INVESTIGATING THE MECHANISMS OF PRB TUMOR SUPPRESSION

视网膜母细胞瘤肿瘤抑制基因是肿瘤抑制基因的原型。 它的蛋白质pRB是一种在绝大多数人类肿瘤中被破坏的完整蛋白质。 因此，了解pRB肿瘤抑制的潜在机制至关重要。pRB作用的分子机制是什么？ 在各种实验环境中，pRB可以介导细胞周期停滞，促进细胞分化，抑制细胞凋亡，抑制某些转录因子（E2 F）的转录，并激活其他转录因子（myoD和C/EBP）的转录。 这些活性主要在人工测定系统中进行了研究，目前还不清楚这些pRB特性中哪一个在体内最重要。 已知pRB的大结构域，即口袋，对于大多数pRB的活性是必要的和足够的。嵌合小鼠髓RB -/-细胞的研究表明，pRB的损失并没有删除细胞周期控制的大多数方面。 在这样的小鼠中，RB -/-细胞对大多数组织有贡献并且具有正常的组织学。 我们假设，pRB通常不是细胞周期控制所必需的，但它必须具有与肿瘤发生相关的非常特定的作用。 戴森实验室的初步数据表明，细胞基因组完整性的维持可能取决于pRB诱导细胞周期停滞以响应DNA损伤的能力。 最近的研究结果还表明，pRB的能力，以促进分化与致瘤性。 该提案将研究pRB途径在体内的意义，导致细胞周期停滞，以响应DNA损伤和细胞分化的pRB肿瘤抑制。具体目标1：鉴定pRB诱导细胞周期阻滞以响应DNA损伤和促进细胞分化的重要结构特征。 这两个途径必不可少的pRB相关蛋白将被确定。 具体目标2：为了确定pRB阻止细胞周期和促进细胞分化的能力对于pRB在体内充当肿瘤抑制剂是否重要，使用目的1中产生的RB突变体来使这些特异性途径失效。 具体目标3：确定E2 F依赖性转录的失调是否与RB -/-细胞体内致瘤潜力相关。

The retinoblastoma tumor suppressor gene is a prototype for tumor suppressor genes. Its protein, pRB, is an integral protein that is disrupted in a vast majority of human tumors. Understanding the underlying mechanisms of pRB tumor suppression is, therefore, of critical importance. What is the molecular mechanism of pRB action? In various experimental settings, pRB can be shown to mediate cell cycle arrest, promote cellular differentiation, suppress apoptosis, repress transcription of some transcription factors (E2F) and activate transcription with others (myoD and C/EBP). These activities have been studied primarily in artificial assay systems and currently it is unclear which of these pRB properties are most important in-vivo. It is known that a large domain of pRB, the pocket, is necessary and sufficient for most of pRB's activities. Studies of chimeric mice pith RB -/- cells demonstrate that loss of pRB does not remove most aspects of cell cycle control. In such mice, RB -/- cells contribute to most tissues and have normal histology. We hypothesize, that pRB is not generally essential for cell cycle control but that it must have very specific roles relevant to tumorigenesis. Preliminary data from the Dyson laboratory suggests that maintenance of the genomic integrity of cells may be dependent upon pRB's ability to induce a cell cycle arrest in response to DNA damage. Recent findings also suggest that pRB's ability to promote differentiation correlates with tumorigenicity. This proposal will investigate the in-vivo significance of pRB pathways leading to cell cycle arrest in response to DNA damage and to cellular differentiation in pRB tumor suppression. Specific Aim number 1: To identify the structural features important for pRB to induce cell cycle arrest in response to DNA damage and to promote cellular differentiation. The pRB-associated proteins essential to these two pathways will be identified. Specific Aim number 2: To determine whether the ability of pRB to arrest cell cycle and promote cellular differentiation is important for pRB to act as a tumor suppressor in-vivo using the RB mutants generated in aim number 1 to disable these specific pathways. Specific Aim number 3: To determine whether the deregulation of E2F dependent transcription is correlated with the tumorigenic potential of RB -/- cells in-vivo.