<i>Mettl3</i> Mutation Disrupts Gamete Maturation and Reduces Fertility in Zebrafish

Abstract

N⁶-methyladenosine (m⁶A), catalyzed by Mettl3 methyltransferase, is a highly conserved epigenetic modification in eukaryotic messenger RNA (mRNA). Previous studies have implicated m⁶A modification in multiple biological processes, but the <i>in vivo</i> function of m⁶A has been difficult to study, because <i>mettl3</i> mutants are embryonic lethal in both mammals and plants. In this study, we have used transcription activator-like effector nucleases and generated viable zygotic <i>mettl3</i> mutant, Z<i>mettl3^m/m</i> , in zebrafish. We find that the oocytes in Z<i>mettl3^m/m</i> adult females are stalled in early development and the ratio of full-grown stage (FG) follicles is significantly lower than that of wild type. Human chorionic gonadotropin-induced ovarian germinal vesicle breakdown <i>in vitro</i> and the numbers of eggs ovulated <i>in vivo</i> are both decreased as well, while the defects of oocyte maturation can be rescued by sex hormone <i>in vitro</i> and <i>in vivo</i> In Z<i>mettl3^m/m</i> adult males, we find defects in sperm maturation and sperm motility is significantly reduced. Further study shows that 11-ketotestosterone (11-KT) and 17β-estradiol (E2) levels are significantly decreased in Z<i>mettl3^m/m</i> , and defective gamete maturation is accompanied by decreased overall m⁶A modification levels and disrupted expression of genes critical for sex hormone synthesis and gonadotropin signaling in Z<i>mettl3^m/m</i> Thus, our study provides the first <i>in vivo</i> evidence that loss of Mettl3 leads to failed gamete maturation and significantly reduced fertility in zebrafish. Mettl3 and m⁶A modifications are essential for optimal reproduction in vertebrates.