Cap-Specific MAm Methyltransferase PCIF1/CAPAM Regulates MRNA Stability of and Through the MA Methyltransferase Activity

Overview

Journal Cells

Publisher MDPI

Specialties Biochemistry
Biophysics
Cell Biology
Molecular Biology

Date 2024 Oct 25

PMID 39451207

Authors

Ai Sugita

Ryoya Kano

Hiroyasu Ishiguro

Natsuki Yanagisawa

Soichiro Kuruma

Shotaro Wani

Aki Tanaka

Yoshiaki Tabuchi

Yoshiaki Ohkuma

Yutaka Hirose

Affiliations

Soon will be listed here.

Abstract

Chemical modifications of cellular RNAs play key roles in gene expression and host defense. The cap-adjacent ,2'--dimethyladenosine (mAm) is a prevalent modification of vertebrate and viral mRNAs and is catalyzed by the newly discovered methyltransferase PCIF1. However, its role in gene expression remains unclear due to conflicting reports on its effects on mRNA stability and translation. In this study, we investigated the impact of siRNA-mediated transient suppression of PCIF1 on global mRNA expression in HeLa cells. We identified a subset of differentially expressed genes (DEGs) that exhibited minimal overlap with previously reported DEGs. Subsequent validation revealed that PCIF1 positively and negatively regulates RAB23 and CNOT6 expression, respectively, at both the mRNA and protein levels. Mechanistic analyses demonstrated that PCIF1 regulates the stability of these target mRNAs rather than their transcription, and rescue experiments confirmed the requirement of PCIF1's methyltransferase activity for these regulations. Furthermore, MeRIP-qPCR analysis showed that PCIF1 suppression significantly reduced the mA levels of and mRNAs. These findings suggest that PCIF1 regulates the stability of specific mRNAs in opposite ways through mA modification, providing new insights into the role of mAm in the regulation of gene expression.

References

Song P, Liu S, Liu D, Keijzers G, Bakula D, Duan S . CNOT6: A Novel Regulator of DNA Mismatch Repair. Cells. 2022; 11(3). PMC: 8833972. DOI: 10.3390/cells11030521. View

Zheng L, Chi S, Li C . Rab23's genetic structure, function and related diseases: a review. Biosci Rep. 2017; 37(2). PMC: 5333778. DOI: 10.1042/BSR20160410. View

Borbolis F, Syntichaki P . Biological implications of decapping: beyond bulk mRNA decay. FEBS J. 2021; 289(6):1457-1475. DOI: 10.1111/febs.15798. View

Jenkins D, Seelow D, Jehee F, Perlyn C, Alonso L, Bueno D . RAB23 mutations in Carpenter syndrome imply an unexpected role for hedgehog signaling in cranial-suture development and obesity. Am J Hum Genet. 2007; 80(6):1162-70. PMC: 1867103. DOI: 10.1086/518047. View

Sendinc E, Valle-Garcia D, Dhall A, Chen H, Henriques T, Navarrete-Perea J . PCIF1 Catalyzes m6Am mRNA Methylation to Regulate Gene Expression. Mol Cell. 2019; 75(3):620-630.e9. PMC: 6688901. DOI: 10.1016/j.molcel.2019.05.030. View

Mauer J, Luo X, Blanjoie A, Jiao X, Grozhik A, Patil D . Reversible methylation of mA in the 5' cap controls mRNA stability. Nature. 2016; 541(7637):371-375. PMC: 5513158. DOI: 10.1038/nature21022. View

Zhang Q, Kang Y, Wang S, Gonzalez G, Li W, Hui H . HIV reprograms host mAm RNA methylome by viral Vpr protein-mediated degradation of PCIF1. Nat Commun. 2021; 12(1):5543. PMC: 8452764. DOI: 10.1038/s41467-021-25683-4. View

Wang L, Wang S, Wu L, Li W, Bray W, Clark A . PCIF1-mediated deposition of 5'-cap ,2'--dimethyladenosine in ACE2 and TMPRSS2 mRNA regulates susceptibility to SARS-CoV-2 infection. Proc Natl Acad Sci U S A. 2023; 120(5):e2210361120. PMC: 9945940. DOI: 10.1073/pnas.2210361120. View

Fan H, Sakuraba K, Komuro A, Kato S, Harada F, Hirose Y . PCIF1, a novel human WW domain-containing protein, interacts with the phosphorylated RNA polymerase II. Biochem Biophys Res Commun. 2003; 301(2):378-85. DOI: 10.1016/s0006-291x(02)03015-2. View

10.

Eggenschwiler J, Espinoza E, Anderson K . Rab23 is an essential negative regulator of the mouse Sonic hedgehog signalling pathway. Nature. 2001; 412(6843):194-8. DOI: 10.1038/35084089. View

11.

Roundtree I, Evans M, Pan T, He C . Dynamic RNA Modifications in Gene Expression Regulation. Cell. 2017; 169(7):1187-1200. PMC: 5657247. DOI: 10.1016/j.cell.2017.05.045. View

12.

Wang L, Wu L, Zhu Z, Zhang Q, Li W, Gonzalez G . Role of PCIF1-mediated 5'-cap N6-methyladeonsine mRNA methylation in colorectal cancer and anti-PD-1 immunotherapy. EMBO J. 2022; 42(2):e111673. PMC: 9841328. DOI: 10.15252/embj.2022111673. View

13.

He P, He C . m A RNA methylation: from mechanisms to therapeutic potential. EMBO J. 2021; 40(3):e105977. PMC: 7849164. DOI: 10.15252/embj.2020105977. View

14.

Sugita A, Kuruma S, Yanagisawa N, Ishiguro H, Kano R, Ohkuma Y . The cap-specific m6A methyltransferase, PCIF1/CAPAM, is dynamically recruited to the gene promoter in a transcription-dependent manner. J Biochem. 2021; 170(2):203-213. DOI: 10.1093/jb/mvab032. View

15.

Song M, Li Y, Kiledjian M . Multiple mRNA decapping enzymes in mammalian cells. Mol Cell. 2010; 40(3):423-32. PMC: 2982215. DOI: 10.1016/j.molcel.2010.10.010. View

16.

Yunokuchi I, Fan H, Iwamoto Y, Araki C, Yuda M, Umemura H . Prolyl isomerase Pin1 shares functional similarity with phosphorylated CTD interacting factor PCIF1 in vertebrate cells. Genes Cells. 2009; 14(9):1105-18. DOI: 10.1111/j.1365-2443.2009.01339.x. View

17.

Sun H, Li K, Zhang X, Liu J, Zhang M, Meng H . mAm-seq reveals the dynamic mAm methylation in the human transcriptome. Nat Commun. 2021; 12(1):4778. PMC: 8346571. DOI: 10.1038/s41467-021-25105-5. View

18.

Sun H, Zhang M, Li K, Bai D, Yi C . Cap-specific, terminal N-methylation by a mammalian mAm methyltransferase. Cell Res. 2018; 29(1):80-82. PMC: 6318281. DOI: 10.1038/s41422-018-0117-4. View

19.

Shi H, Wei J, He C . Where, When, and How: Context-Dependent Functions of RNA Methylation Writers, Readers, and Erasers. Mol Cell. 2019; 74(4):640-650. PMC: 6527355. DOI: 10.1016/j.molcel.2019.04.025. View

20.

Zhang X, Wei L, Wang Y, Xiao Y, Liu J, Zhang W . Structural insights into FTO's catalytic mechanism for the demethylation of multiple RNA substrates. Proc Natl Acad Sci U S A. 2019; 116(8):2919-2924. PMC: 6386707. DOI: 10.1073/pnas.1820574116. View