NCAPG Regulates Myogenesis in Sheep, and SNPs Located in Its Putative Promoter Region Are Associated with Growth and Development Traits

Zehu Yuan; Ling Ge; Pengwei Su; Yifei Gu; Weihao Chen; Xiukai Cao; Shanhe Wang; Xiaoyang Lv; Tesfaye Getachew; Joram M Mwacharo; Aynalem Haile; Wei Sun

doi:10.3390/ani13203173

NCAPG Regulates Myogenesis in Sheep, and SNPs Located in Its Putative Promoter Region Are Associated with Growth and Development Traits

Animals (Basel). 2023 Oct 11;13(20):3173. doi: 10.3390/ani13203173.

Authors

Zehu Yuan^{1

2}, Ling Ge^{1

2

3}, Pengwei Su^{1

2

3}, Yifei Gu^{1

2

3}, Weihao Chen^{1

2

3}, Xiukai Cao^{1

2}, Shanhe Wang^{1

2

3}, Xiaoyang Lv^{1

2}, Tesfaye Getachew⁴, Joram M Mwacharo⁴, Aynalem Haile⁴, Wei Sun^{1

2

3

5}

Affiliations

¹ Joint International Research Laboratory of Agriculture and Agri-Product Safety of Ministry of Education of China, Yangzhou University, Yangzhou 225009, China.
² International Joint Research Laboratory in Universities of Jiangsu Province of China for Domestic Animal Germplasm Resources and Genetic Improvement, Yangzhou University, Yangzhou 225009, China.
³ College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China.
⁴ International Centre for Agricultural Research in the Dry Areas, Addis Ababa 999047, Ethiopia.
⁵ "Innovative China" "Belt and Road" International Agricultural Technology Innovation Institute for Evaluation, Protection, and Improvement on Sheep Genetic Resource, Yangzhou 225009, China.

Abstract

Previously, NCAPG was identified as a candidate gene associated with sheep growth traits. This study aimed to investigate the direct role of NCAPG in regulating myogenesis in embryonic myoblast cells and to investigate the association between single-nucleotide polymorphisms (SNPs) in its promoter region and sheep growth traits. The function of NCAPG in myoblast proliferation and differentiation was detected after small interfering RNAs (siRNAs) knocked down the expression of NCAPG. Cell proliferation was detected using CCK-8 assay, EdU proliferation assay, and flow cytometry cell cycle analysis. Cell differentiation was detected via cell immunofluorescence and the quantification of myogenic regulatory factors (MRFs). SNPs in the promoter region were detected using Sanger sequencing and genotyped using the improved multiplex ligation detection reaction (iMLDR^®) technique. As a result, a notable decrease (p < 0.01) in the percentage of EdU-positive cells in the siRNA-694-treated group was observed. A significant decrease (p < 0.01) in cell viability after treatment with siRNA-694 for 48 h and 72 h was detected using the CCK-8 method. The quantity of S-phase cells in the siRNA-694 treatment group was significantly decreased (p < 0.01). After interfering with NCAPG in myoblasts during induced differentiation, the relative expression levels of MRFs were markedly (p < 0.05 or p < 0.01) reduced compared with the control group on days 5-7. The myoblast differentiation in the siRNA-694 treatment group was obviously suppressed compared with the control group. SNP1, SNP2, SNP3, and SNP4 were significantly (p < 0.05) associated with all traits except body weight measured at birth and one month of age. SNP5 was significantly (p < 0.05) associated with body weight, body height, and body length in six-month-old sheep. In conclusion, interfering with NCAPG can inhibit the proliferation and differentiation of ovine embryonic myoblasts. SNPs in its promoter region can serve as potential useful markers for selecting sheep growth traits.

Keywords: NCAPG; SNP; myoblasts; sheep growth and development.

Abstract

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