Alternative polyadenylation reprogramming of MORC2 induced by NUDT21 loss promotes KIRC carcinogenesis
Yuqin Tan, … , Ke Li, Ning Na
Yuqin Tan, … , Ke Li, Ning Na
Published September 22, 2023
Citation Information: JCI Insight. 2023;8(18):e162893. https://doi.org/10.1172/jci.insight.162893.
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Research Article Oncology

Alternative polyadenylation reprogramming of MORC2 induced by NUDT21 loss promotes KIRC carcinogenesis

Abstract

Alternative polyadenylation (APA), a posttranscriptional mechanism of gene expression via determination of 3′UTR length, has an emerging role in carcinogenesis. Although abundant APA reprogramming is found in kidney renal clear cell carcinoma (KIRC), which is one of the major malignancies, whether APA functions in KIRC remains unknown. Herein, we found that chromatin modifier MORC2 gained oncogenic potential in KIRC among the genes with APA reprogramming, and moreover, its oncogenic potential was enhanced by 3′UTR shortening through stabilization of MORC2 mRNA. MORC2 was found to function in KIRC by downregulating tumor suppressor DAPK1 via DNA methylation. Mechanistically, MORC2 recruited DNMT3A to facilitate hypermethylation of the DAPK1 promoter, which was strengthened by 3′UTR shortening of MORC2. Furthermore, loss of APA regulator NUDT21, which was induced by DNMT3B-mediated promoter methylation, was identified as responsible for 3′UTR shortening of MORC2 in KIRC. Additionally, NUDT21 was confirmed to act as a tumor suppressor mainly depending on downregulation of MORC2. Finally, we designed an antisense oligonucleotide (ASO) to enhance NUDT21 expression and validated its antitumor effect in vivo and in vitro. This study uncovers the DNMT3B/NUDT21/APA/MORC2/DAPK1 regulatory axis in KIRC, disclosing the role of APA in KIRC and the crosstalk between DNA methylation and APA.

Authors

Yuqin Tan, Tong Zheng, Zijun Su, Min Chen, Suxiang Chen, Rui Zhang, Ruojiao Wang, Ke Li, Ning Na

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Figure 6

3′UTR shortening of MORC2 amplifies promoter methylation and downregulation of DAPK1 via enhancing DNMT3A recruitment.

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3′UTR shortening of MORC2 amplifies promoter methylation and downregulat...
(A and B) qPCR (A) and immunoblotting (B) were performed to evaluate DAPK1 expression in KIRC cells transfected with Flag vector or Flag-MORC2 with silencing of indicated gene respectively (n = 3). (C and D) qPCR (C) and immunoblotting (D) were performed to evaluate DAPK1 expression in KIRC cells transfected with Flag vector or Flag-DNMT3A (n = 3). (E and F) qPCR (E) and immunoblotting (F) were performed to evaluate DAPK1 expression in KIRC cells transfected with control or DNMT3A-specific siRNA (n = 3). (G) MeDIP-qPCR was performed to evaluate promoter methylation level of DAPK1 in WT and MORC2-depleted Caki-1 cells transfected with Flag vector or Flag-DNMT3A (n = 3). (H) Co-IP was performed to detect the interaction between MORC2 and DNMT3A in KIRC cells. (I) ChIP-qPCR was performed to determine the abundance of DNMT3A at the DAPK1 promoter in Caki-1 cells transfected with control or MORC2-specific siRNA (n = 3). (J and K) qPCR (J) and immunoblotting (K) were performed to evaluate DAPK1 expression in KIRC cells transfected with vector, short 3′UTR, or long 3′UTR MORC2 plasmid (n = 3). (L and M) qPCR (L) and immunoblotting (M) were performed to evaluate DAPK1 expression in WT and S-KO Caki-1 cells (n = 3). (N) MeDIP-qPCR was performed to evaluate promoter methylation level of DAPK1 in WT and S-KO Caki-1 cells (n = 3). (O) ChIP-qPCR was performed to determine the abundance of DNMT3A at the DAPK1 promoter in WT and S-KO Caki-1 cells (n = 3). All data represent the mean ± SD. Two-tailed t test, 1-way ANOVA with Tukey multiple-comparison test, or 2-way ANOVA with Tukey multiple-comparison test analyses were performed. *P < 0.05; **P < 0.01; ***P < 0.001.