C21, a Transcriptional Regulator in Hematopoiesis

描述（由申请人提供）：C21基因编码在转录调控中发挥作用的蛋白质家族。C21在小鼠造血细胞中的过度表达改变了骨髓的发育，并表明该家族的成员参与调节干细胞分化。在3 T3成纤维细胞中过表达C21增加了它们对凋亡刺激的抗性。C21与一类核辅阻遏物形成复合物，其中最知名的哺乳动物形式是N-CoR（核受体辅阻遏物）和SMRT（类维生素A和甲状腺受体的沉默介体）。C21与辅阻遏物结合，似乎干扰辅阻遏物的泛素介导的蛋白水解，导致辅阻遏物浓度升高。该家族的成员在胎儿造血组织以及许多造血细胞系中以高水平表达。像许多WD 40蛋白一样，C21家族成员似乎通过充当衔接子或桥梁来发挥作用，促进不直接相互作用的蛋白质的相互作用。本建议涉及四个目标。 目标1.鉴定与C21相互作用的蛋白质。C21家族蛋白充当衔接子，将必须在功能上相互作用但彼此缺乏内在亲和力的分子聚集在一起。识别相互作用分子对于理解C21家族如何调控转录至关重要。目标2.确定C21如何改变转录调控。目标3。通过识别这种相互作用调节的途径来分析C21与阻遏物复合物相互作用的后果。微芯片表达阵列将与从成纤维细胞，造血细胞系和胚胎干细胞（ES）细胞，条件性过表达C21的cDNA进行探测，以确定响应C21表达的变化的途径。目标4。确定体内相互作用的功能后果。将研究C21 cDNA在转基因小鼠和培养物中的过表达和靶向缺失的影响。本研究将利用小鼠胚胎干细胞（ESC）分析C21 cDNA过表达对早期造血发育的影响，并通过研究视黄酸诱导HL 60和U937髓系白血病细胞分化来检测C21 cDNA过表达对髓系发育的影响。

DESCRIPTION (provided by applicant): The C21 gene encodes a family of proteins that play a role in transcriptional regulation. Over-expression of C21 in mouse hematopoietic cells alters myeloid development and suggest that members of this family are involved in regulating stem cell differentiation. Over-expressing C21 in 3T3 fibroblasts increases their resistance to apoptotic stimuli. C21 forms a complex with the class of nuclear co repressors of which the best known mammalian forms are N-CoR (nuclear receptor co-repressor) and SMRT (silencing mediator of retinoid and thyroid receptor). C21 binds to the co-repressors and appears to interfere with the ubiquitin-mediated proteolysis of the co-repressors, causing an elevation of the co-repressor concentration. Members of the family are expressed at high levels in fetal hematopoietic tissues as well as in many hematopoietic cell lines. Like many WD40 proteins, C21 family members appear to act by serving as an adaptor or bridge, facilitating the interaction of proteins that do not interact directly. Four Aims are addressed in this proposal. Aim 1. To identify the proteins that interact with C21. C21 family proteins act as adaptors, bringing together molecules that must interact functionally but that lack intrinsic affinity for each other. The identification of the interacting molecules is critical for any understanding of how C21 family to regulate transcription. Aim 2. To determine how C21 alters transcriptional regulation. Aim 3. To analyze the consequences of the interaction of C21 with the repressor complex by identifying the pathways regulated by this interaction. Microchip expression arrays will be probed with cDNAs obtained from fibroblasts, hematopoietic cell lines and embryonic stem (ES) cells that conditionally over-express C21 to identify the pathways that respond to alterations in C21 expression. Aim 4. To identify the functional consequences of the interaction in vivo. The effects of over-expression and targeted deletion of C21 cDNA in transgenic mice and in culture will be studied. Mouse embryonic stem cells (ESC) will be used to analyze the effects of over-expression of C21 cDNA on early hematopoietic development and the effects on myeloid development will be examined by studying the retinoic acid-induced differentiation of HL60 and U937 myeloid leukemia cells.