Aberrant LARG and RhoA activation in human leukemias

描述：(改编自研究人员摘要)与白血病相关的 Rho鸟嘌呤核苷酸交换因子(LARG)最近被鉴定为一种 急性髓系混合系白血病(MLL)蛋白的融合伙伴 白血病。LARG是快速扩展的DBL家族的新成员 通过激活RAS相关蛋白促进恶性转化的癌蛋白 Rho家族GTP酶。与其他DBL家族蛋白一样，LARG含有一个DBL 同源(DH)结构域，其功能是鸟核苷酸交换因子和 Rho GTP酶激活剂。DH域后面跟着一个Pleckstrin同源 (Ph)结构域，可能调节水解酶结构域的功能。LARG还包含一个 G蛋白信号转导(RGS)结构域的调节因子，提示它可能与G Rho GTP酶的蛋白偶联受体信号转导。我们的初步研究 确定LARG是RhoA的激活剂，并可导致 NIH3T3小鼠成纤维细胞。我们提出了四个具体目标来执行 详细分析了LARG的结构和功能。具体目标1将决定 DH域和PH域在RhoA激活LARG中的作用在……里面 具体而言，PH结构域是否调节一个 将确定磷脂酰肌醇3-激酶依赖的方式。特定的 Aim 2将评估RGS结构域在连接LARG和G蛋白中的作用 耦合的受体信号。这包括确定哪些是 异三聚体Gα亚基(S)受RGS结构域调控，其中G α亚基(S)反过来调节LARG水解域的激活。具体目标3 将确定肿瘤相关的MLL-LARG融合蛋白是否是一种异常 活化的LARG能促进上皮细胞的生长转化 细胞和IL-3非依赖于32D髓系细胞的生长。最后，具体目标4 将涉及到确定DH域的晶体结构 LARG与其GTP酶靶标RhoA的络合作用及对RhoA的测定 GTP酶的DH域识别的结构基础。尽管这一数字 DBL家族癌蛋白继续快速增加，到目前为止，LARG是 在人类癌症中发现的唯一突变的功能性DBL蛋白。我们的 研究将提供全面的、结构的、生化的和生物的 LARG功能分析及对LARG异常激活的作用 RhoA在AML开发中的应用。

DESCRIPTION: (Adapted from the investigator's abstract) The leukemia-associated Rho guanine nucleotide exchange factor (LARG) was recently identified as a fusion partner of the mixed lineage leukemia (MLL) protein in acute myeloid leukemia. LARG is a novel member of the rapidly expanding Dbl family of oncoproteins that promote malignant transformation by activating Ras-related Rho family GTPases. Like other Dbl family proteins, LARG contains a Dbl homology (DH) domain that functions as a guanine nucleotide exchange factor and activator of Rho GTPases. The DH domain is followed by a pleckstrin homology (PH) domain that presumably regulates DH domain function. LARG also contains a regulator of G-protein signaling (RGS) domain, suggesting that it may link G protein-coupled receptor signaling to Rho GTPases. Our preliminary studies determined that LARG is an activator of RhoA and can cause transformation of NIH 3T3 mouse fibroblasts. We have proposed four specific aims to perform detailed structure-function analyses of LARG. Specific aim 1 will determine the roles of the DH and PH domains in mediating LARG activation of RhoA. In particular, whether the PH domain regulates DH domain function in a phosphatidylinositol 3-kinase dependent fashion will be determined. Specific aim 2 will evaluate the role of the RGS domain in linking LARG with G protein coupled receptor signaling. This includes a determination of which heterotrimeric G alpha subunit(s) is regulated by the RGS domain and which G alpha subunit(s) in turn regulates LARG DH domain activation. Specific aim 3 will determine if the tumor-associated MLL-LARG fusion protein is an aberrantly activated form of LARG and can promote growth transformation of epithelial cells and lL-3 independent growth of 32D myeloid cells. Finally, Specific Aim 4 will involve a determination of the crystal structure of the DH/PH domains of LARG complexed with its GTPase target, RhoA, and the determination of the structural basis for DH domain recognition of GTPases. Although the number of Dbl family oncoproteins continue to increase at a rapid pace, to date, LARG is the only functional Dbl protein found to be mutated in human cancer. Our studies will provide a comprehensive, structural, biochemical, and biological analysis of LARG function and assess a role for aberrant LARG activation of RhoA in AML development.