| The Shanbei White Cashmere goat is a locally bred breed that is suitable for both meat and cashmere production.This breed has excellent genetic stability,fine cashmere fiber diameter,and strong stress resistance.The fiber（cashmere）produced by the sondary hair follicles of its skin has high economic value.With changes in feeding methods and breeding environments,the cashmere production and fiber diameter of Shanbei White Cashmere goat has decreased,reducing the advantage of this breed.Therefore,improving cashmere production and quality has become an urgent problem in the cashmere goat industry.The SIRT7 gene has the ability to promote r RNA expression,help DNA damage repair,and regulate the periodic changes in hair follicle stem cells（HFSCs）.Studies have shown that the degradation of a transcription factor,NFATc1,mediated by the SIRT7 gene is one of the mechanisms for hair follicles to transition from the resting phase to the growth phase.In addition,compared with wild-type mice,overexpression of the SIRT7 gene in mice significantly improved hair follicle density and hair regeneration.Therefore,this study aimed to investigate the effect of SIRT7 gene overexpression on hair follicle development in Shanbei White Cashmere goat,with the aim of providing a theoretical basis for using molecular breeding methods to improve cashmere production and quality in this breed.In this study,Shanbei White Cashmere g oats and their HFSCs were used as research objects,and a pc DNA3.1-SIRT7 overexpression vector was constructed and packaged into an overexpression adenovirus to overexpress the SIRT7 gene in HFSCs and the skin of goats.The proliferation of HFSCs and the growth of cashmere were analyzed.The main results obtained from this study are as follows:（1）Optimization and identification of specific promoter for goat HFSCsDifferent truncated forms of the goat K15 gene promoter were transfected into goat HFSCs using a dual luciferase reporter vector,and the goat K15 gene core promoter region with a length of 715 bp was screened after relative luciferase activity detection.Subsequently,the activity of the screened core promoter was tested in goat HFSCs,Dermal papilla cells（DPCs）,and Fibroblasts（FBs）,and the results showed that the screened core promoter can specifically express in goat HFSCs.（2）The effect of SIRT7 gene overexpression on goat HFSCsA pc DNA3.1-SIRT7 overexpression vector was constructed and transfected into goat HFSCs.CCK8 and EDU assays were employed to investigate the effects of SIRT7 gene overexpression on the proliferation of Shaanbei Cashmere Goat HFSCs.The results showed that SIRT7 gene overexpression significantly enhanced the proliferation capacity of HFSCs（P<0.05）.Transcriptome sequencing analysis was performed on the SIRT7 gene overexpression group and the control group,revealing 208 differentially expressed genes.By combining the GO and KEGG metabolic pathways related to cell proliferation,18 differentially expressed genes associated with cell proliferation were further enriched.Network interaction analysis identified six candidate genes that affect the proliferation of goat HFSCs:STAT2,IL6,FGF10,IRS2,MYC,CDKN2A,and HDAC1.（3）In vivo injection of AAV to study the effect of SIRT7 gene on goat hair follicle developmentBy analyzing frozen stions of different virus serotypes,a broad-spectrum AAV-9 serotype suitable for goat skin was selected.The core promoter region of the goat K15 gene and the coding region of the SIRT7 gene were constructed into the AAV-9 vector by homologous recombination,and then packaged into adenoviruses.After virus packaging,the virus titer was determined,and the optimal virus injection dose for goat skin was determined to be 5×1011 vg.Three Shanbei White Cashmere goats were selected,and virus injection was performed on their back skin.At the 6th week after virus injection,samples of the hair-like and skin tissue at each injection site were collected for measurement of hair and down length and down fineness.The expression of SIRT7 protein in skin tissue was determined by protein immunoblotting and immunofluorescence staining.The results showed that overexpression of SIRT7 gene significantly increased the length of goat down（P<0.05）,but had no significant effect on the length and fineness of goat hair.At the same time,protein immunoblotting and immunofluorescence results showed that virus injection significantly increased the expression of SIRT7 protein in skin tissue（P<0.05）. |
