The Dynamic Landscape Of Gene Regulation During Bombyx Mori Oogenesis

Oogenesis in the domestic silkworm(Bombyx mori)is a complex process involving previtellogenesis,vitellogenesis and choriogenesis.During this process,follicles show drastic morphological and physiological changes,as a functional unit of oogenesis.Although some studies have been done on the structure and hormonal regulation of silkworm follicles,the genome-wide regulatory profiles of gene expression during oogenesis remain to be determined.In this paper,the time-series transcriptome and MeRIP-seq method were used to analyze the gene regulation mechanism of silkworm oogenesis at transcription level and post-transcriptional level.The main results are as follows:(1)The oogenesis of silkworms involves many cell activities,such as germ cell differentiation,apoptosis,cell proliferation and so on.There is a significant change in ecdysone titer between the early and late stages of egg development.In this study,a total of 1932 genes were identified to be differentially expressed among adjacent stages,most of which occurred during the transition from late vitellogenesis to early choriogenesis,mainly enriched in the reproductive,cell differentiation,morphogenesis and transmembrane transport and other functional classification.Using weighted gene co-expression network analysis,we identified six stage-specific gene modules that correspond to multiple regulatory pathways and every module shows timing changes.Through the functional enrichment and pathway enrichment analysis of each module,the results show that the genes mainly were enriched in the immune,autophagy regulation,cell communication,signal transduction and dorsal ventral axis formation,unsaturated fatty acid synthesis and metabolism,ecdysone synthesis and metabolism pathway.Further analysis of genes involving with ecdysone synthesis and metabolism showed that,the ecdysteroid 20-hydroxylase gene(CYP314A1)of steroidogenesis genes was mainly expressed in previtellogenesis and early vitellogenesis.However,the 20E-inactivated genes,particularly the ecdysteroid 26-hydroxylase encoding gene(Cyp18a1),were highly expressed in late vitellogenesis.These distinct expression patterns between ecdysone synthesis and catabolism-related genes might ensure the rapid changes of the hormone titer during oogenesis.In addition,by in vitro injection of insulin signaling pathway inhibitor experiments,the result initially confirmed that insulin may be an essential factor in the occurrence of normal eggs.(2)Although silkworm(Lepidoptera)and fruit fly(Diptera)belong to different insect classification,they are the main model insects of polytrophic meroistic ovarian.Thus,we compared landscapes of gene regulation between these insects.Our results show that there are four consensus modules of gene co-expression during oogenesis in these insects,involving a number of key genes related to hormone regulation,signal regulation and RNA modification,which provides a new basis for the study of molecular regulation mechanism of polytrophic meroistic ovarian development(3)In this study,nine types of alternative splicing were identified,and splicing donor site changes and exon changes were the main causes of alternative splicing.It was found that histone and RNA modified genes showed similarly molecular switches during oogenesis of silkworms.The dynamic changes of RNA modification around the transition from vitellogenesis to choriogenesis were analyzed by MeRIP-seq.The results showed that the m6A loci were mainly distributed in the genomic coding region(about 85%)and the 5'UTR,CDS and 3'UTR regions,and the 5'UTR region was more.There were significant differences in the m~6A modification site between the early and late stages of oogenesis and the function of the modified genes was different.However,the signal pathways,such as Hippo,Wnt and Notch,were significantly enriched,indicating that these pathways may be the main target pathway of m~6A modification.In addition,the study also found evidence of m~6A modifications in several different types of transposon transcripts,indicating that these types of transposons are active in egg development,and m~6A modifications may be necessary for transposon transposons.Our results provide further understanding of the important role of RNA post-transcription regulation during silkworm oogenesis.The results indicate that there are complex regulations at transcription level and post-transcriptional level during silkworm oogenesis.Develpmental follicles are an excellent model to study the molecular mechanism of insect oogenesis,which is helpful to better understanding the molecular mechanism of insect gametogenesis and reproduction.