SCL GENE AND HEMATOPOIETIC DEVELOPMENT

SCL 基因与造血发育

• 批准号: 2225221
• 负责人: STEPHEN J. BRANDT
• 金额: $ 11.41万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-01-01 至 1997-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2225221
• 关键词: DNA binding protein; RNase protection assay; antisense nucleic acid; cell growth regulation; developmental genetics; embryo /fetus cell /tissue; embryonic stem cell; gene expression; genetic promoter element; genetic transcription; genetic translation; hematopoiesis; hematopoietic stem cells; immunocytochemistry; immunoprecipitation; in situ hybridization; laboratory mouse; messenger RNA; polymerase chain reaction; protein isoforms; protein structure function; tissue /cell culture; transcription factor; transfection; western blottings; DNA binding protein; RNase protection assay; antisense nucleic acid; cell growth regulation; developmental genetics; embryo /fetus cell /tissue; embryonic stem cell; gene expression; genetic promoter element; genetic transcription; genetic translation; hematopoiesis; hematopoietic stem cells; immunocytochemistry; immunoprecipitation; in situ hybridization; laboratory mouse; messenger RNA; polymerase chain reaction; protein isoforms; protein structure function; tissue /cell culture; transcription factor; transfection; western blottings

The proposed research will investigate the expression and functions of the protein products of the SCL gene during hematopoietic development in the mouse. This gene was first identified at the breakpoint of a chromosomal translocation in human T-cell leukemia and is a member of a family of transcription factors whose members are recognized to be regulators of differentiation. The results of these experiments should provide insight into basic mechanisms of hematopoiesis. Since activation of this gene by chromosomal rearrangement is the most common genetic lesion in acute T-cell leukemia, these studies should also contribute to our understanding of leukemogenesis. The first specific aim of this work is to characterize the translational products of the murine SCL gene in hematopoietic cells. The protein products of the murine SCL gene will be characterized in hematopoietic cell lines and mouse tissues by radioimmunoprecipitation and Western blot analysis. Subcellular localization of SCL proteins will be determined by transient expression in COS cells and in cell lines stably expressing individual proteins. The second specific aim is to characterize the transcriptional and translational products of the murine SCL gene during hematopoietic development. SCL mRNA expression will be studied with RNase protection, in situ hybridization, and the polymerase chain reaction (PCR) and SCL protein expression by immunocytochemistry in staged mouse embryos. The transcriptional and translational products of the gene will also be characterized in an in vitro model of hematopoietic development using embryonic stem (ES) cell-derived embryoid bodies. The third specific aim is to define the actions of SCL proteins in hematopoietic development. This will be investigated in gain of function and loss of function mutants of ES cells made to undergo differentiation. The fourth specific aim is to determine the ability of SCL proteins to function as transcriptional regulators during hematopoietic development. The ability of SCL proteins to transactivate artificial and, if identified, physiologic targets will be determined. The possibility that certain SCL proteins may have different transcriptional potencies and act as transcriptional antagonists will be investigated. Cellular targets will be identified using a functional strategy based on the isolation of SCL-inducible promoters, and the ability of different SCL isoforms to activate such promoters will be defined.

拟议的研究将研究 造血性发育过程中SCL基因的蛋白质产物 在鼠标中。 该基因首先是在一个断点 人类T细胞白血病中的染色体易位，是 一个转录因素的家庭，其成员被认为是 差异化的监管机构。 这些实验的结果应 提供对造血的基本机制的见解。 自从 染色体重排激活该基因是最常见的 急性T细胞白血病中的遗传病变，这些研究也应 有助于我们对白血病的理解。 第一个特定 这项工作的目的是表征 造血细胞中的鼠SCL基因。 的蛋白质产物 鼠类SCL基因将在造血细胞系和 小鼠组织通过放射免疫沉淀和蛋白质印迹分析。 SCL蛋白的亚细胞定位将由 COS细胞和细胞系中的瞬时表达稳定表达 单个蛋白质。 第二个具体目的是表征 鼠SCL基因的转录和翻译产物 在造血发育期间。 将研究SCL mRNA表达 使用RNase保护，原位杂交和聚合酶链 通过免疫细胞化学的反应（PCR）和SCL蛋白表达 分阶段的小鼠胚胎。 转录和翻译产品 该基因的体外模型也将在 使用胚胎茎（ES）细胞衍生的造血发育 胚胎体。 第三个具体目的是定义 造血发育中的SCL蛋白。 这将被调查 在功能的获取和ES细胞功能突变体的丧失中 经历差异化。 第四个具体目的是确定 SCL蛋白充当转录调节剂的能力 在造血发育期间。 SCL蛋白的能力 反式激活人工，如果鉴定出生理目标 可以确定。 某些SCL蛋白可能具有的可能性 不同的转录效力并充当转录 对抗者将进行调查。 将确定细胞目标 使用基于SCL诱导的隔离功能策略 启动子，以及不同SCL同工型激活此类的能力 发起人将被定义。