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CTCF genetic alterations in endometrial carcinoma are pro-tumorigenic.

ABSTRACT: CTCF is a haploinsufficient tumour suppressor gene with diverse normal functions in genome structure and gene regulation. However the mechanism by which CTCF haploinsufficiency contributes to cancer development is not well understood. CTCF is frequently mutated in endometrial cancer. Here we show that most CTCF mutations effectively result in CTCF haploinsufficiency through nonsense-mediated decay of mutant transcripts, or loss-of-function missense mutation. Conversely, we identified a recurrent CTCF mutation K365T, which alters a DNA binding residue, and acts as a gain-of-function mutation enhancing cell survival. CTCF genetic deletion occurs predominantly in poor prognosis serous subtype tumours, and this genetic deletion is associated with poor overall survival. In addition, we have shown that CTCF haploinsufficiency also occurs in poor prognosis endometrial clear cell carcinomas and has some association with endometrial cancer relapse and metastasis. Using shRNA targeting CTCF to recapitulate CTCF haploinsufficiency, we have identified a novel role for CTCF in the regulation of cellular polarity of endometrial glandular epithelium. Overall, we have identified two novel pro-tumorigenic roles (promoting cell survival and altering cell polarity) for genetic alterations of CTCF in endometrial cancer.

SUBMITTER: Marshall AD

PROVIDER: S-EPMC5519450 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

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CTCF genetic alterations in endometrial carcinoma are pro-tumorigenic.

Marshall A D AD Bailey C G CG Champ K K Vellozzi M M O'Young P P Metierre C C Feng Y Y Thoeng A A Richards A M AM Schmitz U U Biro M M Jayasinghe R R Ding L L Anderson L L Mardis E R ER Rasko J E J JEJ

Oncogene 20170320 29

CTCF is a haploinsufficient tumour suppressor gene with diverse normal functions in genome structure and gene regulation. However the mechanism by which CTCF haploinsufficiency contributes to cancer development is not well understood. CTCF is frequently mutated in endometrial cancer. Here we show that most CTCF mutations effectively result in CTCF haploinsufficiency through nonsense-mediated decay of mutant transcripts, or loss-of-function missense mutation. Conversely, we identified a recurrent ...[more]

PMID: 28319062

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Project description:BackgroundEndometrial cancer (EC) is a prevalent epithelial malignancy originating in the female endometrium, and its global incidence has been on the rise over the past decade. Despite significant scientific progress has been achieved recently, the genetic factors underlying EC pathogenesis remain poorly understood, warranting further investigation.MethodsWe employed transcriptomic datasets from the Genomic Data Commons database to extract variable and clinical data. Quantile normalization and log2 transformations were applied to obtain a gene expression matrix for the sample cohort. Various assays, such as quantitative real-time polymerase chain reaction (qRT-PCR), western blotting, immunohistochemistry (IHC), wound healing assay, transwell assay, and TUNEL assay, were employed in the study to examine the involvement of CTCF in EC cell biology. Additionally, in vivo experiments were conducted using a subcutaneous transplantation tumor model in athymic nude mice. The potential mechanism of action of CTCF was also illustrated by identifying differentially expressed genes (DEGs) and functions after interfering with CTCF gene expression through the GSPAdb online database.ResultsAfter categorizing 543 samples into cohorts with high and low ImmuneScores, we discovered 1025 genes that were differentially expressed, including 745 genes that were up-regulated and 280 genes that were down-regulated in the high scores group compared to the low scores group. Tumor mutation burden (TMB) analysis highlighted 11 genes with the highest mutation frequency. Furthermore, 16 immune checkpoints and 50 immune regulatory factors exhibited differential expression. Among these, CTCF was up-regulated in EC. We found that CTCF knockdown could diminish EC's invasive ability and metastatic potential while enhancing apoptosis. In vivo experiments corroborated that CTCF knockdown could reduce tumor growth. The GSPAdb online database identified differential expression pathways mainly enriched in cellular metabolism as well as some intracellular classical signaling pathways after interfering with CTCF gene expression. In addition, we identified potential downstream regulators of CTCF through protein interaction networks.ConclusionThis study unveiled comprehensive molecular characteristics and DEGs in EC, emphasizing the up-regulation of CTCF in EC. Our findings collectively suggest that CTCF represents a promising therapeutic target, and our gene molecular typing model offers a novel approach for prognostic evaluation in EC.

Project description:The current study tested the expression and potential functions of Gαi1 in nasopharyngeal carcinoma (NPC). The Cancer Genome Atlas (TCGA) database results demonstrate that Gαi1 transcripts' number in NPC tissues is significantly higher than that in the normal nasal epithelial tissues. Its overexpression correlates with poor survival in certain NPC patients. Moreover, Gαi1 is significantly upregulated in NPC tissues of local primary patients and in different primary human NPC cells. Whereas its expression is relatively low in cancer-surrounding normal tissues and in primary nasal epithelial cells. Genetic silencing (via shRNA strategy) or knockout (via CRISPR-sgRNA method) of Gαi1 substantially suppressed viability, proliferation, cell cycle progression, and migration in primary NPC cells, causing significant caspase-apoptosis activation. Contrarily, ectopic Gαi1 expression exerted pro-tumorigenic activity and strengthened cell proliferation and migration in primary NPC cells. Gαi1 is important for Akt-mTOR activation in NPC cells. Akt-S6K phosphorylation was downregulated after Gαi1 shRNA or KO in primary NPC cells, but strengthened following Gαi1 overexpression. In Gαi1-silenced primary NPC cells, a S473D constitutively-active mutant Akt1 (caAkt1) restored Akt-S6K phosphorylation and ameliorated Gαi1 shRNA-induced proliferation inhibition, migration reduction and apoptosis. Bioinformatics analyses proposed zinc finger protein 384 (ZNF384) as a potential transcription factor of Gαi1. In primary NPC cells, ZNF384 shRNA or knockout (via CRISPR-sgRNA method) decreased Gαi1 mRNA and protein expression, whereas ZNF384 overexpression upregulated it. Importantly, there was an increased binding between ZNF384 protein and the Gαi1 promoter in human NPC tissues and different NPC cells. In vivo studies showed that intratumoral injection of Gαi1-shRNA-expressing adeno-associated virus (AAV) impeded subcutaneous NPC xenograft growth in nude mice. Gαi1 downregulation, Akt-mTOR inactivation, and apoptosis induction were detected in Gαi1-silenced NPC xenograft tissues. Gαi1 KO also effectively inhibited the growth of NPC xenografts in nude mice. Together, overexpressed Gαi1 exerts pro-tumorigenic activity in NPC possibly by promoting Akt-mTOR activation.

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CTCF genetic alterations in endometrial carcinoma are pro-tumorigenic.

Publications

CTCF genetic alterations in endometrial carcinoma are pro-tumorigenic.

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