Genetic Regulation of N6-Methyladenosine-RNA in Mammalian Gametogenesis and Embryonic Development

Abstract

Emerging evidence shows that m⁶A is the most abundant modification in eukaryotic RNA molecules. It has only recently been found that this epigenetic modification plays an important role in many physiological and pathological processes, such as cell fate commitment, immune response, obesity, tumorigenesis, and relevant for the present review, gametogenesis. Notably the RNA metabolism process mediated by m⁶A is controlled and regulated by a series of proteins termed writers, readers and erasers that are highly expressed in germ cells and somatic cells of gonads. Here, we review and discuss the expression and the functional emerging roles of m⁶A in gametogenesis and early embryogenesis of mammals. Besides updated references about such new topics, readers might find in the present work inspiration and clues to elucidate epigenetic molecular mechanisms of reproductive dysfunction and perspectives for future research.

Keywords: embryo development; epigenetics; m6A; oogenesis; spermatogenesis.

FIGURE 1

Methylation and demethylation of m⁶A on RNA. The N ⁶-methyladenosine (m⁶A) modification is imposed by a heterocomplex of two methyltransferases METTL3-METTL14 termed “writers”, assisted by WTAP. ALKBH5 and FTO, termed “erasers” catalyzes the direct removal of m⁶A. YTH family proteins termed “readers” recognize the m⁶A to carry out subsequent functions.

FIGURE 2

The molecular mechanisms of how m⁶A affects splicing/polyadenylation/translation. YTHDF1 and YTHDF3 regulate m⁶A both individually and together. hnRNPG binds to m⁶A-containing 3′-terminal mRNA and interacts with 40S and 60S ribosomal subunits to regulate mRNA translation, and YTHDF3 also facilitates this process. hnRNPG binds to m6a-containing pre-mRNA and interacts with RNA polymerase II (RNAP II) phosphorylated carboxy-terminal domain (CTD) of RNA polymerase II, which in turn regulates alternative splicing of mRNAs. C-YTHDF2 selectively recognizes m⁶A-containing mRNAs, while N-YTHDF2 on the other side binds to mRNA decay machineries and regulates mRNA degradation. YTH domain binds to the m⁶A-containing pre-mRNA and regulates the length of the 3′ UTR through polyadenylation complexes.

FIGURE 3

The m⁶A modification-related proteins exerting stage-dependent functions during spermatogenesis in mammals. Writers: METTL3 (and at lesser extent METTL14) regulates mainly spermatogonial homeostasis; METTL3/METTL14 complexes regulate spermiogenesis, namely the formation of flagellum; METTL16 inhibits spermatogonia proliferation and ensures normal germ cell differentiation; Erasers: FTO regulates cell-cycle in spermatogonia; ALKBH5 improves translation efficiency and rapid protein turnover mainly during spermiogenesis; Readers: YTHDF2 regulates meiotic progression; YTHDC1 is involved in co-transcriptional and/or post-transcriptional regulations in spermatogonia, spermatocytes, and round spermatids; YTHDC2 promotes translation and plays an essential role in gene expression and promotes translation during meiosis. The highlighted color indicates that the proteins mainly play functions at this stage.

FIGURE 4

The function of YTHDC1, YTHDC2 and YTHDF2 in sperm cells. (A) YTHDC2 binds multiple transcripts including Ccna2 and other mitotic transcripts and interacts with RNA granule components, suggesting that proper progression of germ cells through meiosis is licensed by YTHDC2 through post-transcriptional regulation; when YTHDC2 is inactivated, spermatocytes do not pass the zygotene stage. (B) YTHDC1 can recruit SRSF3 and block SRSF10 to regulate alternative splicing of pre-mRNA crucial for maintaining spermatogonia. In addition, YTHDC1 can also promote the output of m⁶A-containing mRNA. YTHDF2 binds to the m⁶A site and degrade RNAs containing m⁶A involved in controlling migration and proliferation of spermatogonia.

FIGURE 5

Regulation of miRNA and circRNA by METTL3 and ALKBH5. Pre-miRNA methylated by METTL3 shortens the mRNA poly(A) tail, inhibits mRNA translation, and forms output tubule cilia. METTL3 and ALKBH5 jointly control the amount of m⁶A on circRNA, which is important for sperm quality and protein supplementation in late sperm stages.

FIGURE 6

m6A regulation in oogenesis. (A) KIAA1429 mediates the m6A modification on pre-mRNA, then YTHDC1 recognizes the m⁶A signal and recruits SRSF3 to the binding regions on the KIAA1429-mediated pre-mRNA and YTHDC1 alters 3′ UTR length through associating with pre-mRNA 3′ end processing factors CPSF6, SRSF3, and SRSF7 which results in splicing of mRNA and then promotes oogenesis. (B) m⁶A affected gene abundance of pluripotent marker Lin28. It interferes with pluripotent regulation of chromosome/spindle tissue and regulates GVBD, PB1 extrusion, cleavage, and blastocyst development of parthenogenesis.

FIGURE 7

The main role of m⁶A in the stage of embryonic development. The level of m6A modification increased slowly in mouse 2-cell, 4-cell and 8-cell embryos but suddenly increased significantly in the transition period from morula to blastocyst which is due to the METTL3 and METTL14 active transcription of many pluripotency genes (includes Nanog, Oct4, Sox2 and so on). In the MZT, METTL3 mediates the m6A modification on pre-mRNA to degrade maternal mRNA and YTHDF2 can promote the process, which is reprogramming zygotic mRNA for zygotic development.