Multiple promoters and targeted micro RNA s direct the expressions of HMGB 3 gene transcripts in dairy cattle

Summary HMGB 3 (high‐mobility group box 3) is an X ‐linked member of a family of sequence‐independent chromatin‐binding proteins and functions as a universal sentinel for nucleic acid–mediated innate immune responses. The splice variant expression, promoter characterization and targeted micro RNA s of the bovine HMGB 3 gene were investigated to explore its expression pattern and possible regulatory mechanism. The results revealed that the expression of HMGB 3 transcript variants 1 and 2 ( HMGB 3‐ TV 1 and HMGB 3 ‐ TV 2) mRNA in the mastitis‐infected mammary gland tissues was up‐regulated by 8.46‐ and 5.31‐fold respectively compared with that in healthy tissues ( P  < 0.05). HMGB 3 ‐ TV 1 was highly expressed in the mammary gland tissues, whereas HMGB 3 ‐ TV 2 was expressed primarily in liver. Functional analyses indicated that HMGB 3 transcription is regulated by three distinct promoters – promoters 1, 2 and 3 ( P 1, P 2 and P 3) – resulting in two alternative transcripts with the same 3′‐untranslated region. Promoter luciferase activity analysis suggested that the core sequences of P 1 and P 2 were mapped in the region of g.1535 to ~g.2076 and g.2074 to ~g.2491 respectively. The g.5880 C > T SNP in P 3 affected its base promoter activity, and different genotypes were associated with the bovine somatic count score. The expression of targets bovine mi R ‐17‐5p , mi R ‐20b and mi R ‐93 of the HMGB 3 gene was down‐regulated 1.56‐, 1.72‐ and 2.94‐fold respectively in mammary gland tissues as compared with that in healthy tissues ( P  <   0.05). The findings suggest that HMGB 3 expression is under complex transcriptional and post‐transcriptional control by alternate promoter usage, alternative splicing mechanism and micro RNA s in dairy cattle.