Screening for phenotypic outliers identifies an unusually low concentration of a β-lactoglobulin B protein isoform in bovine milk caused by a synonymous SNP

• Published in: Genetics selection evolution (Paris) 2022-03, Vol.54 (1), p.22-22, Article 22
• Main Authors: Davis, Stephen R, Ward, Hamish E, Kelly, Van, Palmer, David, Ankersmit-Udy, Alexandra E, Lopdell, Thomas J, Berry, Sarah D, Littlejohn, Mathew D, Tiplady, Kathryn, Adams, Linda F, Carnie, Katie, Burrett, Alayna, Thomas, Natalie, Snell, Russell G, Spelman, Richard J, Lehnert, Klaus
• Format: Article
• Language: English
• Subjects: Animals; Cattle - genetics; Female; Genome-Wide Association Study; Humans; Lactoglobulins - analysis; Lactoglobulins - genetics; Lactoglobulins - metabolism; Life Sciences; Milk - chemistry; Polymorphism, Single Nucleotide; Protein Isoforms - analysis; Protein Isoforms - genetics; Protein Isoforms - metabolism
• ISSN: 1297-9686; 0999-193X; 1297-9686
• DOI: 10.1186/s12711-022-00711-z

Milk samples from 10,641 dairy cattle were screened by a mass spectrometry method for extreme concentrations of the A or B isoforms of the whey protein, β-lactoglobulin (BLG), to identify causative genetic variation driving changes in BLG concentration. A cohort of cows, from a single sire family, was identified that produced milk containing a low concentration of the BLG B protein isoform. A genome-wide association study (GWAS) of BLG B protein isoform concentration in milk from AB heterozygous cows, detected a group of highly significant single nucleotide polymorphisms (SNPs) within or close to the BLG gene. Among these was a synonymous G/A variation at position + 78 bp in exon 1 of the BLG gene (chr11:103256256G > A). The effect of the A allele of this SNP (which we named B') on BLG expression was evaluated in a luciferase reporter assay in transfected CHO-K1 and MCF-7 cells. In both cell types, the presence of the B' allele in a plasmid containing the bovine BLG gene from -922 to + 898 bp (relative to the transcription initiation site) resulted in a 60% relative reduction in mRNA expression, compared to the plasmid containing the wild-type B sequence allele. Examination of a mammary RNAseq dataset (n = 391) identified 14 heterozygous carriers of the B' allele which were homozygous for the BLG B protein isoform (BB'). The level of expression of the BLG B' allele was 41.9 ± 1.0% of that of the wild-type BLG B allele. Milk samples from three cows, homozygous for the A allele at chr11:103,256,256 (B'B'), were analysed (HPLC) and showed BLG concentrations of 1.04, 1.26 and 1.83 g/L relative to a mean of 4.84 g/L in milk from 16 herd contemporaries of mixed (A and B) BLG genotypes. The mechanism by which B' downregulates milk BLG concentration remains to be determined. High-throughput screening and identification of outliers, enabled the discovery of a synonymous G > A mutation in exon 1 of the B allele of the BLG gene (B'), which reduced the milk concentration of β-lactoglobulin B protein isoform, by more than 50%. Milk from cows carrying the B' allele is expected to have improved processing characteristics, particularly for cheese-making.