Abstract
N6-methyladenosine (m6A) is the most common internal modification in eukaryotic mRNA. It is dynamically installed and removed, and acts as a new layer of mRNA metabolism, regulating biological processes including stem cell pluripotency, cell differentiation, and energy homeostasis. m6A is recognized by selective binding proteins; YTHDF1 and YTHDF3 work in concert to affect the translation of m6A-containing mRNAs, YTHDF2 expedites mRNA decay, and YTHDC1 affects the nuclear processing of its targets. The biological function of YTHDC2, the final member of the YTH protein family, remains unknown. We report that YTHDC2 selectively binds m6A at its consensus motif. YTHDC2 enhances the translation efficiency of its targets and also decreases their mRNA abundance. Ythdc2 knockout mice are infertile; males have significantly smaller testes and females have significantly smaller ovaries compared to those of littermates. The germ cells of Ythdc2 knockout mice do not develop past the zygotene stage and accordingly, Ythdc2 is upregulated in the testes as meiosis begins. Thus, YTHDC2 is an m6A-binding protein that plays critical roles during spermatogenesis.
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Acknowledgements
We thank Prof Tao Pan (The University of Chicago) for kindly providing the polysome profiling instrument. We thank Peter Chen and Phil Xie for technical assistance. We also thank the Institutes of Biomedical Sciences at Fudan University for the protein mass spectrometry experiments. This work was supported by the National Science Fund for Excellent Young Scholars (31622039 to BS), the National Key Research and Development Program of China (2016YFC1000600 to BS, 2016YFA0500902 to ML, 2017YFA0103803 to JS), the Science Foundation for Distinguished Young Scholars of Jiangsu Province (BK20160045 to BS), and National Institutes of Health grants HG008935 and GM113194 (to CH). CH is an investigator of the Howard Hughes Medical Institute (HHMI). HM is supported by the Postdoctoral International Exchange Program of the China Postdoctoral Council (CPC). PH is supported by the University of Chicago Medical Scientist Training Program (MSTP; NIH MSTP training grant T32GM007281).
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( Supplementary information is linked to the online version of the paper on the Cell Research website.)
Supplementary information
Supplementary information, Figure S1
Domains of YTHDC2 (https://www.ncbi.nlm.nih.gov/gene/64848). (PDF 39 kb)
Supplementary information, Figure S2
(Supplement to Figure 1) YTHDC2 binds preferentially to m6A-marked RNA transcripts. (PDF 148 kb)
Supplementary information, Figure S3
Characterization of Ythdc2−/− mouse. (PDF 105 kb)
Supplementary information, Figure S4
Female Ythdc2−/− mice demonstrate defects in the ovaries. (PDF 152 kb)
Supplementary information, Figure S5
Expression of Ythdc2 protein in wild type testes over time. (PDF 110 kb)
Supplementary information, Figure S6
Meiotic defects in Ythdc2-deficient meiocytes. (PDF 195 kb)
Supplementary information, Figure S7
Comparative histological analysis of testes of young Ythdc2+/+ and Ythdc2−/− mice. (PDF 436 kb)
Supplementary information, Figure S8
Ythdc2-deficient mice show normal spermatogonia. (PDF 199 kb)
Supplementary information, Figure S9
Ythdc2-deficient mice show normal sertoli cells. (PDF 445 kb)
Supplementary information, Figure S10
(Supplement to Figure 4) YTHDC2 affects the translation efficiency and stability of its targets. (PDF 141 kb)
Supplementary information, Figure S11
m6A-seq of 8.5 d.p.p. Ythdc2+/+ and Ythdc2−/− testes. (PDF 107 kb)
Supplementary information, Figure S12
Effect of YTHDC2 on mRNA abundance and translation efficiency may be m6A dependent. (PDF 108 kb)
Supplementary information, Figure S13
Confirmation of components of protein complex containing YTHDC2 obtained from tandem affinity purification followed by western blot. (PDF 111 kb)
Supplementary information, Table S1
Target transcripts of Ythdc2 in adult mouse testes, determined by CLIP-seq (XLSX 1895 kb)
Supplementary information, Table S2
Target transcripts of Ythdc2 in 16 d.p.p. mouse testes, determined by RIP-seq (XLSX 1406 kb)
Supplementary information, Table S3
Top 40 GO terms of Ythdc2 RIP-seq targets in mouse testes (XLSX 48 kb)
Supplementary information, Table S4
m6A-seq of control and Ythdc2−/− mice (XLSX 8512 kb)
Supplementary information, Table S5
Targets of YTHDC2 in HeLa cells, determined by PAR-CLIP (XLSX 120 kb)
Supplementary information, Table S6
Binding partner proteins of YTHDC2 in HeLa cells, determined by Tandem Affinity Purification followed by mass spectrometry (XLSX 1107 kb)
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Hsu, P., Zhu, Y., Ma, H. et al. Ythdc2 is an N6-methyladenosine binding protein that regulates mammalian spermatogenesis. Cell Res 27, 1115–1127 (2017). https://doi.org/10.1038/cr.2017.99
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DOI: https://doi.org/10.1038/cr.2017.99
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