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Published on: December 28, 2021
MicroRNA-449a Suppresses Mouse Spermatogonia Proliferation via Inhibition of CEP55
Rui Hua1, Qing-Jun Chu1, Yao Zhou1
1Reproductive Medicine Center, Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, China.
Abstract:
At present, infertile patients with maturation arrest (MA) are difficult to obtain mature sperm. Spermatogenesis and its molecular mechanism are still not clear. Patients with MA and normal spermatogenesis (NS) were collected. iTRAQ-based proteomic approach was performed to reveal the different proteins between them. To validate the confidence of proteome data, the individual samples were analyzed by Western blotting (WB), quantitative polymerase chain reaction (qPCR), and immunofluorescence. The miR-449a and CEP55 were determined by Luciferase assay. Mouse GC-1 cells were transfected with CEP55 siRNAs, miR-449a mimic, or inhibitor, and cell proliferation was determined. Compared with NS, 27 proteins were differentially expressed in MA, and CEP55 protein was the most significant difference. WB and qPCR showed that CEP55 levels were significantly elevated in NS than MA. In transfected cells, overexpression of miR-449a and knockdown of CEP55 both downregulated CEP55 expression and decreased cell proliferation. miR-449a suppresses mouse spermatogonia proliferation via inhibition of CEP55.
Insights
Maturation arrest (MA) in infertile men lacks mature sperm due to unclear molecular mechanisms. Researchers found miR-449a suppresses spermatogonia proliferation by inhibiting CEP55, offering new insights into male infertility.
Area of Science:
- Reproductive Biology
- Molecular Endocrinology
- Proteomics
Background:
- Male infertility associated with maturation arrest (MA) presents a significant clinical challenge, with the underlying molecular mechanisms of spermatogenesis remaining largely unknown.
- Identifying molecular differences between MA and normal spermatogenesis (NS) is crucial for understanding disease pathogenesis and developing therapeutic strategies.
Purpose of the Study:
- To investigate the proteomic differences between infertile patients with maturation arrest (MA) and those with normal spermatogenesis (NS).
- To elucidate the molecular mechanism by which microRNA-449a (miR-449a) affects spermatogonia proliferation, focusing on its interaction with Centrosome-associated protein 55 (CEP55).
Main Methods:
- iTRAQ-based proteomic analysis was employed to identify differentially expressed proteins between MA and NS patient groups.
- Western blotting (WB), quantitative polymerase chain reaction (qPCR), immunofluorescence, and Luciferase assays were used for validation and mechanistic studies.
- In vitro experiments involved transfecting mouse GC-1 cells with CEP55 siRNAs, miR-449a mimics, or inhibitors to assess effects on cell proliferation.
Main Results:
- Proteomic analysis revealed 27 differentially expressed proteins between MA and NS, with CEP55 showing the most significant difference.
- WB and qPCR confirmed significantly elevated CEP55 levels in NS compared to MA.
- Overexpression of miR-449a and knockdown of CEP55 in mouse spermatogonia cells led to decreased CEP55 expression and reduced cell proliferation.
Conclusions:
- CEP55 is a key protein with significantly different expression levels in maturation arrest compared to normal spermatogenesis.
- miR-449a plays a suppressive role in mouse spermatogonia proliferation through the inhibition of CEP55.
- These findings provide novel molecular insights into the pathogenesis of maturation arrest and suggest miR-449a/CEP55 as potential targets for male infertility treatment.
