.，et al.J·维罗尔。75：5567-5575,2001)。这些发现构成了这些元素与CTE之间进化联系的直接证据。 我们已经确定Nxf1(以前命名为TAP)是CTE RNA的重要输出受体，也是细胞mRNA的关键转运体。Nxf1是第一个被发现的细胞mRNAs输出受体，因此在mRNA和核孔复合体之间提供了直接的联系。我们进一步发现，线虫中的Nxf1同源基因是一种重要的mRNA输出因子，表明Nxf1功能在后生动物中是保守的(Tan，W.等人。核糖核酸。6：1762-1772,2000)。来自我的实验室的最新数据显示，Nxf1直接与剪接因子U2AF结合(Zolotukhin等人，提交)。这种相互作用在剪接和信使核糖核酸的输出之间提供了新的联系。在与George Pavlakis的合作中，我们发现了一种新的有效的RNA运输元件RTE(Nappi，F.，et al.J·维罗尔。75：45584569,2001年)。我们发现RTE功能在脊椎动物中是保守的。RTE的使用为我们进一步剖析mRNA的转运机制提供了有力的工具。

Our research focuses on regulation of gene expression, in particular the mechanisms controlling cellular and viral mRNA expression. The use of retroviral model systems, pioneered by research on HIV-1, have led to major discoveries in the field of mRNA metabolism. These studies have resulted in the elucidation of the major transport pathways from the nucleus utilized by both mRNAs and proteins. Our work led to the discovery of mechanisms governing posttranscriptional regulation of two key retroviral systems: human immunodeficiency virus type 1 (HIV-1) and simian type D retrovirus (SRV). Whereas the posttranscriptional expression of HIV-1 is mediated through the essential viral Rev protein interacting with the cis-acting Rev-responsive element (RRE), the expression of SRV is mediated through a cellular factor interacting with the cis-acting constitutive transport element (CTE). The mechanisms promoting expression of viral mRNAs also involve cellular factors; therefore, their identification and characterization is important for our understanding of cellular gene expression. My lab investigates the molecular steps mediating cellular mRNA transport and expression of HIV-1 and SRV using a combination of biochemistry and tools of functional genomics and proteomics. The dissection of the mechanisms of posttranscriptional control and nucleocytoplasmic trafficking of macromolecules are relevant to understand the processes involved in carcinogenesis. In previous studies, we have characterized the CTE RNA transportelements of the several simian type D retroviruses and some intracisternal A particle (IAP) retroelements. These studies identified that the overall structure and the sequence of the two internal loops are essential for function. By in vitro RNA selection, we have recently identified a genomic element with the properties of a primordial CTE, which is able to mediate RNA export from Xenopus oocyte nucleus (Zolotukhin, A. S., et al. J Virol. 75:5567-5575, 2001). These findings constitute direct evidence of an evolutionary link between these elements and CTE. We had identified NXF1 (formerly named TAP) as essential export receptor for the CTE RNA and as key transporter of cellular mRNA. NXF1 is the first identified export receptor for cellular mRNAs, thereby providing a direct link between mRNA and the nuclear pore complex. We further found that the NXF1 ortholog in C. elegans is an essential mRNA export factor, demonstrating conservation of NXF1 function in metazoa (Tan, W. et al. RNA. 6:1762-1772, 2000). Recent data from my lab revealed that NXF1 binds directly to the splicing factor U2AF (Zolotukhin et al, submitted). This interaction provides a novel link between splicing and export of mRNA. In collaboration with George Pavlakis, we identified a novel potent RNA transport element RTE (Nappi, F., et al.. J Virol. 75:45584569, 2001). We found that RTE function is conserved in vertebrates. The use of RTE provides us with a powerful tool for further dissection of mRNA transport mechanisms.