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. 2021 Sep 22;10(10):949.
doi: 10.3390/biology10100949.

Bta-miR-2400 Targets SUMO1 to Affect Yak Preadipocytes Proliferation and Differentiation

Yongfeng Zhang  1   2 , Lanhua Ma  1 , Yarong Gu  1 , Yongfang Chang  1 , Chunnian Liang  1 , Xian Guo  1 , Pengjia Bao  1 , Min Chu  1 , Xuezhi Ding  1 , Ping Yan  1
Affiliations

Bta-miR-2400 Targets SUMO1 to Affect Yak Preadipocytes Proliferation and Differentiation

Yongfeng Zhang et al. Biology (Basel). .

Abstract

Yak adipose tissue may have evolved a unique energy metabolism manner to accommodate the organism's seasonal growth rhythms. MiRNAs regulate multiple biological processes including systemic metabolism and energy homeostasis through post-transcriptional regulations. Rare reports have shown that miRNAs regulate lipid metabolism in domestic yaks. Therefore, we investigated the regulatory mechanisms of bta-miR-2400 in modulating yak preadipocytes proliferation and differentiation. We found that bta-miR-2400 was highly expressed in adipose tissue. Overexpression of bta-miR-2400 in yak preadipocytes significantly enhanced cell proliferation, increased the number of EdU fluorescence-stained cells, and promoted the expression of proliferation marker genes (CDK2, CDK4 and PCNA). Besides, overexpression of bta-miR-2400 repressed the expression of adipogenesis-related marker genes, and the content of cellular triglyceride was substantially reduced. Conversely, inhibition of bta-miR-2400 showed opposite effects compared to those of bta-miR-2400 overexpression in yak preadipocytes. Further, luciferase reporter assays revealed that SUMO1 is a target gene of bta-miR-2400, with bta-miR-2400 being able to down-regulate SUMO1 mRNA and protein expression. In conclusion, bta-miR-2400 regulates lipid metabolism and energy homeostasis in yak preadipocytes by directly targeting SUMO1 to promote cell proliferation and inhibit differentiation.

Keywords: SUMO1; bta-miR-2400; differentiation; proliferation; yak adipocyte.

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Conflict of interest statement

The authors have no conflict of interest.

Figures

Figure 1
Figure 1
Expression profiles of bta-miR-2400 in tissues and adipocytes. (A) The bta-miR-2400 expression in different yak tissues and (B) different adipose tissue stages. (C) The expression pattern of bta-miR-2400 during yak preadipocyte differentiation. * p < 0.05. Duncan’s Multiple Range Test was used for multiple comparisons, different lowercase letters represent significant differences (p < 0.05).
Figure 2
Figure 2
Bta-miR-2400 promotes yak preadipocyte proliferation. Cells were transfected with bta-miR-2400 mimic, NC, and inhibitor. (A) Cell Counting kit 8 (CCK-8) assays and (B,C) 5-ethynyl-20-deoxyuridine (EdU) proliferation were performed to measure cell proliferation (n = 6 per treatment per time point, magnification 200×); (D) Relative expression of cyclin-dependent kinases 2 (CDK2), cyclin-dependent kinases 4 (CDK4) and proliferating cell nuclear antigen (PCNA). * p < 0.05; ** p < 0.01.
Figure 3
Figure 3
Bta-miR-2400 inhibits yak preadipocyte differentiation. Yak preadipocytes were transfected with bta-miR-2400 mimics, inhibitors or negative control (NC) for 10 days after induced differentiation. (A) Adipocytes were stained with Oil red O. (B) The cellular triglyceride content was analyzed by spectrophotometer. (C) Expression levels of adipogenic markers (PPARγ, FABP4, and C/EBPα). (D,E) The protein level of PPARγ in adipocytes transfected with bta-miR-2400 mimics, inhibitors and negative control. The expression of genes related to (F) fatty acid transportation (DGAT1, CPT1), (G) fatty acid synthesis (ACC, SCD and FAS), and (H) lipogenic transcription (ELVOL, SREBPF1). * p < 0.05; ** p < 0.01.
Figure 4
Figure 4
Bta-miR-2400 targets the 3′UTR of SUMO1. (A) The schematic of the sequence alignment between bta-miR-2400 and 3′UTR of SUMO1. (B) The expression pattern of bta-miR-2400 during yak adipocyte differentiation. (C) Luciferase reporter assay for bta-miR-2400 and 3′UTR of SUMO1. (D–F) Expression of SUMO1 mRNA and protein for cells transfected with bta-miR-2400 mimics, inhibitors and negative control. * p < 0.05; ** p < 0.01. Different lowercase letters represent significant differences (p < 0.05).
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