p53 acetylation in ferroptosis and tumor suppression

项目摘要 P53肿瘤抑制通路的失活是大多数人类肿瘤形成的关键事件 癌症。尽管P53介导的细胞周期停滞、衰老和凋亡是影响细胞周期的关键因素 癌症的发展，越来越多的证据表明，P53依赖的细胞周期停滞，细胞凋亡， 衰老不足以消除P53的抑瘤活性。几种小鼠模型 提示在缺乏这些典型功能的情况下，P53抑制肿瘤的作用是可以实现的。P53 蛋白质通过作为DNA结合转录因子选择性地实现不同的细胞结果 调节某些P53转录靶基因的表达。大量的研究已经证实 P53乙酰化是启动子特异性激活P53靶基因表达的关键信号。值得注意的是， 我们早期的研究表明p533KR(3KR：K117R+K161R+K162R)乙酰化缺陷突变体 小鼠模型缺乏经历P53介导的细胞凋亡、衰老和细胞周期停滞的能力， 与p53基因缺失的小鼠相比，它们不太容易患上肿瘤。铁性下垂是一种规律性的 非凋亡性细胞死亡的一种形式，由过氧化脂质的积聚驱动。我们发现P53抑制了 通过抑制SLC7A11的表达来摄取胱氨酸并使细胞对铁性下垂敏感 胱氨酸/谷氨酸逆向转运蛋白。事实上，p533KR，一个乙酰化缺陷的突变体，未能诱导细胞周期 抑制衰老和细胞凋亡，完全保留了调节SLC7A11表达和促进 铁性下垂。然而，目前尚不清楚P53介导的铁性下垂是如何调节的，以及P53是否- 介导性下睑下垂是其残留的肿瘤抑制所必需的。在我们的初步研究中，我们 已经在小鼠P53的赖氨酸K98(或人P53的K101)上发现了一个新的P53乙酰化位点。虽然损失很大 K98乙酰化(P53K98R)单独对p53介导的反式激活的影响很小，同时 所有四个乙酰化位点(p534KR：K98R+K117R+K161R+K162R)的突变完全丧失了它的能力 调节TIGAR、SLC7A11等代谢靶点。这里要检验的中心假设是 P53乙酰化在调节P53功能中起主要作用以及P53介导的铁性下垂是否起作用 在没有细胞周期停滞、细胞凋亡和衰老的情况下抑制肿瘤的关键机制。在AIM 1，我们将阐明乙酰化在调节P53介导的铁上链反应中的分子机制。 以及人类癌细胞中的肿瘤生长。在目标2中，我们将研究p53的生理意义。 建立新的p53突变(p534KR/4KR)小鼠体内乙酰化及P53介导的铁下垂 模特。

Project Summary Inactivation of the p53 tumor suppression pathway is a pivotal event in the formation of most human cancers. Although p53-mediated cell-cycle arrest, senescence and apoptosis serve as critical barriers to cancer development, accumulating evidence suggests that loss of p53-dependent cell cycle arrest, apoptosis, and senescence is not sufficient to abrogate the tumor suppression activity of p53. Several mouse models suggest that tumor suppression by p53 can be achieved in the absence of those canonical functions. The p53 protein achieves diverse cellular outcomes by serving as a DNA-binding transcription factor that selectively modulates the expression of certain p53 transcriptional target genes. Numerous studies have established that acetylation of p53 acts as a key signal in promoter-specific activation of p53 target gene expression. Notably, our earlier studies demonstrate that the p533KR (3KR:K117R+K161R+K162R) acetylation-deficient mutant mouse model, which lacks the ability to undergo p53-mediated apoptosis, senescence and cell cycle arrest, are not significantly prone to developing tumors when compared to p53-null mice. Ferroptosis is a regulated form of non-apoptotic cell death driven by accumulation of lipid hydroperoxides. We found that p53 inhibits cystine uptake and sensitizes cells to ferroptosis by repressing expression of SLC7A11, a key component of the cystine/glutamate antiporter. Indeed, p533KR, an acetylation-defective mutant that fails to induce cell-cycle arrest, senescence and apoptosis, fully retains the ability to regulate SLC7A11 expression and promote ferroptosis. Nevertheless, it remains unclear how p53-mediated ferroptosis is regulated and whether p53- mediated ferroptosis is absolutely required for its remaining tumor suppression. In our preliminary studies, we have identified a novel p53 acetylation site at lysine K98 in mouse p53 (or K101 for human p53). While the loss of K98 acetylation (p53K98R) alone has very modest effects on p53-mediated transactivation, simultaneous mutations at all four acetylation sites (p534KR: K98R+K117R+K161R+K162R) completely abolish its ability to regulate metabolic targets such as TIGAR and SLC7A11. The central hypothesis to be tested here is whether p53 acetylation plays a major role in modulating p53 functions and whether p53-mediated ferroptosis is acts as a key mechanism in tumor suppression in the absence of cell-cycle arrest, apoptosis and senescence. In Aim 1, we will elucidate the molecular mechanisms of acetylation in modulating p53-mediated ferroptotic responses and tumor growth in human cancer cells. In Aim 2, we will examine the physiological significance of p53 acetylation as well as p53-mediated ferroptosis in vivo by establishing a new p53-mutant (p534KR/4KR) mouse model.