Regulation of RNA metabolism and cell growth control by SR protein kinases

转录后基因表达调控对细胞生长控制和动物生长至关重要 发展。该项目攻击SRPK1和SRPK2，这两个进化保守的激酶对应于 哺乳动物细胞中的剪接因子和调节因子SR家族的主要激酶活性。SR蛋白质 参与RNA代谢的许多方面，所有这些似乎都受到磷酸化的调节。 令人惊讶的是，我们最近发现SRPK1基因缺失的小鼠胚胎成纤维细胞转化为 软琼脂试验和裸鼠试验。此外，SRPKs定位于细胞质中，可以 被诱导对特定信号作出反应而移位到核仁。因此，SRPK从根本上说是 对细胞生长控制很重要，而激酶系统可能是由信号调节的。 基于我们在本项目前几个资助期建立的专业知识，长期 对基因表达的转录后调控感兴趣，并广泛发表和未发表 根据调查结果，我们建议在下一阶段研究的三个具体目标下继续这一项目。目标1 是建立条件性基因敲除小鼠模型，以确定SRPK1和SRPK1的功能需求 2在小鼠发育过程中，鉴定了小鼠胚胎成纤维细胞中的激酶基因敲除，为其提供 SRPKs调节哺乳动物细胞中RNA代谢中SR蛋白功能的遗传学证据。目标 2是通过测试三个特定的假设来解决SRPK1可能的肿瘤抑制活性：(1) SRPK1可能通过SR蛋白调节癌基因和抑癌基因的选择性剪接； 介导的磷酸化可能作用于重组新输出的mRNA-蛋白质复合体以调节 在细胞质中的翻译；(3)SRPK1介导的磷酸化在维持 通过调节转录和剪接之间的耦合来实现基因组的稳定性。目标3是了解如何 SRPKs可能受信号调节。一组与SRPK相互作用的蛋白质已经被鉴定并 许多都是各种信号转导途径的已知组成部分。我们设计了具体的实验来 了解一些特定的信号如何通过SRPKs转导来调节基因表达 转录水平。总而言之，该项目将剖析一条前所未有的细胞转化途径。

Post-transcriptional regulation of gene expression is critical for cell growth control and animal development. This project attacks SRPK1 and 2, two evolutionary conserved kinases that correspond to the major kinase activity for the SR family of splicing factors and regulators in mammalian cells. SR proteins are involved in many aspects of RNA metabolism, all of which appear to be regulated by phosphorylation. Strikingly, we recently found that mouse embyo fibroblasts genetically deleted of SRPK1 are transformed in the soft agar assay and in nude mice. Furthermore, SRPKs are localized in the cytoplasm and can be induced to translocate to the nucelus in response to specific signals. Thus, SRPKs are fundamentally important for cell growth control and the kinase system may be regulated by signaling. Building upon our expertise established in the previous funding periods of this project, long-term interest in post-transcriptional regulation of gene expression, and extensive published and unpublished findings, we propose to continue this project under three specific aims for the next phase of research. Aim 1 is to develop conditional knockout mice models to determine the functional requirement for both SRPK1 and 2 during mouse development and characterize the kinase knockout mouse embryo fibroblasts to provide genetic evidence that SRPKs regulate the function SR proteins in RNA metabolism in mammalian cells. Aim 2 is to address the putative tumor suppressor activity of SRPK1 by testing three specific hypotheses: (1) SRPK1 may regulate alternative splicing of oncogenes and tumor supressors via SR proteins; (2) SRPK1- mediated phosphorylation may act to reorganize newly exported mRNA-protein complex to regulate translation in the cytoplasm; and (3) SRPK1-mediated phosphorylation may play a critical role in maintaining genomic stability by modulating the coupling between transcription and splicing. Aim 3 is to understand how SRPKs might be regulated by signaling. A panel of SRPK-interacting proteins have been identified and many are known components of various signal transduction pathways. We design specific experiments to understand how some specific signals may be transduced via SRPKs to regulate gene expression at post- transcriptional levels. Together, the project will dissect an unprecedented cellular transformation pathway.