Long-chain acyl-CoA synthetase 1 (ACSL1) plays an important role in fatty acid metabolism and fat deposition. The transcription of the ACSL1 gene is regulated specifically among cells and physiological processes, and transcriptional regulation of ACSL1 in adipogenesis remains elusive. Here, we characterize transcription factors (TFs) associated with adipogenesis in the porcine ACSL1 gene. CCAAT-enhancer binding protein (C/EBP)α, a well-known adipogenic marker, was found to enhance the expression of the ACSL1 gene via binding two tandem motifs in the promoter. Further, we demonstrate that ACSL1 mediates C/EBPα effects on adipogenesis in preadipocytes cultured from subcutaneous fat tissue of pigs via gain- and loss-of-function analyses. The cAMP-response element binding protein, another TF involved in adipogenesis, was also identified in the regulation of ACSL1 gene expression. Additionally, single nucleotide polymorphisms (SNPs) were screened in the promoter of ACSL1 among four breeds including the Chinese indigenous Min, and Duroc, Berkshire, and Yorkshire pigs through sequencing of PCR products. Two tightly linked SNPs, −517G>T and −311T>G, were found exclusively in Min pigs. The haplotype mutation decreases promoter activity in PK-15 and ST cells, and in vivo the expression of ACSL1, illustrating a possible role in adipogenesis regulated by C/EBPα/ACSL1 axis. Additionally, a total of 24 alternative splicing transcripts were identified, indicating the complexity of alternative splicing in the ACSL1 gene. The results will contribute to further revealing the regulatory mechanisms of ACSL1 during adipogenesis and to the characterization of molecular markers for selection of fat deposition in pigs.

Transcription factor Homeobox C8 (HOXC8) is identified as a white adipose gene as revealed by expression profile analysis in fat tissues. However, the specific role of HOXC8 in fat accumulation remains to be identified. This study was designed to reveal the effects of HOXC8 on preadipocyte proliferation and differentiation. We first make clear that the expression of HOXC8 is associated with fat contents in muscles, highlighting a role of HOXC8 in fat accumulation. Next, it is demonstrated that HOXC8 promotes the proliferation and differentiation of preadipocytes through gain- and loss-of-function assays in primary cultured porcine preadipocytes. Then, mechanisms underlying the regulation of HOXC8 on preadipocyte proliferation and differentiation are identified with RNA sequencing, and a number of differentially expressed genes (DEGs) in response to HOXC8 knockdown are identified. The top GO (Gene Ontology) terms enriched by DEGs involved in proliferation and differentiation, respectively, are identical. IL-17 signaling pathway is the common one significantly enriched by DEGs involved in preadipocyte proliferation and differentiation, respectively, indicating its importance in mediating fat accumulation regulated by HOXC8. Additionally, we find that the inhibition of proliferation is one of the main processes during preadipocyte differentiation. The results will contribue to further revealing the mechanisms underlying fat accumulation regulated by HOXC8.