Let-7g induces granulosa cell apoptosis by targeting MAP3K1 in the porcine ovary

Rui Cao; Wangjun Wu; Xiaolong Zhou; Kaiqing Liu; Bojiang Li; Xianju Huang; Yu Zhang; Honglin Liu

doi:10.1016/j.biocel.2015.08.011

Let-7g induces granulosa cell apoptosis by targeting MAP3K1 in the porcine ovary

Int J Biochem Cell Biol. 2015 Nov:68:148-57. doi: 10.1016/j.biocel.2015.08.011. Epub 2015 Aug 20.

Authors

Rui Cao¹, Wangjun Wu¹, Xiaolong Zhou¹, Kaiqing Liu¹, Bojiang Li¹, Xianju Huang¹, Yu Zhang¹, Honglin Liu²

Affiliations

¹ Department of Animal Genetics, Breeding and Reproduction, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, 210095, China.
² Department of Animal Genetics, Breeding and Reproduction, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, 210095, China. Electronic address: [email protected].

PMID: 26299328
DOI: 10.1016/j.biocel.2015.08.011

Abstract

Follicular atresia mainly results from apoptosis of granulosa cells (GCs). Our previous microRNA array data indicated that the miRNA let-7g level increases significantly during porcine ovary follicular atresia. It is uncertain if GCs apoptosis is mediated by microRNA let-7g. In this study, the expression levels of the apoptosis-associated genes CASP3, BAX and BIM were significantly upregulated when let-7g mimic was transfected into porcine GCs, and the anti-apoptotic genes BCL-2 and MCL-1 were significantly downregulated. The apoptosis rate was measured by flow cytometry, and our results indicated that let-7g significantly enhanced GCs apoptosis. In further studies, we found that overexpression of let-7g induced the expression of FoxO1 in GCs and led to nuclear accumulation of dephosphorylated FoxO1. In addition, the effect of let-7g on FoxO1 expression and dephosphorylation resulted from repression of the expression of the MAP3K1 gene in porcine GCs. The site on MAP3K1 mRNA targeted by let-7g was confirmed by luciferase reporter assay. The anti-apoptotic effect of MAP3K1 was validated by silencing MAP3K1 using small interfering RNA technology. In conclusion, our data indicate that let-7g induces porcine GCs apoptosis by inhibiting the MAP3K1 gene, which promotes FoxO1 expression and dephosphorylation with nuclear accumulation.

Keywords: Apoptosis; FoxO1; Granulosa cell; Let-7g; MAP3K1.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Active Transport, Cell Nucleus
Animals
Apoptosis
Apoptosis Regulatory Proteins / genetics
Apoptosis Regulatory Proteins / metabolism
Base Sequence
Bcl-2-Like Protein 11
Binding Sites
Caspase 3 / genetics
Caspase 3 / metabolism
Female
Follicular Atresia / genetics*
Follicular Atresia / metabolism
Forkhead Transcription Factors / genetics*
Forkhead Transcription Factors / metabolism
Gene Expression Regulation
Granulosa Cells / cytology
Granulosa Cells / metabolism*
MAP Kinase Kinase Kinase 1 / antagonists & inhibitors
MAP Kinase Kinase Kinase 1 / genetics*
MAP Kinase Kinase Kinase 1 / metabolism
Membrane Proteins / genetics
Membrane Proteins / metabolism
MicroRNAs / antagonists & inhibitors
MicroRNAs / genetics*
MicroRNAs / metabolism
Molecular Sequence Data
Myeloid Cell Leukemia Sequence 1 Protein / genetics
Myeloid Cell Leukemia Sequence 1 Protein / metabolism
Oligonucleotides, Antisense / genetics
Oligonucleotides, Antisense / metabolism
Primary Cell Culture
Protein Binding
Proto-Oncogene Proteins / genetics
Proto-Oncogene Proteins / metabolism
RNA, Messenger / genetics*
RNA, Messenger / metabolism
RNA, Small Interfering / genetics
RNA, Small Interfering / metabolism
Signal Transduction
Swine
bcl-2-Associated X Protein / genetics
bcl-2-Associated X Protein / metabolism

Substances

Apoptosis Regulatory Proteins
Bcl-2-Like Protein 11
Forkhead Transcription Factors
Membrane Proteins
MicroRNAs
Myeloid Cell Leukemia Sequence 1 Protein
Oligonucleotides, Antisense
Proto-Oncogene Proteins
RNA, Messenger
RNA, Small Interfering
bcl-2-Associated X Protein
MAP Kinase Kinase Kinase 1
Caspase 3