Identification of a noval oncogene using validation-based insertional mutagenesis

描述（由申请人提供）：使用人乳腺上皮细胞（hmec），我们进行了有助于恶性转化的改变的正向遗传筛选，并鉴定了一种新的致癌基因，命名为FAM83B。FAM83B的n端编码一个PLD基序，这对于FAM83B介导的转化是必要和充分的。PLD活性的升高通过增加磷脂酸（PA）的产生来促进肿瘤的发生，磷脂酸是一种有效的第二信使，参与调节mTOR和MAPK信号，这两种信号在表达fam83b的细胞中都被显著激活。从EGFR或HER2升高的肿瘤来源细胞，或从活化RAS转化的HMECs中消融FAM83B，抑制其增殖，锚定不依赖生长和致瘤性，支持FAM83B作为关键的EGFR/RAS-信号介质的作用。通过对人类肿瘤标本中FAM83B表达的分析，发现其在多种人类癌症中过表达，我们的研究结果表明FAM83B是一种参与上调MAPK和mTOR信号的新型癌蛋白。本提案的目标是确定FAM83B在MAPK和mTOR信号传导中的参与，并检查人类癌症标本，以将FAM83B的表达与这些高度重要的信号级联的激活联系起来。我们的假设是fam83b介导的PA的产生导致RAF-MEK-ERK信号的激活和pi3k - akt独立的mTOR信号的激活，从而驱动乳腺癌的发展。上述研究的成功完成将为FAM83B在癌细胞中的抑制是否会同时消融MAPK和mTOR信号提供重要信息，验证FAM83B作为治疗干预的未来靶点。本提案的目的是：(1)确定FAM83B参与EGFR/ ras介导的转化和效应物激活。(2)确定FAM83B在mTOR激活中的作用，独立于典型的PI3K-AKT信号轴。(3)确定FAM83B表达的升高是否为癌症中MAPK和mTOR信号的过度激活提供了另一种机制。

DESCRIPTION (provided by applicant): Using human mammary epithelial cells (HMECs), we performed a forward genetic screen for alterations that contribute to malignant transformation and identified a novel oncogene, named FAM83B. The N-terminus of FAM83B encodes a PLD motif, which is both necessary and sufficient for FAM83B-mediated transformation. Elevated PLD activity contributes to tumorigenesis through the increased production of phosphatidic acid (PA), a potent second messenger involved in regulating mTOR and MAPK signaling, both of which are significantly activated in FAM83B-expressing cells. Ablation of FAM83B from tumor-derived cells with elevated EGFR or HER2, or from HMECs transformed by activated RAS, inhibited their proliferation, anchorage-independent growth and tumorigenicity, supporting a role for FAM83B as a critical EGFR/RAS- signaling intermediary. Together with the analysis of FAM83B expression in human tumor specimens, which revealed its overexpression in a variety of human cancers, our results suggest that FAM83B represents a novel oncoprotein involved in upregulating MAPK and mTOR signaling. The goals of this proposal are to define the involvement of FAM83B in MAPK and mTOR signaling, and examine human cancer specimens to correlate FAM83B expression with the activation of these highly important signaling cascades. Our hypothesis is that the FAM83B-mediated production of PA results in the activation of RAF-MEK-ERK signaling and the PI3K-AKT-independent activation of mTOR signaling, which drives breast cancer development. Successful completion of the proposed studies will provide important information regarding whether FAM83B inhibition in cancer cells will simultaneously ablate MAPK and mTOR signaling, validating FAM83B as a future target for therapeutic intervention. The aims of this proposal are to (1) Determine the involvement of FAM83B in EGFR/RAS-mediated transformation and effector activation. (2) Define the role of FAM83B in mTOR activation, independently of the canonical PI3K-AKT signaling axis. (3) Determine whether elevation of FAM83B expression provides an alternative mechanism for hyperactivating MAPK and mTOR signaling in cancer. PUBLIC HEALTH RELEVANCE: Successful identification of novel proteins involved in HMEC transformation, such as FAM83B, is critical to expanding our understanding of the oncogene signaling interactions involved in driving breast epithelial hyperplasia. Novel proteins also provide opportunities for drug development that will expand the arsenal of therapies necessary for eradicating cancer and extending the life of patients.