Control of T Lymphopoiesis and Growth by Mad Genes

描述(由申请人提供)：抗原激活的淋巴细胞，或白血病中的转化淋巴细胞，必须将其大小和含量增加一倍(称为细胞生长)，才能分裂成相同大小的子细胞。尽管细胞生长在正常细胞增殖和癌症中很重要，但在分裂的淋巴细胞或其他哺乳动物细胞中控制细胞生长的分子事件仍然是一个谜。我们在小鼠身上的初步研究表明，Mad家族的碱性螺旋-环-螺旋转录因子(Mad1、Mxi1、Mad3、Mad4)被认为是Myc癌蛋白的拮抗剂，部分通过抑制细胞生长来抑制T细胞的增殖和发育。此外，人类Mad1和mxi1基因分别定位于与淋巴细胞白血病、霍奇金氏病和前列腺癌相关的染色体区域，这表明MAD基因在淋巴细胞生物学和癌症中的重要性。本研究的主要目的是确定Mad家族成员在T淋巴细胞发育和扩增中的正常作用和作用机制。具体地说，我们打算：(1)验证Mad家族成员调节T淋巴细胞成熟的假说。我们将通过在小鼠中选择Mad家族成员的靶向缺失和转基因过表达来研究Mad家族成员(S)在T淋巴细胞发育中的作用。(2)验证Mad家族成员在T细胞活化过程中控制T淋巴细胞增殖和细胞生长(细胞大小、蛋白质合成)的假设。我们将研究Mad过度表达或丢失对细胞分裂、细胞大小、RNA处理和激活后立即合成的初级淋巴细胞的功能影响。(3)验证Mad直接结合和调节参与细胞生长控制的必要基因表达的假设。我们将确定Mad1是否抑制了与细胞生长控制有关的几个必要基因的表达。然后，我们将使用染色质免疫沉淀分析来确定这些基因的调节区是否直接与Mad蛋白结合。总而言之，这些目标将检验Mad-Max复合体通常部分通过控制生长调节基因的表达来调节淋巴细胞增殖和发育的总体假设。这些研究的结果将确定目标基因，这些基因可以被操纵来调节Myc和Mad之间的平衡，从而抑制淋巴瘤或自身免疫性疾病中的淋巴细胞增殖，或者在初级免疫反应中或在骨髓移植后促进抗原特异性淋巴细胞的克隆性扩张。

DESCRIPTION (provided by applicant): Antigen-activated lymphocytes, or transformed lymphocytes in leukemias, must double their size and contents (termed cell growth) before they can divide into equal sized daughter cells. Despite the importance of cell growth in normal cell proliferation and cancer, the molecular events that control cell growth in dividing lymphocytes or other mammalian cells remain an enigma. Our preliminary studies in mice suggest that the Mad family of basic helix-loop-helix transcription factors (Mad1, Mxi1, Mad3, Mad4), considered to be antagonists of the Myc oncoprotein, inhibit T cell proliferation and development in part by inhibiting cell growth. Furthermore, human mad1 and mxi1 genes each localize to separate chromosome regions associated with lymphocytic leukemias, Hodgkin's disease, and prostatic carcinomas suggesting the importance of mad genes in lymphocyte biology and cancer. The broad objective of this proposal is to determine the normal roles and mechanism of action of Mad family members in the development and expansion of T Iymphocytes Specifically, we intend to: (1) Test the hypothesis that Mad family members modulate the maturation of T lymphocytes. We will examine the role(s) of Mad family members in T lymphocyte development by employing targeted deletion, and transgenic overexpression, of selected Mad family members in mice. (2) Test the hypothesis that Mad family members control the proliferation and cell growth (cell size, protein synthesis) of T Iymphocytes during T cell activation. We will examine the functional consequences of Mad overexpression or loss on cell division, cell size, RNA processing, and protein synthesis in primary lymphocytes immediately following activation. (3) Test the hypothesis that Mad directly binds and modulates the expression of essential genes involved in cell growth control. We will determine if Mad1 inhibits the expression of several essential genes involved in cell growth control. We will then use chromatin immunoprecipitation assays to determine if the regulatory regions of these genes are directly bound by Mad proteins. Together, these aims will test the overall hypothesis that Mad-Max complexes normally modulate lymphocyte proliferation and development in part by controlling the expression of growth-regulating genes. Results of these studies will identify target genes that could be manipulated to regulate the balance between Myc and Mad in order to inhibit lymphocyte proliferation in lymphomas or autoimmune disease, or to enhance clonal expansion of antigen-specific lymphocytes in a primary immune response, or following bone marrow transplantation.