PO.MCB07.01 · 分子与细胞生物学
CREBBP在EP300改变的膀胱癌中通过异常的IL-1alpha信号传导驱动致瘤性
CREBBP drives tumorigenicity via aberrant IL-1alpha signaling in EP300 altered bladder cancer
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摘要 Abstract
中文摘要
EP300和CREBBP分别编码p300和CBP，二者均为H3K27乙酰化酶。EP300和CREBBP均在一部分尿路上皮癌中发生突变（EP300或CREBBP各约15%）。EP300和CREBBP基因具有约60%的序列相似性，因此一直被推测在细胞稳态和癌症发病机制中具有很大程度上重叠的功能作用。为了表征EP300突变在尿路上皮癌发病机制中的作用并鉴定这两个旁系同源基因的非冗余作用，我们构建了EP300和CREBBP同基因（isogenic）敲除的尿路上皮癌细胞系，并表征了EP300和CREBBP突变型及野生型的低传代尿路上皮癌来源患者来源类器官。EP300敲除（KO）和功能缺失突变与软琼脂中细胞生长增强、体外侵袭潜能增加以及细胞代谢改变相关。这些功能获得性表型由IL-6反式信号传导（trans-signaling）所致的JAK-STAT3激活增强所介导，其近端驱动因素是IL-1á转录和产生的增加。值得注意的是，CREBBP敲除的同基因BLCA细胞并未导致IL-1á过度分泌或IL-6/JAK1/STAT3信号轴的过度激活，表明这一现象特异于EP300功能缺失。对亲本、EP300 KO和CREBBP KO的RT112克隆进行的转录组分析显示，CREBBP KO显著降低了IL1A转录本水平，而EP300 KO则显著升高了该水平。此外，与亲本和EP300缺失细胞系相比，CREBBP失活的BLCA细胞系在基因毒性应激后无法上调IL1A表达。使用靶向CREBBP的诱导型短发夹RNA构建体，我们还发现CREBBP敲低可挽救EP300 KO克隆中IL1A的上调，同时伴随显著的生长缺陷。通过p300/CBP特异性抑制剂A485进行药理学抑制也消除了IL1A的上调，并在EP300 KO细胞中引起显著的生长缺陷。EP300缺失克隆对A485治疗的敏感性也显著高于CREBBP缺失的尿路上皮细胞或亲本细胞系。总之，我们的结果确定了IL-1á-JAK-STAT3信号的调控是EP300和CREBBP之间一种新的非冗余关系，可在具有EP300功能缺失突变的患者中加以治疗性利用。
查看英文原文 English abstract
EP300 and CREBBP encode p300 and CBP respectively, which are H3K27 acetylases. Both EP300 and CREBBP mutated in a subset of urothelial cancers (~15% for EP300 or CREBBP ). The EP300 and CREBBP genes have ~60% sequence similarity and thus have been presumed to have largely overlapping functional roles in cell homeostasis and cancer pathogenesis. To characterize the role of EP300 mutations in urothelial cancer pathogenesis and to identify non-redundant roles of these paralogues, we generated EP300 and CREBBP isogenic knockout urothelial cancer cell lines and characterized low passage mutant and wild type for EP300 and CREBBP urothelial cancer-derived patient derived organoids. EP300 KO and loss-of-function mutation was associated with enhanced cell growth in soft agar, increased invasive potential in vitro and altered cellular metabolism. These gain-of-function phenotypes were mediated by enhanced JAK-STAT3 activation resulting from IL-6 trans-signaling, the proximal driver of which was increased transcription and production of IL-1á. Notably, isogenic BLCA cells with CREBBP knockout did not confer IL-1á hypersecretion or hyperactivation of the IL-6/JAK1/STAT3 signaling axis indicating that this is a phenomenon was specific to EP300 loss-of-function. Transcriptomic analysis of Parental, EP300 KO, and CREBBP KO RT112 clones revealed that CREBBP KO significantly depressed IL1A transcript levels which EP300 KO significantly elevated. Additionally, CREBBP inactivated BLCA lines could not upregulate IL1A expression following genotoxic stress in contrast to parental and EP300 null cell lines. Using an inducible short hairpin RNA construct targeting CREBBP , we also find that CREBBP knockdown rescued IL1A upregulation in EP300 ko clones that coincided with a significant growth defect. Pharmacologic inhibition via the p300/CBP specific inhibitor A485 also abolished IL1A upregulation and caused significant growth defect in EP300 KO cells. EP300 null clones were also significantly more sensitive to A485 treatment than either CREBBP null urothelial cell or parental cell lines. In sum, our results identify regulation of IL-1á-JAK-STAT3 signaling as a novel non-redundancy between EP300 and CREBBP that could be exploited therapeutically in patient with EP300 loss-of-function mutations.
利益披露 Disclosure
J. A. Rodrigues, None..
D. Li, None..
D. Schoeps, None..
S. Shukla, None..
Z. Jacobs, None..
W. Cho, None.