The ocnus (ocn) gene encodes a protein abundant in the testes, implying its role in testis development. When Drosophila melanogaster is infected with the endosymbiont wMel Wolbachia, which affects the spermatogenesis of its hosts, ocn is downregulated in the third-instar larval testes, suggesting a role of ocn in spermatogenesis. In this study, we knocked down ocn in the testes and found that the hatch rates of embryos derived from ocn-knockdown males were significantly decreased, and 84.38% of the testes were much smaller in comparison to controls. Analysis of the smaller testes showed no germ cells but they had an extended hub. Using RNA-sequencing (RNA-Seq), we identified 69 genes with at least a twofold change (q-value < 5%) in their expression after ocn knockdown; of these, eight testes-specific and three reproduction-related genes were verified to be significantly downregulated using quantitative reverse transcription-PCR. Three genes (orientation disruptor, p24-2 and CG13541) were also significantly downregulated in the presence of Wolbachia. Furthermore, 98 genes were not expressed when ocn was knocked down in testes. These results suggest that ocn plays a crucial role in male germ cell development in Drosophila, possibly by regulating the expression of multiple spermatogenesis-related genes. Our data provide important information to help understand the molecular regulatory mechanisms underlying spermatogenesis.
Activating transcription factor 5 (Atf5) is a member of the ATF/CREB family of transcription factors and involved in diverse cellular functions and diseases in mammals. However, the function of atf5 remains largely unknown in fish. Here, we report the expression pattern and function of duplicated atf5 genes in zebrafish. The results showed that the gene structures of zebrafish atf5a and atf5b were similar to their mammalian orthologs. Zebrafish Atf5a and Atf5b shared an amino acid sequence identity of 40.7%. Zebrafish atf5a and atf5b had maternal origin with dynamic expression during embryonic development. Zebrafish atf5a mRNA is mainly enriched in olfactory epithelium, midbrain, and hindbrain, while zebrafish atf5b mRNA is mainly detected in midbrain, hindbrain, and liver during embryogenesis. The results of acute hypoxia experiment showed that atf5a mRNA was significantly upregulated in the brain, liver, and muscle, while atf5b mRNA was just increased significantly in the brain. Functional analysis showed that knockdown of atf5a affects the development of the ciliated neurons in zebrafish embryos. The effect was enhanced when atf5a MO was co-injected with atf5b MO. The development of ciliated neurons in zebrafish embryos was not affected by injection of atf5b MO alone. atf5a knockdown also affects the development of early-born olfactory neurons. The effects caused by atf5a knockdown could be rescued by atf5b mRNA.These results suggest that the duplicated atf5 genes may have evolved divergently and play redundant biological roles in the development of olfactory sensory neurons in zebrafish.
Arginine methylation is an important posttranslational modification and catalyzed by a family of protein arginine methyltransferases (PRMTs). PRMT7 is the type III PRMT and produces solely monomethylarginine products. PRMT7 has been found to play important roles in multiple biological processes in mammals. However, the expression pattern and function of Prmt7 remain largely unknown in fish. In this study, we characterized the medaka prmt7 gene and determined its expression pattern and function during embryogenesis and germ cell development. The results showed that the chromosomal location and gene structure of medaka prmt7 were similar to its mammalian orthologs. Comparisons of deduced amino acid sequences indicated that medaka Prmt7 was a homolog of human PRMT7 with two methyltransferase domains. Reverse transcription‐polymerase chain reaction (RT‐PCR) and real time RT‐PCR revealed that medaka prmt7 had maternal origin with continuous and dynamical expression during embryonic development. Whole‐mount in situ hybridization analysis observed that the transcripts of prmt7 were ubiquitous at morula and gastrula stage, and were later riched in the brain and otic vesicles during embryogenesis. In the adult stage, prmt7 messenger RNA was detected in all examined tissues with the high levels in the ovary and testis. The expression of prmt7 in the gonads was restricted to oocytes of the ovary and spermatids/sperm of the testis. Functional analysis showed that knockdown of medaka prmt7 did not reduce the total number of primordial germ cells (PGCs) in vivo but significantly affected PGCs distribution during embryonic development. These results indicate that prmt7 may be involved in germ cell development in medaka.
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