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Analysis of protein-protein interaction network based on transcriptome profiling of ovine granulosa cells identifies candidate genes in cyclic recruitment of ovarian follicles.

ABSTRACT: After pubertal, cohort of small antral follicles enters to gonadotrophin-sensitive development, called recruited follicles. This study was aimed to identify candidate genes in follicular cyclic recruitment via analysis of protein-protein interaction (PPI) network. Differentially expressed genes (DEGs) in ovine granulosa cells of small antral follicles between follicular and luteal phases were accumulated among gene/protein symbols of the Ensembl annotation. Following directed graphs, PTPN6 and FYN have the highest indegree and outdegree, respectively. Since, these hubs being up-regulated in ovine granulosa cells of small antral follicles during the follicular phase, it represents an accumulation of blood immune cells in follicular phase in comparison with luteal phase. By contrast, the up-regulated hubs in the luteal phase including CDK1, INSRR and TOP2A which stimulated DNA replication and proliferation of granulosa cells, they known as candidate genes of the cyclic recruitment.

SUBMITTER: Talebi R

PROVIDER: S-EPMC5994657 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Analysis of protein-protein interaction network based on transcriptome profiling of ovine granulosa cells identifies candidate genes in cyclic recruitment of ovarian follicles.

Talebi Reza R Ahmadi Ahmad A Afraz Fazlollah F

Journal of animal science and technology 20180611

After pubertal, cohort of small antral follicles enters to gonadotrophin-sensitive development, called recruited follicles. This study was aimed to identify candidate genes in follicular cyclic recruitment via analysis of protein-protein interaction (PPI) network. Differentially expressed genes (DEGs) in ovine granulosa cells of small antral follicles between follicular and luteal phases were accumulated among gene/protein symbols of the Ensembl annotation. Following directed graphs, <i>PTPN6</i ...[more]

PMID: 29992036

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Project description:BackgroundThe major function of the ovary is to produce oocytes for fertilisation. Oocytes mature in follicles surrounded by nurturing granulosa cells and all are enclosed by a basal lamina. During growth, granulosa cells replicate and a large fluid-filled cavity (the antrum) develops in the centre. Only follicles that have enlarged to over 10 mm can ovulate in cows. In mammals, the number of primordial follicles far exceeds the numbers that ever ovulate and atresia or regression of follicles is a mechanism to regulate the number of oocytes ovulated and to contribute to the timing of ovulation. To better understand the molecular basis of follicular atresia, we undertook transcriptome profiling of granulosa cells from healthy (n = 10) and atretic (n = 5) bovine follicles at early antral stages (< 5 mm).ResultsPrincipal Component Analysis (PCA) and hierarchical classification of the signal intensity plots for the arrays showed primary clustering into two groups, healthy and atretic. These analyses and size-frequency plots of coefficients of variation of signal intensities revealed that the healthy follicles were more heterogeneous. Examining the differentially-expressed genes the most significantly affected functions in atretic follicles were cell death, organ development, tissue development and embryonic development. The overall processes influenced by transcription factor gene TP53 were predicted to be activated, whereas those of MYC were inhibited on the basis of known interactions with the genes in our dataset. The top ranked canonical pathway contained signalling molecules common to various inflammatory/fibrotic pathways such as the transforming growth factor-β and tumour necrosis factor-α pathways. The two most significant networks also reflect this pattern of tissue remodelling/fibrosis gene expression. These networks also contain molecules which are present in the canonical pathways of hepatic fibrosis/hepatic stellate cell activation and transforming growth factor-β signalling and were up regulated.ConclusionsSmall healthy antral follicles, which have a number of growth outcomes, exhibit greater variability in gene expression, particularly in genes associated with cell division and other growth-related functions. Atresia, on the other hand, not only involves cell death but clearly is an active process similar to wound healing.

Project description:BackgroundAt later stages of folliculogenesis, the mammalian ovarian follicle contains layers of epithelial granulosa cells surrounding an antral cavity. During follicle development granulosa cells replicate, secrete hormones and support the growth of the oocyte. In cattle, the follicle needs to grow > 10 mm in diameter to allow an oocyte to ovulate, following which the granulosa cells cease dividing and differentiate into the specialised cells of the corpus luteum. To better understand the molecular basis of follicular growth and granulosa cell maturation, we undertook transcriptome profiling of granulosa cells from small (< 5 mm; n = 10) and large (> 10 mm, n = 4) healthy bovine follicles using Affymetrix microarrays (24,128 probe sets).ResultsPrincipal component analysis for the first two components and hierarchical clustering showed clustering into two groups, small and large, with the former being more heterogeneous. Size-frequency distributions of the coefficient of variation of the signal intensities of each probe set also revealed that small follicles were more heterogeneous than the large. IPA and GO enrichment analyses revealed that processes of axonal guidance, immune signalling and cell rearrangement were most affected in large follicles. The most important networks were associated with: (A) Notch, SLIT/ROBO and PI3K signalling, and (B) ITGB5 and extracellular matrix signalling through extracellular signal related kinases (ERKs). Upstream regulator genes which were predicted to be active in large follicles included STAT and XBP1. By comparison, developmental processes such as those stimulated by KIT, IHH and MEST were most active in small follicles. MGEA5 was identified as an upstream regulator in small follicles. It encodes an enzyme that modifies the activity of many target proteins, including those involved in energy sensing, by removal of N-acetylglucosamine from serine and threonine residues.ConclusionsOur data suggest that as follicles enlarge more genes and/or pathways are activated than are inactivated, and gene expression becomes more uniform. These findings could be interpreted that either the cells in large follicles are more uniform in their gene expression, or that follicles are more uniform or a combination of both and that additional factors, such as LH, are additionally controlling the granulosa cells.

Project description:BackgroundLAPTM4B is a member of the lysosome-associated transmembrane protein superfamily that is differentially expressed in normal human tissues and upregulated in various types of carcinomas. These proteins are thought to be involved in the regulation of cell proliferation and survival. The objective of this study was to investigate the expression of bovine LAPTM4B during ovarian follicular development and in various bovine tissues.Methods and resultsNorthern blot analysis revealed a 1.8 kb transcript, with highly variable steady state levels among tissues. RT-PCR analysis showed that LAPTM4B mRNA transcripts were low in granulosa cells of small antral follicles, increased in large dominant follicles, and decreased in ovulatory follicles following injection of human chorionic gonadotropin (hCG; P < 0.003). Ovulatory follicles collected at various times after hCG injection revealed a significant reduction of LAPTM4B mRNA starting at 18 h post-hCG (P < 0.029). Immunoblotting analysis using antibodies generated against bovine LAPTM4B recognized proteins of 26.3 and 31.5 kDa in granulosa cells of developing follicles and corpus luteum. Further analyses of affinity-purified His-tag LAPTM4B overexpressed in HEK cells showed that the 31.5 kDa protein represented the ubiquinated isoform of the 26.3 kDa native protein. The 26.3 kDa protein was differentially expressed showing highest amounts in dominant follicles and lowest amounts in ovulatory follicles 24 h post-hCG. Immunohistochemical analyses of LAPTM4B showed marked heterogeneity of labeling signal among tissues, with LAPTM4B mainly localized to perinuclear vesicles, in keeping with its putative lysosomal membrane localization.ConclusionThis study reports for the first time that bovine LAPTM4B in granulosa cells is present in both unubiquinated and ubiquinated forms, and is differentially expressed in developing ovarian follicles, suggesting a possible role in terminal follicular growth.

Project description:Follicle development is characterized by the recruitment, growth, selection, and dominance of follicles, and follicle selection determines the lifetime reproductive performance. However, in birds, the molecular mechanisms underlying follicle selection still remain elusive. This study analyzed genome-wide changes in the mRNA and miRNA expression profiles in both the granulosa and theca layers of geese ovarian follicles before selection (4-6- and 8-10-mm follicles) and after selection (F5). The sequencing results showed that a higher number of both differentially expressed (DE) mRNAs and DE miRNAs were identified between 8-10-mm and F5 follicles compared with those between the 4-6- and 8-10-mm follicles, especially in the granulosa layer. Moreover, a Short Time-series Expression Miner analysis identified a large number of DE mRNAs and DE miRNAs that are associated with follicle selection. The functional enrichment analysis showed that DE genes in the granulosa layer during follicle selection were mainly enriched in five pathways related to junctional adhesion and two pathways associated with lipid metabolism. Additionally, an interaction network was constructed to visualize interactions among protein-coding genes, which identified 53 junctional adhesion- and 15 lipid regulation-related protein-coding genes. Then, a co-expression network between mRNAs and miRNAs in relation to junctional adhesion was also visualized and mainly included acy-miR-2954, acy-miR-218, acy-miR-2970, acy-miR-100, acy-miR-1329, acy-miR-199, acy-miR-425, acy-miR-181, and acy-miR-147. Furthermore, miRNA-mRNA interaction pairs related to lipid regulation were constructed including acy-miR-107, acy-miR-138, acy-miR-130, acy-miR-128, and acy-miR-101 during follicular selection. In summary, these data highlight the key roles of junctional adhesion and lipid metabolism during follicular selection and contribute to a better understanding of the mechanisms underlying follicle selection in birds.

Dataset Information

Analysis of protein-protein interaction network based on transcriptome profiling of ovine granulosa cells identifies candidate genes in cyclic recruitment of ovarian follicles.

Publications

Analysis of protein-protein interaction network based on transcriptome profiling of ovine granulosa cells identifies candidate genes in cyclic recruitment of ovarian follicles.

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