bims-rimeca 2022-06-12 papers

Clin Transl Med. 2022 Jun;12(6): e906

ZNF677 suppresses renal cell carcinoma progression through N6-methyladenosine and transcriptional repression of CDKN3.

Aolin Li, Congcong Cao, Ying Gan, Xiaofei Wang, Tianyu Wu, Quan Zhang, Yuchen Liu, Lin Yao, Qian Zhang.

BACKGROUND: Studies on biological functions of N6-methyladenosine (m6 A) modification in mRNA have sprung up in recent years. Previous studies have reported m6 A can determine mRNA fate and play a pivotal role in tumour development and progression. The zinc finger protein 677 (ZNF677) belongs to the zinc finger protein family and possesses transcription factor activity by binding sequence-specific DNA.
METHODS: The expression of ZNF677 and its clinicopathological impact were evaluated in renal cell carcinoma (RCC) patients. The m6 A level of ZNF677 was determined by m6 A methylated RNA immunoprecipitation-sequencing (MeRIP-seq) and MeRIP-qPCR in RCC tissues and adjacent normal tissues. RNA immunoprecipitation-qPCR (RIP-qPCR) and luciferase assays were performed to identify the targeted effect of IGF2BP2 and YTHDF1 on ZNF677. RCC cells and subcutaneous models uncovered the role of ZNF677 methylated by CRISPR/dCas13b-METTL3 in tumour growth. ZNF677-binding sites in the CDKN3 promoter were investigated by chromatin immunoprecipitation (ChIP) and luciferase assays.
RESULTS: ZNF677 is frequently downregulated in RCC tissues and its low expression is associated with unfavourable prognosis and decreased m6 A modification level. Further, we find the m6 A-modified coding sequence (CDS) of ZNF677 positively regulates its translation and mRNA stability via binding with YTHDF1 and IGF2BP2, respectively. Targeted specific methylation of ZNF677 m6 A by CRISPR/dCas13b-METLL3 system can significantly increase the m6 A and expression level of ZNF677, and dramatically inhibit cell proliferation and induce cell apoptosis of RCC cells. In addition, ZNF677 exerted its tumour suppressor functions in RCC cells through transcriptional repression of CDKN3 via binding to its promoter. In vitro and clinical data confirm the negative roles of ZNF677/CDKN3 in tumour growth and progression of RCC.
CONCLUSION: ZNF677 functions as a tumour suppressor and is frequently silenced via m6 A modification in RCC, which may highlight m6 A methylation-based approach for RCC diagnosis and therapy.

Keywords: CDKN3; RCC; ZNF677; dCas13b; m6A

DOI: https://doi.org/10.1002/ctm2.906

J Clin Lab Anal. 2022 Jun 11. e24514

RNA demethylase ALKBH5 regulates hypopharyngeal squamous cell carcinoma ferroptosis by posttranscriptionally activating NFE2L2/NRF2 in an m6 A-IGF2BP2-dependent manner.

Jing Ye, Xiaozhen Chen, Xiaohua Jiang, Zhihuai Dong, Sunhong Hu, Mang Xiao.

BACKGROUND: Having emerged as the most abundant posttranscriptional internal mRNA modification in eukaryotes, N6-methyladenosine (m6 A) has attracted tremendous scientific interest in recent years. However, the functional importance of the m6 A methylation machinery in ferroptosis regulation in hypopharyngeal squamous cell carcinoma (HPSCC) remains unclear.
METHODS: We herein performed bioinformatic analysis, cell biological analyses, transcriptome-wide m6 A sequencing (m6 A-seq, MeRIP-seq), RNA sequencing (RNA-seq), and RNA immunoprecipitation sequencing (RIP-seq), followed by m6 A dot blot, MeRIP-qPCR, RIP-qPCR, and dual-luciferase reporter assays.
RESULTS: The results revealed that ALKBH5-mediated m6 A demethylation led to the posttranscriptional inhibition of NFE2L2/NRF2, which is crucial for the regulation of antioxidant molecules in cells, at two m6 A residues in the 3'-UTR. Knocking down ALKBH5 subsequently increased the expression of NFE2L2/NRF2 and increased the resistance of HPSCC cells to ferroptosis. In addition, m6 A-mediated NFE2L2/NRF2 stabilization was dependent on the m6 A reader IGF2BP2. We suggest that ALKBH5 dysregulates NFE2L2/NRF2 expression in HPSCC through an m6 A-IGF2BP2-dependent mechanism.
CONCLUSION: Together, these results have revealed an association between the ALKBH5-NFE2L2/NRF2 axis and ferroptosis, providing insight into the functional importance of reversible mRNA m6 A methylation and its modulators in HPSCC.

Keywords: ALKBH5; HPSCC; NFE2L2/NRF2; ferroptosis; m6A modification

DOI: https://doi.org/10.1002/jcla.24514

Dev Cell. 2022 May 26. pii: S1534-5807(22)00349-5. [Epub ahead of print]

PDGF signaling inhibits mitophagy in glioblastoma stem cells through N6-methyladenosine.

Deguan Lv, Ryan C Gimple, Cuiqing Zhong, Qiulian Wu, Kailin Yang, Briana C Prager, Bhaskar Godugu, Zhixin Qiu, Linjie Zhao, Guoxin Zhang, Deobrat Dixit, Derrick Lee, Jia Z Shen, Xiqing Li, Qi Xie, Xiuxing Wang, Sameer Agnihotri, Jeremy N Rich.

Dysregulated growth factor receptor pathways, RNA modifications, and metabolism each promote tumor heterogeneity. Here, we demonstrate that platelet-derived growth factor (PDGF) signaling induces N6-methyladenosine (m6A) accumulation in glioblastoma (GBM) stem cells (GSCs) to regulate mitophagy. PDGF ligands stimulate early growth response 1 (EGR1) transcription to induce methyltransferase-like 3 (METTL3) to promote GSC proliferation and self-renewal. Targeting the PDGF-METTL3 axis inhibits mitophagy by regulating m6A modification of optineurin (OPTN). Forced OPTN expression phenocopies PDGF inhibition, and OPTN levels portend longer survival of GBM patients; these results suggest a tumor-suppressive role for OPTN. Pharmacologic targeting of METTL3 augments anti-tumor efficacy of PDGF receptor (PDGFR) and mitophagy inhibitors in vitro and in vivo. Collectively, we define PDGF signaling as an upstream regulator of oncogenic m6A regulation, driving tumor metabolism to promote cancer stem cell maintenance, highlighting PDGF-METTL3-OPTN signaling as a GBM therapeutic target.

Keywords: (m(6)A); METTL3; N6-methyladenosine; OPTN; PDGF; PDGFRβ; cancer stem cell; glioblastoma; glioblastoma stem cell; mitophagy; optineurin

DOI: https://doi.org/10.1016/j.devcel.2022.05.007

Nat Commun. 2022 Jun 07. 13(1): 3257

Structural basis for MTA1c-mediated DNA N6-adenine methylation.

Jiyun Chen, Rong Hu, Ying Chen, Xiaofeng Lin, Wenwen Xiang, Hong Chen, Canglin Yao, Liang Liu.

DNA N6-adenine methylation (6 mA) has recently been found to play a crucial role in epigenetic regulation in eukaryotes. MTA1c, a newly discovered 6 mA methyltransferase complex in ciliates, is composed of MTA1, MTA9, p1 and p2 subunits and specifically methylates ApT dinucleotides, yet its mechanism of action remains unknown. Here, we report the structures of Tetrahymena thermophila MTA1 (TthMTA1), Paramecium tetraurelia MTA9 (PteMTA9)-TthMTA1 binary complex, as well as the structures of TthMTA1-p1-p2 and TthMTA1-p2 complexes in apo, S-adenosyl methionine-bound and S-adenosyl homocysteine-bound states. We show that MTA1 is the catalytically active subunit, p1 and p2 are involved in the formation of substrate DNA-binding channel, and MTA9 plays a structural role in the stabilization of substrate binding. We identify that MTA1 is a cofactor-dependent catalytically active subunit, which exhibits stable SAM-binding activity only after assembly with p2. Our structures and corresponding functional studies provide a more detailed mechanistic understanding of 6 mA methylation.

DOI: https://doi.org/10.1038/s41467-022-31060-6

Mol Cancer Res. 2022 Jun 08. pii: molcanres.0122.2022-2-15 13:39:56.450. [Epub ahead of print]

Ecdysoneless overexpression drives mammary tumorigenesis through upregulation of c-MYC and glucose metabolism.

Ecdysoneless (ECD) protein is essential for embryogenesis, cell cycle progression and cellular stress mitigation with an emerging role in mRNA biogenesis. We have previously shown that ECD protein as well as its mRNA are overexpressed in breast cancer (BC) and ECD overexpression predicts shorter survival in BC patients. However, the genetic evidence for an oncogenic role of ECD has not been established. Here, we generated transgenic mice with mammary epithelium-targeted overexpression of an inducible human ECD transgene (ECDTg) mouse. Significantly, ECDTg mice develop mammary hyperplasia, pre-neoplastic lesions, and heterogeneous tumors with occasional lung metastasis. ECDTg tumors exhibit epithelial to mesenchymal transition and cancer stem cells characteristics. Organoid cultures of ECDTg tumors showed ECD dependency for in vitro oncogenic phenotype and in vivo growth when implanted in mice. RNA-Seq analysis of ECDTg tumors showed a c-MYC signature, and alterations in ECD levels regulated c-MYC mRNA and protein levels as well as glucose metabolism. ECD knockdown-induced decrease in glucose uptake was rescued by overexpression of mouse ECD as well as c-MYC. Publicly available expression data analyses showed a significant correlation of ECD and c-MYC overexpression in BC, and ECD and c-MYC co-expression exhibits worse survival in BC patients. Taken together, we establish a novel role of overexpressed ECD as an oncogenesis driver in the mouse mammary gland through upregulation of c-MYC-mediated glucose metabolism. Implications: We demonstrate ECD overexpression in the mammary gland of mice led to the development of a tumor progression model through upregulation of c-MYC signaling and glucose metabolism.

DOI: https://doi.org/10.1158/1541-7786.MCR-22-0122