Zc3h13 Regulates Nuclear RNA mA Methylation and Mouse Embryonic Stem Cell Self-Renewal
Authors: Fangfang Jiao
Hang Liu
Li Tan
Yang Shi
Jiahua Wang
Ruitu Lv
Chenxi He
Jing Wen
Hongjie Shen
Pengyuan Yang
Honghui Ma
Yujiang Geno Shi
Fei Lan
Chuan He
Jianbo Diao
Journal: Molecular cell (2018)
Discipline: MOLECULAR BIOLOGY & GENETICS
Keywords: Zc3h13; m(6)A; mESC self-renewal; nuclear localization.
Affiliations: Institute of Clinical Science, Zhongshan Hospital, and Institutes of Biomedical Sciences, Fudan University, Shanghai 200032, P.R. China., Department of Systems Biology, Institutes of Biomedical Sciences, Fudan University, 138 Yixueyuan Road, Shanghai 200032, China., Institute of Clinical Science, Zhongshan Hospital, and Institutes of Biomedical Sciences, Fudan University, Shanghai 200032, P.R. China; Key Laboratory of Birth Defect, Children's Hospital of Fudan University, Shanghai 201102, China., Institute of Clinical Science, Zhongshan Hospital, and Institutes of Biomedical Sciences, Fudan University, Shanghai 200032, P.R. China; Key Laboratory of Birth Defect, Children's Hospital of Fudan University, Shanghai 201102, China; Division of Endocrinology, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA. Electronic address: ., Department of Chemistry, and Department of Biochemistry and Molecular Biology, Institute for Biophysical Dynamics, University of Chicago, 929 East 57th Street, Chicago, IL 60637, USA; Howard Hughes Medical Institute, University of Chicago, 929 East 57th Street, Chicago, IL 60637, USA. Electronic address: ., Institute of Clinical Science, Zhongshan Hospital, and Institutes of Biomedical Sciences, Fudan University, Shanghai 200032, P.R. China; Key Laboratory of Birth Defect, Children's Hospital of Fudan University, Shanghai 201102, China; Newborn Medicine Division, Boston Children's Hospital, Boston, MA 02115, USA; Department of Cell Biology, Harvard Medical School, Boston, MA 02115, USA. Electronic address: ., Institute of Clinical Science, Zhongshan Hospital, and Institutes of Biomedical Sciences, Fudan University, Shanghai 200032, P.R. China; Key Laboratory of Metabolism and Molecular Medicine, Ministry of Education, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China; Key Laboratory of Epigenetics, Department of Cellular and Genetic Medicine, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China; The Fifth People's Hospital of Shanghai, Fudan University, Shanghai 200240, China. Electronic address: .

Abstract:  N6-methyladenosine (m6A) is an abundant modification in eukaryotic mRNA, regulating mRNA dynamics by influencing mRNA stability, splicing, export, and translation. However, the precise m6A regulating machinery still remains incompletely understood. Here we demonstrate that ZC3H13, a zinc-finger protein, plays an important role in modulating RNA m6A methylation in the nucleus. We show that knockdown of Zc3h13 in mouse embryonic stem cell significantly decreases global m6A level on mRNA. Upon Zc3h13 knockdown, a great majority of WTAP, Virilizer, and Hakai translocate to the cytoplasm, suggesting that Zc3h13 is required for nuclear localization of the Zc3h13-WTAP-Virilizer-Hakai complex, which is important for RNA m6A methylation. Finally, Zc3h13 depletion, as does WTAP, Virilizer, or Hakai, impairs self-renewal and triggers mESC differentiation. Taken together, our findings demonstrate that Zc3h13 plays a critical role in anchoring WTAP, Virilizer, and Hakai in the nucleus to facilitate m6A methylation and to regulate mESC self-renewal. 

Retracted: No

PubPeer comments:
 **Figure S3J and K:  (J-L) Western blot analysis of cytoplasmic and nuclear fractions of Mettl3 and Mettl14 in WTAP kd (J), Virilizer kd (K), or Hakai kd (L) mESCs and control cells.  ** Marked areas (in green and blue) display a very high degree of similarity despite the fact that these two panels were supposed to represent different cell treatments. ![file] , 
 **Figure S3D:  (D) Western blot analysis of cytoplasmic and nuclear fractions of WTAP, Virilizer, Hakai, Mettl3 and Mettl14 in Zc3h13 kd, rescue and control mESCs. ** All the bands in the Mettl3 and Mettl14 lanes (in red) show a highly similar running pattern. ![file] , 
 #1, #2 The panels you have noted are similar but they are not identical. In all cases, numerous distinct features (e.g., shapes of bands, etc.) are readily apparent after a few seconds of close inspection. I see no basis for concern. , 
 The images are not identical, but they could certainly be different exposures of the same blot. It would be useful for the authors to check their records to ensure that these images come from different blots. I agree with #3 Leucanella Acutissima, that the images do not appear identical. However, that doesn t mean that there wasn t a mix-up and the incorrect blots were assembled into the figure. The patterns of the shapes of the bands is remarkably similar in the instances highlighted above. To me they look like different exposure times of the same western blot. ,