Differences in the ribosomes prepared from lactating and non-lactating bovine mammary gland.
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ABSTRACT:
SUBMITTER: Herrington MD
PROVIDER: S-EPMC1176567 | biostudies-other | 1971 Jan
REPOSITORIES: biostudies-other
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Project description:BackgroundThe mammary gland is a dynamic organ that undergoes important physiological changes during reproductive cycles. Until now, data regarding the characterisation of miRNA in the mammary gland have been scarce and mainly focused on their abnormal expression in breast cancer. Our goal was to characterise the microRNA (miRNA) involved in mechanisms regulating the mammary function, with particular focus on the lactation stage.Methodology/principal findingsUsing high-throughput sequencing technology, the exhaustive repertoires of miRNA expressed (miRNome) in mouse and bovine mammary glands during established lactation were identified, characterized and compared. Furthermore, in order to obtain more information on miRNA loading in the RNA-induced silencing complex (RISC), the miRNome was compared with that obtained from RNA associated with the AGO2 protein (AGO2-miRNome) in mouse lactating mammary gland. This study enabled the identification of 164 and 167 miRNA in mouse and bovine, respectively. Among the 30 miRNA most highly expressed in each species, 24 were common to both species and six of them were preferentially highly expressed in lactating than non-lactating mammary gland. The potential functional roles of these 24 miRNA were deduced using DIANA-miRPath software, based on miRNA/mRNA interactions. Moreover, seven putative novel miRNA were identified. Using DAVID analysis, it was concluded that the predicted targets of two of these putative novel miRNA are involved in mammary gland morphogenesis.Conclusion/significanceOur study provides an overview of the characteristics of lactating mouse and bovine mammary gland miRNA expression profiles. Moreover, species-conserved miRNA involved in this fundamental biological function were identified. These miRNomes will now be used as references for further studies during which the impact of animal breeding on the miRNA expression will be analysed.
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Project description:Small-Tailed Han (STH) sheep are known for their high fecundity, but the survival of lambs is compromised and influences the commercial return from farming these sheep, with this being attributed in part to starvation from insufficient milk production by the ewes. In this study, the transcriptome profiles of the mammary gland of lactating and non-lactating STH ewes were investigated using paired-end RNA sequencing (RNA-Seq). An average of 14,447 genes were found to be expressed at peak-lactation in the STH sheep, while 15,146 genes were expressed in non-lactating ewes. A total of 4,003 differentially expressed genes (DEGs) were identified. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed that the DEGs were associated with a wide range of cellular components, biological processes and metabolic pathways, including binding activities, signaling pathways, cellular structures, and immune responses. The most highly expressed genes at peak-lactation included CSN2, LGB, LALBA, CSN1S1, CSN1S2, and CSN3, and the 10 most highly expressed genes accounted for 61.37% of the total Reads Per Kilobase of transcript, per Million mapped reads (RPKM). The most highly expressed genes in the mammary gland of non-lactating ewes included IgG, THYMB4X, EEF1A1, IgA, and APOE, and the 10 most highly expressed genes accounted for only 12.97% of the total gene RPKM values. This suggests that the sheep mammary gland undergoes a substantial development in milk protein synthesis infrastructure and promotion of protein transportation during lactation.
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Project description:The objective of this study was to examine five different sources of RNA, namely mammary gland tissue (MGT), milk somatic cells (SC), laser microdissected mammary epithelial cells (LCMEC), milk fat globules (MFG) and antibody-captured milk mammary epithelial cells (mMEC) to analyze the bovine mammary gland transcriptome using RNA-Sequencing. Our results provide a comparison between different sampling methods (invasive and non-invasive) to define the transcriptome of mammary gland tissue and milk cells. This information will be of value to investigators in choosing the most appropriate sampling method for different research applications to study specific physiological states during lactation. One of the simplest procedures to study the transcriptome associated with milk appears to be the isolation of total RNA directly from SC or MFG released into milk during lactation. Our results indicate that the SC and MFG transcriptome are representative of MGT and LCMEC and can be used as effective and alternative samples to study mammary gland expression without the need to perform a tissue biopsy.
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Project description:Background: The mammary gland is a dynamic organ that undergoes important physiological changes during reproductive cycles. Until now, data regarding the characterisation of miRNA in the mammary gland have been scarce and mainly focused on their abnormal expression in breast cancer. Our goal was to characterise the microRNA (miRNA) involved in mechanisms regulating the mammary function, with particular focus on the lactation stage Methodology/principal findings: Using high-throughput sequencing technology, an exhaustive repertoire of miRNA (miRNome) expressed in mouse and bovine mammary glands during established lactation was identified, characterised and compared. Furthermore, in order to obtain more information on miRNA loading in the RNA-induced silencing complex (RISC), the miRNome was compared with that obtained from RNA associated with the AGO2 protein (AGO2-miRNome) in mouse lactating mammary gland. This study enabled the identification of 164 and 167 miRNA in mouse and bovine, respectively. Among the 30 miRNA most highly expressed in each species, 24 were common to both species and six of them tended to be more highly expressed in lactating than in non-lactating mammary gland. The potential functional roles of these 24 miRNA were deduced using DIANA-miRPath software, based on miRNA/mRNA interactions. Moreover, seven putative novel miRNA were identified. Using DAVID analysis, it was concluded that the predicted targets of two of these putative novel miRNA are involved in mammary gland morphogenesis. Conclusion/significance: Our study provides an overview of the characteristics of lactating mouse and bovine mammary gland miRNA expression profiles. Moreover, species-conserved miRNA involved in this fundamental biological function were identified. These miRNomes will new be used as references for further studies during which the impact of animal breeding on the miRNA expression will be analysed.
2014-03-24 | GSE53511 | GEO
Project description:BackgroundMicroRNAs (miRNAs) have been implicated in the regulation of milk protein synthesis and development of the mammary gland (MG). However, the specific functions of miRNAs in these regulations are not clear. Therefore, the elucidation of miRNA expression profiles in the MG is an important step towards understanding the mechanisms of lactogenesis.ResultsTwo miRNA libraries were constructed from MG tissues taken from a lactating and a non-lactating Holstein dairy cow, respectively, and the short RNA sequences (18-30 nt) in these libraries were sequenced by Solexa sequencing method. The libraries included 885 pre-miRNAs encoding for 921 miRNAs, of which 884 miRNAs were unique sequences and 544 (61.5%) were expressed in both periods. A custom-designed microarray assay was then performed to compare miRNA expression patterns in the MG of lactating and non-lactating dairy cows. A total of 56 miRNAs in the lactating MG showed significant differences in expression compared to non-lactating MG (P<0.05). Integrative miRNA target prediction and network analysis approaches were employed to construct an interaction network of lactation-related miRNAs and their putative targets. Using a cell-based model, six miRNAs (miR-125b, miR-141, miR-181a, miR-199b, miR-484 and miR-500) were studied to reveal their possible biological significance.ConclusionOur study provides a broad view of the bovine MG miRNA expression profile characteristics. Eight hundred and eighty-four miRNAs were identified in bovine MG. Differences in types and expression levels of miRNAs were observed between lactating and non-lactating bovine MG. Systematic predictions aided in the identification of lactation-related miRNAs, providing insight into the types of miRNAs and their possible mechanisms in regulating lactation.
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Project description:miRNAs have been implicated in the regulation of milk protein synthesis and development of mammary gland. However, the function of miRNAs in regulating lactation is unclear. Therefore, the elucidation of miRNA expression profiles in MG provides a crucial entry into the understanding of the mechanisms of lactation initiation. Our present work is to examine miRNA expression profiles in bovine mammary gland, and to evaluate miRNAs function through the identification of differentially expressed miRNA between lactation and non-lactation mammary gland. Identification of novel miRNAs highlights the important function of low abundance and less conserved miRNAs. An interaction network of known miRNAs and their target genes around the lactation function was constructed to postulate the functional roles of miRNAs in mammary gland. This integrated analysis provides important information that will inspire further experimental investigations into the field of miRNAs and their targets during lactation. Examination of 2 different miRNA expression profilings in bovine mammary gland
2012-11-26 | E-GEOD-39000 | biostudies-arrayexpress
Project description:Citrullination is a post-translational modification (PTM) in which positively charged peptidyl-arginine is converted into neutral peptidyl-citrulline by peptidylarginine deiminase (PAD or PADI) enzymes. The full protein citrullinome in many tissues is unknown. Herein, we used mass spectrometry and identified 107 citrullinated proteins in the lactation day 9 (L9) mouse mammary gland including histone H2A, α-tubulin, and β-casein. Given the importance of prolactin to lactation, we next tested if it stimulates PAD-catalyzed citrullination using mouse mammary epithelial CID-9 cells. Stimulation of CID-9 cells with 5 µg/mL prolactin for 10 min induced a 2-fold increase in histone H2A citrullination and a 4.5-fold increase in α-tubulin citrullination. We next investigated if prolactin-induced citrullination regulates the expression of lactation genes β-casein (Csn2) and butyrophilin (Btn1a1). Prolactin treatment for 12 h increased β-casein and butyrophilin mRNA expression; however, this increase was significantly inhibited by the pan-PAD inhibitor, BB-Cl-amidine (BB-ClA). We also examined the effect of tubulin citrullination on the overall polymerization rate of microtubules. Our results show that citrullinated tubulin had a higher maximum overall polymerization rate. Our work suggests that protein citrullination is an important PTM that regulates gene expression and microtubule dynamics in mammary epithelial cells.
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Project description:Background: The mammary gland is a dynamic organ that undergoes important physiological changes during reproductive cycles. Until now, data regarding the characterisation of miRNA in the mammary gland have been scarce and mainly focused on their abnormal expression in breast cancer. Our goal was to characterise the microRNA (miRNA) involved in mechanisms regulating the mammary function, with particular focus on the lactation stage Methodology/principal findings: Using high-throughput sequencing technology, an exhaustive repertoire of miRNA (miRNome) expressed in mouse and bovine mammary glands during established lactation was identified, characterised and compared. Furthermore, in order to obtain more information on miRNA loading in the RNA-induced silencing complex (RISC), the miRNome was compared with that obtained from RNA associated with the AGO2 protein (AGO2-miRNome) in mouse lactating mammary gland. This study enabled the identification of 164 and 167 miRNA in mouse and bovine, respectively. Among the 30 miRNA most highly expressed in each species, 24 were common to both species and six of them tended to be more highly expressed in lactating than in non-lactating mammary gland. The potential functional roles of these 24 miRNA were deduced using DIANA-miRPath software, based on miRNA/mRNA interactions. Moreover, seven putative novel miRNA were identified. Using DAVID analysis, it was concluded that the predicted targets of two of these putative novel miRNA are involved in mammary gland morphogenesis. Conclusion/significance: Our study provides an overview of the characteristics of lactating mouse and bovine mammary gland miRNA expression profiles. Moreover, species-conserved miRNA involved in this fundamental biological function were identified. These miRNomes will new be used as references for further studies during which the impact of animal breeding on the miRNA expression will be analysed. microRNA profiles of mammary glands from 2 FVB/N mice at lactation day-12 and mammary biopsies from 4 Holstein dairy cows at mid-lactation, generated by deep sequencing, using Illumina HiSeq 2000.
2014-03-24 | E-GEOD-53511 | biostudies-arrayexpress
Project description:Enhancers are transcription factor platforms that synergize with promoters to control gene expression. Here, we investigate enhancers that activate gene expression several hundred-fold exclusively in the lactating mouse mammary gland. Using ChIP-seq for activating histone marks and transcription factors, we identify two candidate enhancers and one super-enhancer in the Csn1s2b locus. Through experimental mouse genetics, we dissect the lactation-specific distal enhancer bound by the mammary-enriched transcription factors STAT5 and NFIB and the glucocorticoid receptor. While deletions of canonical binding motifs for NFIB and STAT5, individually or combined, have a limited biological impact, a non-canonical STAT5 site is essential for enhancer activity during lactation. In contrast, the intronic enhancer contributes to gene expression only in late pregnancy and early lactation, possibly by interacting with the distal enhancer. A downstream super-enhancer, which physically interacts with the distal enhancer, is required for the functional establishment of the Csn1s2b promoter and gene activation. Lastly, NFIB binding in the promoter region fine-tunes Csn1s2b expression. Our study provides comprehensive insight into the anatomy and biology of regulatory elements that employ the JAK/STAT signaling pathway and preferentially activate gene expression during lactation.
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Project description:1. The conditions that promoted the solubilization of particulate lactose synthetase were effective for solubilizing the thiamine pyrophosphatase of the Golgi apparatus but differed from those effective for beta-glucuronidase or acid phosphatase of lysosomes. 2. Lactose synthetase-containing particles did not bind Mg(2+) or Cs(+) ions, suggesting that they are not related to endoplasmic reticulum membranes. 3. Intact lactose synthetase and thiamine pyrophosphatase particles banded isopycnically at a density of 1.143 in a sucrose gradient. The dissociated ;A' sub-unit of lactose synthetase, UDP-galactose hydrolase, p-nitrophenyl phosphate acid phosphatase, alkaline phosphatase and phosphodiesterase I were associated with particles of a broad density range from 1.12 to 1.20. Lysosomal enzymes beta-glucuronidase, arylsulphatase and beta-glycerophosphate acid phosphatase were associated with particles of density 1.20, 1.175 and 1.15 respectively. 4. Rate-zonal sedimentation studies indicated that lactose synthetase particles have S(20,w) values exceeding 24000s, corresponding to spherical particles of diameter exceeding 5.4x10(-5)cm. 5. Electron micrographs of lactose synthetase particles purified over 20-fold revealed small spherical bodies (0.1-0.5mu) resembling lysosomes, the smaller of which were attached to membranes, and larger heterogeneous spherical or oval bodies (0.7-1.8mu) resembling lipofuscin secretory granules. 6. The relationship between lactose synthetase particles and the Golgi origin of secretion granules is discussed.
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