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Mammary transcriptome analysis of lactating dairy cows

ABSTRACT: Growth hormone (GH) has a well established galactopoietic effect on ruminant lactation, however the molecular mechanisms that mediate these effects are not fully understood. This study aimed to define the effects of GH on the production of milk protein by the bovine mammary gland and to establish the potential molecular mechanisms mediating these effects. Elevated levels of GH increased milk yield with corresponding increases in protein, fat and lactose yield, while milk composition remained unaffected by treatment. Increased yield (but not concentration) of all the individual milk proteins was not accompanied by changes in the levels of the milk protein transcripts, with the exception of S2-casein, suggesting post-transcriptional regulation of the milk protein genes is a main factor in the GH-mediated effects on bovine lactation. There were significant transcriptional changes in a wide range of pathways including cell signaling, cell death, cell growth and proliferation, cell cycle regulation and metabolism, consistent with the broad effects of GH. Differential expression of genes involved in the PI3K, ERK/MAPK, insulin receptor, JAK/STAT and IGF-1 signaling pathways were also evident in response to GH treatment supporting potential cross-talk between the GH and insulin signaling pathways in the bovine mammary gland. Transcript levels of the SHC1 gene were elevated in GH-treated cows. SHC1 activates many pathways including those associated with translational regulation and gene transcription and has been implicated in mediating cross-talk between the insulin and GH signaling pathways, providing some insights into the potential pathways mediating the effects of GH on milk protein synthesis in the bovine mammary gland.

ORGANISM(S): Bos taurus

PROVIDER: GSE11571 | GEO | 2009/09/04

SECONDARY ACCESSION(S): PRJNA106193

REPOSITORIES: GEO

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Project description:The objective of this study was to determine the effect of the DGAT1 K232A polymorphism on the global mRNA expression pattern of genes in the mammary gland tissue of grazing dairy cows in order to get more insight into the effects of this polymorphism on the physiology of the mammary glandgland of grazing dairy cows. Microarray analysis was used to identify genes whose expression was affected by the DGAT1 polymorphism in the mammary gland biopsies of 9 A232A cows, 13 K232A cows, and 4 K232K cows. The Microarray Analysis of Variance (MAANOVA) and Factor Analysis for Multiple Testing method (FAMT) were used to associate the expression of the genes present on Affymettrix Bovine Genome Arrays with the DGAT1 polymorphism. The data was also analysed at the level of functional modules by gene set enrichment analysis. In this experimental setting, DGAT1 polymorphism did not modify milk yield and composition significantly, although changes occurred in the yields of C14:0, C16:1cis-9, and some long chain fatty acids in milk. The DGAT1 polymorphism resulted in 30 differentially expressed genes related to cell growth, proliferation and development, signalling, remodelling and immune system. At the functional level, the pathways most affected by DGAT1 polymorphism were related to lipid biosynthesis, which likely reflected counter mechanisms of mammary tissue to respond to changes in milk FA composition, signalling, as well as immune system responses. A total of 28 Holstein-Friesian dairy cows in mid-lactation (DIM; 153 M-BM-1 32.8 days), milk yield (25.7 M-BM-1 3.08 kg/d) and fat content (4.3 M-BM-1 0.12%) were used in the study. Two consecutive milk samples (a.m. and p.m. milking) were obtained and pooled. One aliquot was stored at 4M-BM-0C until analysis of fat, protein and lactose percentage, and another aliquot was frozen at -20M-BM-0C until analysis for FA composition by gas chromatography. Specific details regarding the analysis of FA in milk are presented in Mach et al. (2011). Approximately 750 to 1,000 mg of mammary tissue from each cow was obtained by surgical biopsy. One part of the tissue was used for isolation of DNA and the other for extraction of the RNA. Genotyping of the DGAT1 polymorphism was performed using a TaqMan allelic discrimination method in an Applied Biosystems 7500 Real-Time PCR System (Applied Biosystems) as described by Schennink et al. (2007). The genotype at the DGAT1 locus was designated KK, KA, or AA for homozygous Lysine, heterozygous Lysine/Alanine, or homozygous Alanine, respectively. Microarray analysis was used to identify genes whose expression was affected by the DGAT1 polymorphism in the mammary gland biopsies of 11 A232A cows, 13 K232A cows, and 4 K232K cows. Total RNA from mammary gland tissue (50-100 mg) was isolated using TRIzol reagent (Invitrogen, Breda, The Netherlands), following the manufacturerM-bM-^@M-^Ys instructions. The RNA purity and concentrations were determined using a NanoDrop ND-1000 spectrophotometer (Isogen, Maarssen, the Netherlands), and the RNA quality was assessed using the BioAnalyzer 2100 (Agilent Technologies, Amsterdam, the Netherlands). The RNA of each biopsy was amplified, biotin-labeled, and hybridized to single-dye Affymetrix GeneChipM-BM-. Bovine Genome Array (#900493) by ServiceXS (Leiden, the Netherlands)

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Mammary transcriptome analysis of lactating dairy cows

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