MOLECULAR BASIS OF ACUTE PROMYELOCYTIC LEUKEMIA

急性早幼粒细胞白血病的分子基础

• 批准号: 6522917
• 负责人: J DON CHEN
• 金额: $ 5.61万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-01 至 2003-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6522917
• 关键词: DNA binding protein; acute myelogenous leukemia; apoptosis; cell differentiation; chimeric proteins; chromosome translocation; gene induction /repression; genetic transcription; immunofluorescence technique; immunoprecipitation; intermolecular interaction; molecular oncology; neoplasm /cancer genetics; oncoproteins; retinoid binding proteins; tissue /cell culture; transcription factor; transport proteins; yeast two hybrid system

DESCRIPTION: (Adapted from the investigator's abstract) The t(15;17) translocation accounts for over 95 percent of all human acute promyelocytic leukemia (APL) cases. This translocation fuses the PML gene with the nuclear receptor for retinoic acid (RARa) and creates an oncogenic fusion protein, PML-RARa. It is thought that PML-RARa may interfere with the function of wild-type PML and RARa. Immunocytochemistry studies reveal the localization of PML at discrete nuclear domains known as PML oncogenic domains (PODs), Kr bodies, ND10, or nuclear bodies. While the exact function of the PODs remains unclear, its structure is disrupted in t(15;17)-translocation APL cells, suggesting an important roles for the PODs in APL. To investigate the function of the PODs, they have screened for PML-interacting factors using the yeast two-hybrid system. They have isolated three specific interacting proteins including a previously demonstrated PML-modifier, SUMO-1, and two novel proteins that are implicated in transcriptional regulation. In this project, they will investigate the interactions of PML with these two novel cofactors. Standard techniques including the yeast two-hybrid assay, GST pull down, Far-Western blot, and immunofluorescence microscopy will be utilized to establish the interaction of PML with these cofactors. Based on their preliminary studies, they propose that these two cofactors are transcriptional repressors. Experiments will be conducted to determine the mechanism of transcriptional repression by these proteins. They propose that the roles of PML and the PODs are to recruit these cofactors to the restricted PML domains by protein-protein interaction with PML. They further propose that localization of these cofactors to the PODs will lead to inactivation of their transcriptional repressor function. These hypotheses will be tested by a combination of transient transfection and immunocytochemistry studies. In addition, they will investigate the role of these PML cofactors in APL, based on observations that these two proteins also interact with the APL oncogenic fusion protein, PML-RARa. Together, these studies should shed new lights into the function of PML and its nuclear domains, as well as providing new insights to the function and regulation of the new cofactors. A better understanding of mechanism of APL at the molecular and cellular levels should lead to a more effective diagnosis and treatment of this disease, as well as providing implications for other malignancies.

描述：（改编自研究者的摘要）t(15;17) 易位占所有人类急性早幼粒细胞疾病的 95% 以上 白血病（APL）病例。这种易位将 PML 基因与核融合 视黄酸（RARa）受体并产生致癌融合蛋白， PML-RARa。据认为 PML-RARa 可能会干扰 野生型 PML 和 RARa。免疫细胞化学研究揭示了 PML 位于离散核域，称为 PML 致癌域 (POD)，Kr 体、ND10 或核体。虽然 POD 的确切功能仍然存在 尚不清楚，其结构在 t(15;17)-易位 APL 细胞中被破坏， 表明 POD 在 APL 中发挥重要作用。研究该函数 在 POD 中，他们使用酵母筛选了 PML 相互作用因子 双混合系统。他们分离出了三种特定的相互作用蛋白 包括先前演示的 PML 修改器 SUMO-1 和两个新颖的 与转录调控有关的蛋白质。在这个项目中， 他们将研究 PML 与这两种新型辅助因子的相互作用。 标准技术包括酵母双杂交测定、GST pull down、 Far-Western印迹和免疫荧光显微镜将用于 建立 PML 与这些辅助因子的相互作用。基于他们的 初步研究，他们提出这两个辅助因子是转录的 阻遏物。将进行实验以确定其机制 这些蛋白质的转录抑制。他们建议的角色 PML 和 POD 将招募这些辅助因子到受限的 PML 域 通过蛋白质-蛋白质与 PML 的相互作用。他们进一步建议本土化 这些 POD 辅助因子的加入将导致其失活 转录抑制功能。这些假设将由 瞬时转染和免疫细胞化学研究的结合。在 此外，他们还将研究这些 PML 辅因子在 APL 中的作用，基于 观察到这两种蛋白也与 APL 致癌蛋白相互作用 融合蛋白，PML-RARa。总之，这些研究应该为以下问题提供新的线索： PML 及其核域的功能，并提供新的见解 新辅助因子的功能和调节。更好地理解 APL在分子和细胞水平上的机制应该导致更多 对该疾病进行有效的诊断和治疗，并提供 对其他恶性肿瘤的影响。