Roles of p63 in regulation of miRNA and LincRNA targets in metastatic cancer

描述（由申请人提供）：癌症死亡通常是由于远处转移的发生，而对此几乎没有有效的全身治疗方法。因此，体内转移调控相关基因的研究具有重要意义。 p63（p53 家族成员）的功能在 p53 功能已得到充分确立的背景下开始被理解，包括细胞凋亡和肿瘤抑制（Su 等人，Nature 2010）以及其他作用，如转移（Su 等人，Nature 2010）、皮肤发育和干细胞维持（Su 等人，Cell Stem Cell 2009）。 p63 功能的复杂性部分归因于以前无法独立研究的多种亚型的存在。 p63 的全长 TA 同工型包含一个反式激活结构域，在结构和功能上与 p53 相似，而 p63 的 ?N 同工型在也具有反式激活活性的同时，拮抗 p53 以及 p63 和 p73（另一个 p53 家族成员）的 TA 同工型的活性。有趣的是，存在于多种人类癌症中的突变 p53 已被证明与 p63 亚型相互作用并使其功能失活。 通过生成和研究具有 p63 异构体特异性无效等位基因的小鼠、TAp63-/- 和 ?Np63-/- 小鼠，我们发现这些基因在抑制肿瘤发生和转移中具有独特的作用。 TAp63-/- 小鼠对自发性转移性肿瘤高度敏感，并且 TAp63 通过 Dicer 和几种 miRNA 的直接协调转录调节来有效抑制肿瘤转移（Su 等人，Nature 2010）。此外，我们还将这些发现扩展到多种人类肿瘤类型，包括鳞状细胞癌、肺腺癌和乳腺癌，表明 TAp63 的这种功能非常重要。我们还表明，?Np63 可以抑制侵袭并转录调节 DGCR8，DGCR8 是 miRNA 加工所必需的另一种至关重要的酶。我们假设 TAp63 和 ?Np63 调节不同细胞区室中的不同 miRNA，以抑制肿瘤发生和转移并参与 DNA 损伤反应。我们将通过提出以下具体目标来解决我们的假设： 具体目标 1：了解癌起源细胞（表皮细胞）中 p63 亚型对 miRNA 和 LincRNA 靶标的机制调节。具体目标 2：了解皮肤癌和转移干细胞中 p63 亚型对 miRNA 和 LincRNA 的机制调节。具体目标 3：了解 p63 同工型调控的 miRNA 和 LincRNA 在 DNA 损伤反应和 p53 补偿中的作用。 公共健康相关性：考虑到转移性癌症的高死亡率，体内转移调控相关基因的研究非常重要，而 p63（p53 家族成员）通过协调 Dicer 和 microRNA (miRNA) 的转录调控，在抑制转移方面发挥着关键作用。该拨款提案的目标是确定在抑制转移和 DNA 损伤反应中受 p63 及其亚型调节的 miRNA 和长非编码 RNA (LincRNA) 靶标。我们将使用我的实验室生成的 p63 亚型特异性敲除转移性鳞状细胞癌小鼠模型和人类患者样本来实现这一目标。

DESCRIPTION (provided by applicant): Mortality in cancer is most often due to the development of distant metastases for which almost no effective systemic treatments exist. The study of genes relevant for the regulation of metastasis in vivo is therefore of great importance. The functions of p63, a p53 family member, are beginning to be understood in contexts in which p53 function has been well established, including apoptosis and tumor suppression (Su et al., Nature 2010) as well as in other roles such as metastasis (Su et al., Nature 2010), skin development, and stem cell maintenance (Su et al., Cell Stem Cell 2009). The complexity of p63 function is due in part to the existence of multiple isoforms that previously could not be studied independently. Full length TA isoforms of p63 contain a transactivation domain, structurally and functionally resembling p53, whereas the ?N isoforms of p63, while also possessing transactivation activity, antagonize the activities of p53 and the TA isoforms of p63 and p73, the other p53 family member. Interestingly, mutant p53, present in a wide variety of human cancers, has been shown to interact with p63 isoforms and inactivate their function. By generating and studying mice with isoform-specific null alleles of p63, the TAp63-/- and ?Np63-/- mice, we have shown that these genes have distinct roles in suppression of tumorigenesis and metastasis. TAp63-/- mice are highly susceptible to spontaneous metastatic tumors, and TAp63 potently suppresses tumor metastasis by direct coordinate transcriptional regulation of Dicer and several miRNAs (Su et al., Nature 2010). In addition, we have extended these findings to multiple human tumor types including squamous cell carcinoma, lung adenocarcinoma, and breast adenocarcinoma, indicating this function of TAp63 is widely important. We have also shown that ?Np63 can suppress invasion and transcriptionally regulates DGCR8, another critically important enzyme necessary for miRNA processing. We hypothesize that TAp63 and ?Np63 regulate distinct miRNAs in different cellular compartments to suppress tumorigenesis and metastasis and to engage the DNA damage response. We will address our hypothesis by proposing the following specific aims: Specific Aim 1: To understand the mechanistic regulation of miRNA and LincRNA targets by p63 isoforms in the cell of origin of carcinomas (epidermal cells). Specific Aim 2: To understand the mechanistic regulation of miRNAs and LincRNAs by p63 isoforms in stem cells of skin cancer and metastasis. Specific Aim 3: To understand the roles of miRNAs and LincRNAs regulated by p63 isoforms in the DNA damage response and p53 compensation. PUBLIC HEALTH RELEVANCE: The study of genes relevant for the regulation of metastasis in vivo is of great importance given the high mortality of metastatic cancer, and p63, a p53 family member, plays critical roles to suppress metastasis through coordinate transcriptional regulation of Dicer and microRNAs (miRNAs). The goals of this grant proposal are to identify the miRNA and long non-coding RNA (LincRNA) targets that are regulated by p63 and its isoforms in the suppression of metastasis and the DNA damage response. We will use p63 isoform specific knock out mouse models of metastatic squamous cell carcinoma generated in my laboratory and human patient samples to address this goal.