No abstract

N-methyladenosine (mA), catalyzed by Mettl3 methyltransferase, is a highly conserved epigenetic modification in eukaryotic messenger RNA (mRNA). Previous studies have implicated mA modification in multiple biological processes, but the function of mA has been difficult to study, because mutants are embryonic lethal in both mammals and plants. In this study, we have used transcription activator-like effector nucleases and generated viable zygotic mutant, Z , in zebrafish. We find that the oocytes in Z 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 and the numbers of eggs ovulated are both decreased as well, while the defects of oocyte maturation can be rescued by sex hormone and In Z 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 , and defective gamete maturation is accompanied by decreased overall mA modification levels and disrupted expression of genes critical for sex hormone synthesis and gonadotropin signaling in Z Thus, our study provides the first evidence that loss of Mettl3 leads to failed gamete maturation and significantly reduced fertility in zebrafish. Mettl3 and mA modifications are essential for optimal reproduction in vertebrates.

We showed that the Ad-sig-TAA/ecdCD40L vaccine induces a tumor suppressive immune response to the hMUC-1 and rH2N tumor-associated self Ags (TAA) and to the Annexin A1 tumor vascular Ag, even in mice in which anergy exists to these Ags. When the TAA/ecdCD40L protein is given s.c. as a boost following the Ad-sig-TAA/ecdCD40L vector, the levels of the TAA-specific CD8 T cells and Abs increase dramatically over that seen with vector alone, in young (2-mo-old) as well as old (18-mo-old) mice. The Abs induced against hMUC-1 react with human breast cancer. This vaccine also induces a 4-fold decrement of negative regulatory CD4CD25FOXP3-T cells in the tumor tissue of 18-mo-old mice. These results suggest that the Ad-sig-TAA/ecdCD40L vector prime-TAA/ecdCD40L protein boost vaccine platform may be valuable in reducing postsurgery recurrence in a variety of epithelial neoplasms.