MiR-202-5p is identified as the maker microRNA(miRNA)of germplasm in fish.However,the role of miR-202-5p in gonadal development is still unclear.Here we investigated the expression patterns of miR-202-5p in Japanese f...MiR-202-5p is identified as the maker microRNA(miRNA)of germplasm in fish.However,the role of miR-202-5p in gonadal development is still unclear.Here we investigated the expression patterns of miR-202-5p in Japanese flounder(Paralichthys olivaceus).Real-time PCR displayed that miR-202-5p was specifically expressed in bisexual gonads,and its transcription in the testis was relatively abundant than that in the ovary.Moreover,it was gradually increased in testis from stage II to stage IV.Fluorescent in situ hybridization(FISH)illustrated that miR-202-5p was co-localized with vasa(a marker gene of germ cells)in testis,and it was predominately expressed in spermatogonia and spermatocytes,implying that miR-202-5p might participate in spermatogenesis.Considering the important role of retinoic acid(RA)on regulating meiosis during spermatogenesis,we treated Japanese flounders by injecting all-trans retinoic acid(ATRA).MiR-202-5p in testis was shown to be up-regulated,and ccnd1,a key cell cycle regulator,was found to be down-regulated after ATRA treatment.Furthermore,ccnd1 was demonstrated to be a direct regulatory target of miR-202-5p using Dual-luciferase assay.In the primary testis cell from Japanese flounder,we further confirmed that ccnd1 was down-regulated by miR-202-5p.The results suggest that miR-202-5p exerts a crucial role in gonadal development by inhibiting ccnd1 in Japanese flounder.展开更多
In many lower animals,germ cell formation,migration,and maintenance depend on maternally provided determinants in germ plasm.In zebrafish,these processes have been extensively studied in terms of RNA-binding proteins ...In many lower animals,germ cell formation,migration,and maintenance depend on maternally provided determinants in germ plasm.In zebrafish,these processes have been extensively studied in terms of RNA-binding proteins and other coding genes.The role of small non-coding RNAs in the regulation of primordial germ cell(PGC)development remains largely unknown and poorly investigated,even though growing interests for the importance of miRNAs involved in a wide variety of biological processes.Here,we reported the role and mechanism of the germ plasm-specific miRNA miR-202-5p in PGC migration:(i)both maternal loss and knockdown of miR-202-5p impaired PGC migration indicated by the mislocalization and reduced number of PGCs;(ii)cdc42se1 was a direct target gene of miR-202-5p,and overexpression of Cdc42se1 in PGCs caused PGC migration defects similar to those observed in loss of miR-202-5p mutants;(iii)Cdc42se1 not only interacted with Cdc42 but also inhibited cdc42 transcription,and overexpression of Cdc42 could rescue PGC migration defects in Cdc42se1 overexpressed embryos.Thus,miR-202-5p regulates PGC migration by directly targeting and repressing Cdc42se1 to protect the expression of Cdc42,which interacts with actin to direct PGC migration.展开更多
基金supported by the National Natural Science Foundation of China(No.31972772)the Laboratory for Marine Fisheries Science and Food Production Processes,Qingdao National Laboratory for Marine Science and Technology,China(ZZ-A11).
文摘MiR-202-5p is identified as the maker microRNA(miRNA)of germplasm in fish.However,the role of miR-202-5p in gonadal development is still unclear.Here we investigated the expression patterns of miR-202-5p in Japanese flounder(Paralichthys olivaceus).Real-time PCR displayed that miR-202-5p was specifically expressed in bisexual gonads,and its transcription in the testis was relatively abundant than that in the ovary.Moreover,it was gradually increased in testis from stage II to stage IV.Fluorescent in situ hybridization(FISH)illustrated that miR-202-5p was co-localized with vasa(a marker gene of germ cells)in testis,and it was predominately expressed in spermatogonia and spermatocytes,implying that miR-202-5p might participate in spermatogenesis.Considering the important role of retinoic acid(RA)on regulating meiosis during spermatogenesis,we treated Japanese flounders by injecting all-trans retinoic acid(ATRA).MiR-202-5p in testis was shown to be up-regulated,and ccnd1,a key cell cycle regulator,was found to be down-regulated after ATRA treatment.Furthermore,ccnd1 was demonstrated to be a direct regulatory target of miR-202-5p using Dual-luciferase assay.In the primary testis cell from Japanese flounder,we further confirmed that ccnd1 was down-regulated by miR-202-5p.The results suggest that miR-202-5p exerts a crucial role in gonadal development by inhibiting ccnd1 in Japanese flounder.
基金supported by the National Natural Science Foundation of China(31771587 and 31970535)Fundamental Research Funds for the Central Universities(17lgpy66)+2 种基金the Pearl River S&T Nova Program of Guangzhou(201806010047)the Scince and Technology Planning Project of Guangdong Province(2017A030303010)the Guangdong Natural Science Foundation(2015A030308012).
文摘In many lower animals,germ cell formation,migration,and maintenance depend on maternally provided determinants in germ plasm.In zebrafish,these processes have been extensively studied in terms of RNA-binding proteins and other coding genes.The role of small non-coding RNAs in the regulation of primordial germ cell(PGC)development remains largely unknown and poorly investigated,even though growing interests for the importance of miRNAs involved in a wide variety of biological processes.Here,we reported the role and mechanism of the germ plasm-specific miRNA miR-202-5p in PGC migration:(i)both maternal loss and knockdown of miR-202-5p impaired PGC migration indicated by the mislocalization and reduced number of PGCs;(ii)cdc42se1 was a direct target gene of miR-202-5p,and overexpression of Cdc42se1 in PGCs caused PGC migration defects similar to those observed in loss of miR-202-5p mutants;(iii)Cdc42se1 not only interacted with Cdc42 but also inhibited cdc42 transcription,and overexpression of Cdc42 could rescue PGC migration defects in Cdc42se1 overexpressed embryos.Thus,miR-202-5p regulates PGC migration by directly targeting and repressing Cdc42se1 to protect the expression of Cdc42,which interacts with actin to direct PGC migration.
