Background: Follistatin (FST) is a secretory glycoprotein and belongs to the TGF-β superfamily. Previously, we found that two single nucleotide polymorphisms (SNPs) of sheep FST gene were significantly associated with wool quality traits in Chinese Merino sheep (Junken type), indicating that FST is involved in the regulation of hair follicle development and hair trait formation. The transcription regulation of human and mouse FST genes has been widely investigated, and many transcription factors have been identified to regulate FST gene, such as erythroid 2-related factor 2 (Nrf2), Estrogen-related receptor-β (ERRβ), β-catenin/transcription factor 4 (TCF4) and transcription factor Sp1. However, to date, the transcriptional regulation of sheep FST is largely unknown. The objective of this study was to investigate the transcriptional regulation of sheep FST gene in hair follicles.
Results: Genome walking analysis revealed that the gap region upstream of sheep genomic FST gene was 775 bp long. Transcription factor binding site analysis showed sheep FST promoter region contained a conserved putative binding site for signal transducer and activator of transcription 3 (STAT3), located at nucleotides -423 to -416 relative to the first nucleotide (A, +1) of the initiation codon (ATG). The dual-luciferase reporter assays showed that STAT3 inhibited the activity of the FST promoter reporter, and the mutation of the putative STAT3 binding site attenuated the inhibitory effect of STAT3 on the FST promoter activity. Furthermore, chromatin immunoprecipitation assay (ChIP) indicated that STAT3 directly binds to the FST promoter. The further functional study displayed that FST and STAT3 played opposite roles in cell proliferation. Overexpression of FST significantly promoted the proliferation of sheep fetal fibroblasts (SFFs) and human keratinocyte (HaCaT) cells, and overexpression of STAT3 significantly inhibited the proliferation of SFFs and HaCaT cells, which was accompanied by a significantly reduced expression of FST gene (P < 0.05).
Conclusions: STAT3 directly negatively regulates sheep FST gene and inhibits cell proliferation. The findings will contribute to understanding molecular mechanisms that underlie hair follicle development and wool trait formation.