CITED1 nuclear localization and its role in Wilms' tumor pathogenesis

描述(由申请者提供)：这一独立之路奖概述了最初为期两年的指导培训计划，然后是为期三年的独立阶段，以加快洛夫沃恩博士向科学独立的过渡。在最初的指导阶段，洛夫沃恩博士建议在范德比尔特大学高生产率的原理研究员和受人尊敬的导师马克·德·凯斯特克博士的指导下，通过一个多学科项目来扩大他在癌症和发育生物学领域的科学技能和知识。在系主任的认可下，洛夫沃恩博士目前正在实践一种工作努力公式，将他工作日75%的时间用于研究目的。本文详述的研究计划将为洛夫沃恩博士提供基本生化技术方面的严格培训，以测试Wilms瘤中转录辅助因子CITED1的核质穿梭的功能意义和调节机制。在De Caestecker博士的指导下，Lovvorn博士发现，CITED1在发育中正常活跃，但在成熟肾脏中缺失，在人和大鼠肾母细胞瘤中持续表达。他还表明，CITED1的亚细胞定位从后肾间充质的细胞质转移到恶性胚芽的细胞核，并且CITED1在原发人类肾母细胞瘤中的表达水平增加与疾病的晚期相关。Lovvorn博士从这些观察中提出了两个假说：1.CITED1的核定位促进了Wilms的肿瘤发生；2.CITED1的核质穿梭在Wilms的肿瘤发生中是失调的。在他的最初目标中，洛夫沃恩博士将测试将CITED1限制在Wilms1肿瘤细胞核上的致病意义。他将建立两个Wilms肿瘤细胞系，这两个细胞系在人类Wilms肿瘤细胞系WIT49中错误表达CITED1的主要胞浆或核部分，并将评估这些操作对Wilms肿瘤生物学的影响。在他的第二个目标中，洛韦德博士将利用质谱学，确定CITED1结合伙伴是任何一个亚细胞室所特有的。这些数据将有助于确定CITED1的修饰和调节其核保留在Wilms‘s肿瘤胚芽中的机制。这些研究不仅将为了解改变的CITED1核质穿梭及其在Wilms肿瘤发病机制中的作用提供一个框架，而且还将为未来的独立研究提供必要的培训。

DESCRIPTION (provided by applicant): This Pathway to Independence Award outlines an initial two-year mentored training program followed by a three-year independent phase to expedite the transition to scientific independence for Dr. Lovvorn. In the initial mentored phase, Dr. Lovvorn proposes to expand his scientific skills and knowledge in the fields of Cancer and Developmental Biology through a multidisciplinary program under the supervision of Dr. Mark de Caestecker, a highly productive principle investigator and respected mentor at Vanderbilt University. With the endorsement of his Department Chair, Dr. Lovvorn currently is practicing a work effort formula that affords 75% of his weekday time for research purposes. The research plan detailed herein will provide Dr. Lovvorn with rigorous training in basic biochemistry techniques to test the functional significance of and mechanisms regulating nucleocytoplasmic shuttling of the transcriptional co-factor CITED1 in Wilms' tumor. Under Dr. de Caestecker's guidance, Dr. Lovvorn has shown that expression of,CITED1, normally active in the developing but absent in the mature kidney, persists in both human and rat Wilms' tumors. He also has shown that the subcellular localization of CITED1 shifts from the cytoplasm in metanephric mesenchyme to the nucleus of malignant blastema, and that increasing levels of CITED1 expression in primary human Wilms' tumors correlate with advanced stages of disease. Dr. Lovvorn proposes two hypotheses from these observations: 1. CITED1 nuclear localization facilitates Wilms' tumor pathogenesis, and 2. Nucleocytoplasmic shuttling of CITED1 is dysregulated in Wilms' tumorigenesis. In his initial aim, Dr. Lovvorn will test the pathogenic significance of restricting CITED1 to the nucleus of Wilms1 tumor cells. He will generate two Wilms' tumor cell lines that misexpress either a predominantly cytosolic or nuclear fraction of CITED1 in the human Wilms' tumor cell line, WIT49, and will assess the effects of these manipulations on Wilms' tumor biology. In his second aim, Dr. Loworn, using mass spectrometry, will identify CITED1 binding partners that are specific to either subcellular compartment. These data will facilitate identification of CITED1 modifications and mechanisms regulating its nuclear retention in Wilms' tumor blastema. These studies will provide a framework not only to understand altered CITED1 nucleocytoplasmic shuttling in, and its contributions to, Wilms' tumor pathogenesis, but also the necessary training to pursue future independent investigation.