title: Role of an SNP in Alternative Splicing of Bovine NCF4 and Mastitis Susceptibility.
abstract: Neutrophil cytosolic factor 4 (NCF4) is component of the nicotinamide dinucleotide phosphate oxidase complex, a key factor in biochemical pathways and innate immune responses. In this study, splice variants and functional single-nucleotide polymorphism (SNP) of NCF4 were identified to determine the variability and association of the gene with susceptibility to bovine mastitis characterized by inflammation. A novel splice variant, designated as NCF4-TV and characterized by the retention of a 48 bp sequence in intron 9, was detected in the mammary gland tissues of infected cows. The expression of the NCF4-reference main transcript in the mastitic mammary tissues was higher than that in normal tissues. A novel SNP, g.18174 A>G, was also found in the retained 48 bp region of intron 9. To determine whether NCF4-TV could be due to the g.18174 A>G mutation, we constructed two mini-gene expression vectors with the wild-type or mutant NCF4 g.18174 A>G fragment. The vectors were then transiently transfected into 293T cells, and alternative splicing of NCF4 was analyzed by reverse transcription-PCR and sequencing. Mini-gene splicing assay demonstrated that the aberrantly spliced NCF4-TV with 48 bp retained fragment in intron 9 could be due to g.18174 A>G, which was associated with milk somatic count score and increased risk of mastitis infection in cows. NCF4 expression was also regulated by alternative splicing. This study proposes that NCF4 splice variants generated by functional SNP are important risk factors for mastitis susceptibility in dairy cows.
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, 250131, China.
authors.forename: Zhihua
authors.initials: Z
authors.lastname: Ju
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, 250131, China.
authors.forename: Changfa
authors.initials: C
authors.lastname: Wang
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, 250131, China.
authors.forename: Xiuge
authors.initials: X
authors.lastname: Wang
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, 250131, China.
authors.forename: Chunhong
authors.initials: C
authors.lastname: Yang
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, 250131, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Sun
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, 250131, China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Jiang
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, 250131, China.
authors.forename: Fei
authors.initials: F
authors.lastname: Wang
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, 250131, China.
authors.forename: Mengjiao
authors.initials: M
authors.lastname: Li
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, 250131, China.
authors.forename: Jifeng
authors.initials: J
authors.lastname: Zhong
authors.affiliation: Dairy Cattle Research Center, Shandong Academy of Agricultural Sciences, Jinan, Shandong, 250131, China.
authors.forename: Jinming
authors.initials: J
authors.lastname: Huang
mesh_terms: D017398:Alternative Splicing / Q000235:genetics*; D000818:Animals; D002417:Cattle; D005260:Female; D020022:Genetic Predisposition to Disease; D008321:Mammary Glands, Animal / Q000378:metabolism; D008414:Mastitis, Bovine / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D010750:Phosphoproteins / Q000235:genetics* / Q000502:physiology; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*
chemical_list: D010750:Phosphoproteins; C085072:neutrophil cytosol factor 40K