Gene Expression At Beginning Of Mammalian Development

哺乳动物发育初期的基因表达

• 批准号: 7201708
• 负责人: Melvin DePamphilis
• 金额: --
• 依托单位: EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7201708
• 关键词: CpG islands; DNA methylation; DNA replication; RNA binding protein; acrosome; cell differentiation; cell line; developmental genetics; early embryonic stage; embryonic stem cell; genetic regulatory element; immunoprecipitation; laboratory mouse; mammalian embryology; nucleolus; phosphorylation; protein localization; protein protein interaction; protein quantitation /detection; transcription factor

Kohn, M.J., K.J. Kaneko and M.L. DePamphilis (2005) DkkL1(Soggy), A Dickkopf Family Member, Localizes To The Acrosome During Mammalian Spermatogenesis, Mol. Reprod. Dev. 71:516-522. Dickkopf-like 1 (DkkL1) is related to the Dickkopf gene family, a group of proteins that are characterized as secreted antagonists of Wingless signal transduction proteins. DkkL1 mRNA is found in preimplantation mouse embryos and in developing neural tissue, but in adults it is found primarily in the testes. In an effort to elucidate its function, the distribution of Dickkopf-like 1 protein (DkkL1) in mouse testis and mature sperm was analyzed by immuno-histochemistry and immuno-blotting techniques. DkkL1 first appeared in the developing spermatocytes in seminiferous tubules as early as Stage XII, coincident with the appearance of DkkL1 mRNA. Surprisingly, however, DkkL1 localized to the developing acrosome in spermatocytes and spermatids and to the acrosome in mature sperm. Furthermore, DkkL1 was N-glycosylated in the testis, but it did not appear to be excreted, and the DkkL1 in mature sperm was no longer N-glycosylated, suggesting that additional post-translational modifications occurred during the final stages of spermatogenesis. These results identify a member of the Dickkopf family as a novel acrosomal protein that may be involved in acrosome assembly or function, a unique role for a secreted signaling molecule. Kaneko, K.J., T. Rein, Z-S Guo, K. Latham and M.L. DePamphilis. (2004) DNA Methylation May Restrict But Does Not Determine Differential Gene Expression at the Sgy/Tead2 locus During Mouse Development, Mol. Cell. Biol. 24:1968-1982. Soggy (Sgy) and Tead2, two closely linked genes with CpG islands, were coordinately expressed in mouse preimplantation embryos and embryonic stem (ES) cells, but were differentially expressed in differentiated cells. Analysis of established cell lines revealed that Sgy gene expression could be fully repressed by methylation of the Sgy promoter, and that DNA methylation acted synergistically with chromatin deacetylation. Differential gene expression correlated with differential DNA methylation, resulting in sharp transitions from methylated to unmethylated DNA at the open promoter in both normal cells and tissues, as well as in established cell lines. However, neither promoter was methylated in normal cells and tissues even when its transcripts were undetectable. Moreover, the Sgy promoter remained unmethylated as Sgy expression was repressed during ES cell differentiation. Therefore, DNA methylation was not the primary determinant of Sgy/Tead2 expression. Nevertheless, Sgy expression was consistently restricted to basal levels whenever downstream regulatory sequences were methylated, suggesting that DNA methylation restricts but does not regulate differential gene expression during mouse development. Intine, R. V., M. Dundr, A. Vassilev, E. Schwartz, Y. Zhou, Y. Zhao, M. L. DePamphilis, and R. J. Maraia (2004) Nonphosphorylated Human La Antigen Interacts With Nucleolin At Nucleolar Sites Involved In rRNA Biogenesis, Mol. Cell. Biol. 24:10894-904. La is an abundant RNA-binding protein that has been proposed to function in multiple pathways involved in the production of tRNAs, ribosomal proteins and other components of the translational machinery (Kenan and Keene, 2004 Nat Struct & Mol Biol, 11:306). Human (h)La is either phosphorylated or nonphosphorylated on serine-366 (S366, pLa & npLa, respectively). In addition to its conserved N-terminal domain, hLa contains a C-terminal domain that contains an atypical RNA recognition motif (RRM) and short basic motif (SBM), whose distinct mode of RNA binding is inhibited by S366 phosphorylation. We report that npLa is most concentrated in nucleolar foci that harbor fibrillarin, nucleolin, and nascent pol I transcripts. Immunoprecipitation of native or tagged hLa as well as yeast two hybrid analysis and fluorescence resonance energy transfer (FRET) in the nucleolus, reveals association with nucleolin. HLa lacking the SBM fails to localize to nucleoli and does not exhibit FRET with nucleolin. The data indicate that the nonphosphorylated SBM directs interaction with nucleolin, and provides evidence that the npLa specifically is involved in nucleolar biology related to ribosome biogenesis. DePamphilis, M.L. (2004) Mammalian Development, Regulation of Gene Expression in, in Meyers, R.A. (Ed.): Encyclopedia of Molecular Cell Biology and Molecular Medicine, volume 6, pp. ??, WILEY-VCH Verlag, Weinheim, Germany. (http://www.wiley-vch.de/books/emcbmm2/index.html)

科恩，M.J.，K.J.金子和 M.L. DePamphilis (2005) DkkL1(Soggy)，迪克科普夫家族成员，在哺乳动物精子发生过程中定位于顶体，分子生物学。重现。开发。 71：516-522。 Dickkopf-like 1 (DkkL1) 与 Dickkopf 基因家族相关，该家族是一组以 Wingless 信号转导蛋白的分泌拮抗剂为特征的蛋白质。 DkkL1 mRNA 存在于植入前小鼠胚胎和发育中的神经组织中，但在成人中，它主要存在于睾丸中。为了阐明其功能，通过免疫组织化学和免疫印迹技术分析了 Dickkopf 样 1 蛋白 (DkkL1) 在小鼠睾丸和成熟精子中的分布。 DkkL1 早在第 XII 阶段就首次出现在生精小管中正在发育的精母细胞中，与 DkkL1 mRNA 的出现一致。然而，令人惊讶的是，DkkL1 定位于精母细胞和精子细胞中正在发育的顶体以及成熟精子中的顶体。此外，DkkL1在睾丸中被N-糖基化，但似乎没有被排出体外，并且成熟精子中的DkkL1不再被N-糖基化，这表明在精子发生的最后阶段发生了额外的翻译后修饰。这些结果将 Dickkopf 家族的成员鉴定为一种新型顶体蛋白，可能参与顶体组装或功能，这是分泌信号分子的独特作用。 Kaneko, K.J.、T. Rein、Z-Suo、K. Latham 和 M.L.德潘菲利斯。 (2004) DNA 甲基化可能限制但不决定小鼠发育过程中 Sgy/Tead2 基因座的差异基因表达，Mol。细胞。生物。 24：1968-1982。 Soggy (Sgy) 和 Tead2 是两个与 CpG 岛紧密相连的基因，在小鼠植入前胚胎和胚胎干 (ES) 细胞中协调表达，但在分化细胞中差异表达。对已建立细胞系的分析表明，Sgy 启动子的甲基化可以完全抑制 Sgy 基因的表达，并且 DNA 甲基化与染色质脱乙酰化具有协同作用。差异基因表达与差异DNA甲基化相关，导致正常细胞和组织以及已建立的细胞系中开放启动子处的DNA从甲基化到非甲基化的急剧转变。然而，即使在正常细胞和组织中无法检测到其转录本，这两个启动子都没有甲基化。此外，Sgy 启动子保持未甲基化，因为 Sgy 表达在 ES 细胞分化过程中受到抑制。因此，DNA甲基化不是Sgy/Tead2表达的主要决定因素。然而，每当下游调控序列甲基化时，Sgy 表达始终限制在基础水平，这表明 DNA 甲基化限制但不调节小鼠发育过程中的差异基因表达。 Intine, R. V.、M. Dundr、A. Vassilev、E. Schwartz、Y. Zhou、Y. Zhao、M. L. DePamphilis 和 R. J. Maraia (2004) 非磷酸化人 La 抗原在参与 rRNA 生物发生的核仁位点与核仁蛋白相互作用，分子生物学。细胞。生物。 24：10894-904。 La 是一种丰富的 RNA 结合蛋白，已被认为在涉及 tRNA、核糖体蛋白和翻译机器其他组件的产生的多种途径中发挥作用（Kenan 和 Keene，2004 Nat Struct & Mol Biol，11：306）。人 (h)La 在丝氨酸 366 上被磷酸化或非磷酸化（分别为 S366、pLa 和 npLa）。除了保守的 N 端结构域外，hLa 还包含一个 C 端结构域，其中包含非典型 RNA 识别基序 (RRM) 和短基本基序 (SBM)，其独特的 RNA 结合模式受到 S366 磷酸化的抑制。我们报告 npLa 最集中于含有纤维蛋白、核仁素和新生 pol I 转录本的核仁灶。天然或标记 hLa 以及酵母两种杂交分析和核仁中荧光共振能量转移 (FRET) 的免疫沉淀揭示了与核仁蛋白的关联。缺乏 SBM 的 HLa 无法定位到核仁，并且不表现出与核仁素的 FRET。数据表明，非磷酸化的 SBM 指导与核仁素的相互作用，并提供了 npLa 特别参与与核糖体生物发生相关的核仁生物学的证据。 德潘菲利斯，M.L. (2004) 哺乳动物发育，基因表达调控，Meyers, R.A. （主编）：分子细胞生物学和分子医学百科全书，第 6 卷，第 10 页，WILEY-VCH Verlag，魏因海姆，德国。 （http://www.wiley-vch.de/books/emcbmm2/index.html）