Chi-miR-3031 regulates beta-casein via the PI3K/AKT-mTOR signaling pathway in goat mammary epithelial cells (GMECs)

Abstract

BackgroundMicroRNAs can regulate gene expression at the posttranscriptional level through translational repression or target degradation. Our previous investigations examined the differential expression levels of chi-miR-3031 in caprine mammary gland tissues in colostrum and common milk stages.ResultsThe present study detected the role of chi-miR-3031 in the lactation mechanisms of GMECs. High-throughput sequencing was used to analyze transcriptomic landscapes of GMECs transfected with chi-miR-3031 mimics (MC) and a mimic negative control (NC). In the MC and NC groups, we acquired 39,793,503 and 36,531,517 uniquely mapped reads, respectively, accounting for 85.85 and 81.66% of total reads. In the MC group, 180 differentially expressed unigenes were downregulated, whereas 157 unigenes were upregulated. KEGG pathway analyses showed that the prolactin, TNF and ErbB signaling pathways, including TGFα, PIK3R3, IGF2, ELF5, IGFBP5 and LHβ genes, played important roles in mammary development and milk secretion. Results from transcriptome sequencing, real-time PCR and western blotting showed that chi-miR-3031 suppressed the expression of IGFBP5 mRNA and protein. The expression levels of β-casein significantly increased in the MC and siRNA-IGFBP5 groups. We observed that the down-regulation of IGFBP5 activated mTOR at the Ser2448 site in GMECs transfected with MC and siRNA-IGFBP5. Previous findings and our results showed that chi-miR-3031 activated the PI3K-AKT-mTOR pathway and increased β-casein expression by down-regulating IGFBP5.ConclusionsThese findings will afford valuable information for improving milk quality and contribute the development of potential methods for amending lactation performance.Electronic supplementary materialThe online version of this article (10.1186/s12917-018-1695-6) contains supplementary material, which is available to authorized users.

Phosphoproteome Reveals the Lactation Mechanism in Caprine Mammary Gland Tissues in Peak and Late Lactation Stages

“…A total of 1,938 differentially modi ed modi cation sites of 1,172 proteins were found between PL and LL. The western blot result of phosphorylation levels of ACACA, EIF4EBP1, IRS1 and JUN showed that the results of phosphoproteome were reliable and these results were consistent with this studies [16,17]. Previous studies showed APOC3, FASN and ACACA proteins were highly correlated with the decomposition and utilisation of fat [18][19][20].…”

CircRNA8220 Sponges MiR-8516 to Regulate Cell Viability and Milk Synthesis via Ras/MEK/ERK and PI3K/AKT/mTOR Pathways in Goat Mammary Epithelial Cells

“…Human embryonic kidney cell line (HEK293T) was cultured in high-glucose DMEM (Dulbecco’s Modified Eagle Medium) supplemented with 10% fetal bovine serum, 100 μg/mL penicillin and 100 μg/mL streptomycin at 37 °C and 5% CO 2 to proliferate. The GMECs were purified and cultured depending on previous reports [ 18 ].…”

Epigenetics: New Insights into Mammary Gland Biology

“…Bioinformatics analysis suggested that miR-3031 and Igfbp5 are key signalling factors that regulate cell proliferation and protein synthesis in goat mammary epithelial cells. Chen et al [ 161 ], using miR-3031 mimics, showed that miR-3031 activated the PI3K-AKT-mTOR pathway and increased β-casein expression by downregulating Igfbp5 . In bovine mammary epithelium, Li et al [ 162 ] showed that the function of miR-486 is indispensable for regulating Phosphatase and tensin homologue deleted on chromosome ten ( Pten ), which reduces the differentiation of mammary epithelial cells [ 163 ].…”

“…Moreover, the overexpression of miR-152 leads to a strong decrease in DNMT1 expression, as well as a reduction in the global rate of 5-meC in mammary epithelial cells [ 170 ]. Studies describing miRNAs involved in the expression of milk protein genes, such as casein ( miR-3031 [ 161 ], miR-15 [ 159 ], miR-139 [ 130 ], miR-101a [ 127 ], miR-142-3p [ 131 ]), have highlighted the aforementioned genes as markers of the differentiation status of mammary epithelial cells (see before) rather than with the goal of understanding the role of miRNAs in their regulation. Indirectly, by showing that miR-24-3p regulates genes involved in the PI3K/Akt/mTOR and JAK/Stat5 pathways, which regulate milk protein synthesis, Qiaoqiao et al [ 156 ] highlighted the importance of this miRNA in this process.…”