| Poultry follicle development is a dynamically biological process,and temporally specific expression of large number of functional genes precisely regulate this process.Hi-C,the high-throughput chromosome conformation capture technology,first appeared in 2009,which makes it possible to study the changes of chromatin spatial organization during development process through a three-dimensional genomic perspective.It was found that higher-order chromatin structure has become an important regulator of gene expression,but its changes during follicle development are still unknown.In this study,we used RNA-seq to study the m RNA and mi RNA expression profiles of the chicken follilcular granulosa cells from 10 developmental stages,including pre-hierarchical follicle(SWF,LWF,SYF and LYF),hierarchical follicle(F5,F4,F3,F2 and F1)and post-ovulatory follicle(POF).We identified the temporal expression profiles of m RNAs and mi RNAs during follicle development and constructed the mi RNA-m RNA targeted network.According to the results of stage-specific gene expression and number of differentially expression genes(DEGs),we then selected SWF,F1 and POF to generate genome-wide contact maps of chicken follicle development using Hi-C technology.We obtained the matrix of chromatin contact across developmental stages.Compartment A/B,topological associated domain(TAD),and enhancer-promoter interaction(PEI)were explored in relation to the regulation of gene expression.The dynamic changes of chromatin spatial structure during follicle development were analyzed by functional gene set analysis.The main results are listed as follows:1.During chicken follicle development process,there was little difference in the m RNA expression profile of granulosa cells at the internal adjacent time points of the prehierarchical stage and hierarchical stage,but the distinction among the three major stages was obvious.The number of specific genes in top 100 was the largest at SWF,F1 and POF stages.While expression profile of mi RNA at 10 stages could be clearly divided into three stages: pre-hierarchical,preovulatory and post-ovulatory stage.Among them,mi R-148a-3p,with the highest expression abundacnce at all stages,showed significant dynamic changes across development.2.Ma Sig Pro analysis showed that four clusters with temporal changes were identified in both m RNA and mi RNA,and a temporal mi RNA-m RNA regulatory network was constructed.Functional enrichment analysis showed that temporally expressed m RNA and mi RNA function variation during follicular development.3.The proportions of compartment A and B at each stage of follicle development were roughly equal,but 36% of the genome changes compartments during follicle development,and the A/B compartments had a contributory but not deterministic role on gene expression,just modest correlation may be due to the possibility that only a subset of genes may be affected by compartment changes.4.A total of 2000 TADs were identified for each sample.The average size of TADs was465 kb,occupying more than 84.75% of the genome.More than 98% of the TAD boundaries were stable and conserved during follicle development.In the boundary with stage-specific changes,acquisition of new boundary was linked to the upregulation of gene expression,while the boundary losting caused a certain level of decline in gene expression,and the intensity of interaction within TAD was positively correlated with gene expression.5.Enhancer number was positively correlated with level of gene expression,and it had additive effect on gene expression.The specific RP Index of genes at different stages were matched with the corresponding biological function during follicle development.6.The structure of the chromatin 3D model showed that the large chromosomes(such as chromosomes 1,2,and 3)are located in the periphery of the cell nucleus,and the small chromosomes are more concentrated near the center of the nucleus(such as chromosomes22,23,24,25,27),and A/B compartments and gene expression levels were related to relative nuclear position.To sum up,the spatial organization of granulosa cell at different hierarchical structural units has undergone significant changes during chicken follicle development process.The perturbation caused by these local contact changes leads to the increase or decrease of the number of enhancers,which leads to the change of gene expression.This study analyzed the dynamic change of chicken follicle development through the perspective of threedimensional(3D)genomics and provided new insights for the study of gene transcriptional regulation. |
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