TGF-β1 resulting in differential microRNA expression in bovine granulosa cells

Yefen Xu; Jiaqiang Niu; Guangying Xi; Xuezhi Niu; Yuheng Wang; Ming Guo; Qiangba Yangzong; Yilong Yao; Suo Lang Sizhu; Jianhui Tian

doi:10.1016/j.gene.2018.04.036

TGF-β1 resulting in differential microRNA expression in bovine granulosa cells

Gene. 2018 Jul 15:663:88-100. doi: 10.1016/j.gene.2018.04.036. Epub 2018 Apr 14.

Authors

Affiliations

¹ Department of Animal Science, Tibet Agricultural and Animal Husbandry College, Nyingzhi, Tibet 860000, China.
² Ministry of Agriculture Key Laboratory of Animal Genetics, Breeding and Reproduction, National Engineering Laboratory for Animal Breeding, College of Animal Sciences and Technology, China Agricultural University, Haidian, Beijing 100193, China.
³ Department of Mechanical and Biomedical Engineering, College of Science and Engineering, City University of Hong Kong, Hong Kong 999077, China.
⁴ College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, Jiangsu 210095, China.
⁵ Department of Animal Science, Tibet Agricultural and Animal Husbandry College, Nyingzhi, Tibet 860000, China. Electronic address: 574428982@qq.com.

PMID: 29665451
DOI: 10.1016/j.gene.2018.04.036

Abstract

To explore the expression profile of the cellular miRNAs in bovine ovarian granulosa cells responding to transforming growth factor-β1 (TGF-β1), the effect of TGF-β1 on cell proliferation was firstly investigated by CCK-8 method and the results showed that there was a significant inhibitory effect on bovine granulosa cell proliferation treated with 5/10 ng/mL human recombinant TGF-β1 for 24 h compared to the control (P < 0.05). Then, we performed high-throughput sequencing of two small RNA libraries prepared from cultured bovine granulosa cells stimulated with or without 10 ng/mL human recombinant TGF-β1. A total of 13,257,248 and 138,726,391 clean reads per library were obtained from TGF-β1 and control groups, respectively. There were 498 and 499 bovine-specific exist miRNAs (exist miRNAs), 627 and 570 conserved known miRNAs (known miRNAs), and 593 and 585 predicted novel miRNAs in TGF-β1 and control groups, respectively. A total of 78 miRNAs with significant differential expression, including 39 up-regulated miRNAs and 39 down-regulated miRNAs were identified in the TGF-β1 group compared with the control. Real-time quantitative PCR analyses of bta-miR-106a and bta-miR-1434-5p showed that their up-expressions were interrupted by SB431542, an inhibitor that blocks TGFβ1/Smad signaling, which supported the sequencing data. GO analysis showed involvement of the predicted genes of the differentially expressed miRNAs in a broad spectrum of cell biological processes, cell components, and molecular functions. KEGG pathway analysis of the predicted miRNA targets further indicated that these differentially expressed miRNAs are involved in various signaling pathways, such as Wnt, MAPK, and TGF-β signaling, which might be involved in follicular development. These results provide valuable information on the composition, expression, and function of miRNAs in bovine granulosa cells responding to TGF-β1, and will aid in understanding the molecular mechanisms of TGF-β1 in granulosa cells.

Keywords: Bovine granulosa cells; High-throughput sequencing; MicroRNA; TGF-β1.

MeSH terms

Animals
Cattle
Cell Proliferation / drug effects
Cells, Cultured
Female
Gene Expression Regulation / drug effects
Gene Regulatory Networks
Granulosa Cells / cytology*
Granulosa Cells / drug effects
High-Throughput Nucleotide Sequencing / methods*
High-Throughput Nucleotide Sequencing / veterinary
Humans
MicroRNAs / genetics*
Sequence Analysis, RNA / methods*
Sequence Analysis, RNA / veterinary
Signal Transduction
Transforming Growth Factor beta1 / pharmacology*

Substances

MicroRNAs
TGFB1 protein, human
Transforming Growth Factor beta1