The role of keratinocyte p53 in development of pigmented lesions

 描述(申请人提供)：皮肤恶性黑色素瘤是最致命的皮肤癌形式。在黑色素瘤的发生发展过程中，阳光下的紫外线辐射(UVR)既是BRAF等癌基因突变激活的始发者，又是黑素细胞迁移和增殖的促进剂。虽然导致晒伤的强烈紫外线照射与黑色素瘤风险有关，但流行病学表明，持续较低水平的长期暴露可能会预防黑色素瘤。这项建议集中在转录因子P53，一个基因组和细胞压力的传感器，以及它在UVR后角质形成细胞中的作用。UVR激活角质形成细胞中的P53，根据损伤的程度或类型，诱导角质形成细胞凋亡(晒伤细胞)，并增加黑素细胞生长因子的表达，如KIT配体(KITLG)，控制黑素细胞的增殖和功能；因此，P53可能很好地调节黑色素瘤的风险。事实上，在我们的角质形成细胞中p53水平持续高的煤油脚共济失调(SFA)小鼠中，我们观察到角质形成细胞来源的黑素细胞生长因子的表达增加，黑素细胞数量增加。重要的是，这些小鼠在接受致癌方案时会患上痣和黑色素瘤。这些数据强烈表明角质形成细胞P53具有促进肿瘤的作用。最近，我们在KITLG基因的p53反式激活位点上发现了一个关键的人类癌症风险SNP，它导致了UVR后KITLG表达的增加。综合这些数据，我们提出假设，角质形成细胞中强大的P53激活刺激了一个富含生长因子的环境，促进了色素病变向黑色素瘤的开始和发展；而持续的低P53激活则防止了黑色素瘤的发展。P53的这一新作用将在我们的结构性角质形成细胞P53过表达的SFA小鼠和我们的多西环素诱导的Tet-on角质形成细胞P53过表达的小鼠中进行测试。在目标1中，我们将研究角质形成细胞中P53的激活对旁分泌因子的释放和黑素细胞增殖的影响。在不同水平的P53存在的情况下，对P53途径活性(包括调节基因和靶基因)的详细分析将使我们能够构建角质形成细胞中P53作用的模型，该模型可能解释急性强烈UVR与黑色素瘤风险之间的明显联系，以及慢性低UVR与黑色素瘤保护之间的联系。在目标2中，我们将使用我们的Tet-on P53小鼠结合BRAFV600E黑色素瘤小鼠模型来研究角质形成细胞中不同水平的P53激活对黑色素瘤和黑色素瘤发生的旁分泌作用。我们将研究急性强烈的P53激活作为黑色素瘤的促进剂，以及低的慢性P53激活作为保护UVR诱导的黑色素瘤的介体。这一建议将使我们更好地了解角质形成细胞中的P53信号及其对黑素细胞和黑色素瘤的影响，最终可能导致黑色素瘤化学预防的新策略。本建议书中生成的数据将用于支持调查这些策略的R01应用程序。

 DESCRIPTION (provided by applicant): Cutaneous malignant melanoma is the deadliest form of skin cancer. Ultraviolet radiation (UVR) in sunlight plays a causative role in melanoma development both as an initiator, through mutational activation of oncogenes such as BRAF, and as a promoter through effects on melanocyte migration and proliferation. While intense UVR resulting in sunburns is associated with melanoma risk, epidemiology shows that persistent lower level chronic exposure may protect against melanoma. This proposal focuses on the transcription factor p53, a sensor of genomic and cellular stress, and its action in keratinocytes after UVR. UVR activates p53 in keratinocytes, and depending on the level or type of damage, induces keratinocyte apoptosis (sunburn cells), and increases the expression of melanocyte growth factors, like KIT Ligand (KITLG), that control melanocyte proliferation and function; therefore p53 may well modulate melanoma risk. Indeed, in our Sooty Foot Ataxia (SFA) mice with constitutively high p53 levels in keratinocytes, we observed increased expression of keratinocyte-derived melanocyte growth factors and elevated melanocyte numbers. Importantly, these mice develop nevi and melanomas when subjected to a carcinogenesis protocol. These data strongly suggest a tumor promotional role for keratinocyte p53. Recently, we identified a key human cancer risk SNP in the p53 transactivation site at the KITLG locus that resulted in increased KITLG expression post-UVR. Together these data led us to hypothesize that strong p53 activation in keratinocytes stimulates a growth factor-rich environment that promotes the initiation and progression of pigmented lesions to melanoma; whereas persistent low p53 activation protects against melanoma development. This new role for p53 will be tested using both our constitutive keratinocyte p53 overexpressing SFA mice and our Doxycycline-inducible Tet-ON keratinocyte p53 overexpressing mice. In Aim 1, we will examine the impact of p53 activation in keratinocytes on the release of paracrine factors and on melanocyte proliferation. A detailed analysis of p53 pathway activity, including regulators and target genes, in the presence of different levels of p53 will allow us to construct a model of p53 action in keratinocytes that may explain the apparent link between acute intense UVR and melanoma risk and the association of chronic low UVR with protection from melanoma. In Aim 2, we will use our Tet-ON p53 mice in combination with the BrafV600E melanoma mouse model to study the paracrine effects of different levels of p53 activation in keratinocytes on nevus and melanoma development. We will investigate both acute intense p53 activation as a melanoma promoter, and low chronic p53 activation as a mediator of protection against UVR-induced melanoma. This proposal will yield a greater understanding of p53 signaling in keratinocytes and its impact on melanocytes and melanoma that may ultimately lead to new strategies for melanoma chemoprevention. The data generated in this proposal will be used to support an R01 application investigating these strategies.