The glucocorticoid receptor as signal integrator: studying ALL drug resistance

描述(由申请人提供)：蛋白质位于信号通路的节点，他们的任务是整合这些信号以指导特定的输出--一种根据细胞当时的需要量身定做的输出。迈尔斯·普法尔博士研究了使用糖皮质激素受体(GR)的信号整合，特别是DNA序列调节变构的机制基础。K99/R00奖项将为他提供资源、时间和培训，以开发系统，将GR变构整合的结构和生物物理基础转化为对导致儿童前B期急性淋巴细胞白血病(B-ALL)治疗耐药的信号异常的理解。这项工作将使选择性干预措施的发展成为可能，这些干预措施可以指导蛋白质中的变构网络。这一奖项将使普法尔博士能够实现以下职业发展目标：1)为造血发展和疾病奠定基础；2)学习使用初级组织；3)获得基因组数据集方面的经验，以发展未来的研究；4)开发一种测试信号整合原理的体外系统。 糖皮质激素受体通过在基因组中特定的DNA反应元件上组装调控复合体来协调一系列基因表达，以响应细胞信号。在B-ALL中，针对GR的合成糖皮质激素是一种有效的治疗方法，与其他药物联合使用，可诱导细胞凋亡。然而，对糖皮质激素无效的患者将面临严峻的预后。普法尔博士假设，调节GR的变构信号通路已经被破坏，改变了受体的正常功能，并阻止了细胞凋亡。他将从三个方面验证这一假说：1)确定糖皮质激素诱导的基因表达程序、GR结合和活性染色质标记在糖皮质激素敏感和耐药的急性淋巴细胞白血病中的变化；2)从选定的反应元件中提纯GR调节复合体并鉴定成分；以及3)识别来自GR DNA结合域的变构网络，并确定信号如何影响这些网络。 与公共健康相关：糖皮质激素通过其受体GR调节基因表达。激活GR的药物用于多种疾病，从哮喘到器官移植再到癌症，如急性淋巴细胞性白血病。然而，这些药物并不总是有效的，它们也有严重的副作用，包括糖尿病、骨质疏松症和高血压。因此，了解GR如何整合和响应细胞信号对健康以及检测、治疗和治愈疾病具有重要意义。

DESCRIPTION (provided by applicant): Proteins lie at the nodes of signaling pathways, and it is their task to integrate these signals to direct a specific output - one that is tailored to the needs of the cell at the time. Dr. Miles Pufall has studied signal integration using the glucocorticoid receptor (GR), in particular the mechanistic basis of allosteric regulation by DNA sequence. This K99/R00 award will provide him with the resources, time, and training to develop systems that translate the structural and biophysical basis of GR allosteric integration into an understanding of the signaling aberrations that result in childhood pre-B acute lymphoblastic leukemia (B-ALL) treatment resistance. This work will enable development of selective interventions that direct allosteric networks in the protein. This award will allow Dr. Pufall to achieve the following career development goals: 1) Develop a grounding in hematopoietic development and disease; 2) Learn to work with primary tissue; 3) Gain experience with genomic data sets to develop future studies; and 4) Develop an in vitro system to test signal integration principles. The glucocorticoid receptor orchestrates a program of gene expression in response to cellular signals by nucleating the assembly of regulatory complexes at specific DNA response elements throughout the genome. In B-ALL, synthetic glucocorticoids directed against GR are an effective treatment, which in combination with other drugs, work to induce an apoptotic program. However those who do not respond to glucocorticoids face a grim prognosis. Dr. Pufall hypothesizes that signaling pathways that allosterically regulate GR have been disrupted, changing the normal function of the receptor, and blocking apoptosis. He will test this hypothesis in three aims: 1) Identify changes in glucocorticoid induced gene expression programs, GR binding, and marks for active chromatin in glucocorticoid sensitive and resistant acute lymphoblastic leukemia; 2) Purify GR regulatory complexes from select response elements and identify components; and 3) Identify allosteric wires emanating from the DNA binding domain of GR, and determine how signals impinge on these wires. PUBLIC HEALTH RELEVANCE: Glucocorticoid hormones act through their receptor, GR, to regulate gene expression. Drugs that activate GR are used for a wide range of conditions, from asthma to organ transplants to cancer, such as acute lymphoblastic leukemia. However, these drugs are not always effective, and they also have serious side effects, including diabetes, osteoporosis, and hypertension. Thus, understanding how GR integrates and responds to cellular cues has important implications for health, and for detecting, treating and curing disease.