RNA-Seq explores the functional role of the fibroblast growth factor 10 gene in bovine adipocytes differentiation
2
조회
0
다운로드
0
인용
0
참고문헌
키워드
초록
Objective: The present study was executed to explore the molecular mechanism of fibroblast growth factor 10 (<i>FGF10</i>) gene in bovine adipogenesis.Methods: The bovine <i>FGF10</i> gene was overexpressed through Ad-FGF10 or inhibited through siFGF10 and their negative control (NC) in bovine adipocytes, and the multiplicity of infection, transfection efficiency, interference efficiency were evaluated through quantitative real-time polymerase chain reaction, western blotting and fluorescence microscopy. The lipid droplets, triglycerides (TG) content and the expression levels of adipogenic marker genes were measured during preadipocytes differentiation. The differentially expressed genes were explored through deep RNA sequencing.Results: The highest mRNA level was found in omasum, subcutaneous fat, and intramuscular fat. Moreover, the highest mRNA level was found in adipocytes at day 4 of differentiation. The results of red-oil o staining showed that overexpression (Ad-FGF10) of the <i>FGF10</i> gene significantly (p<0.05) reduced the lipid droplets and TG content, and their downregulation (siFGF10) increased the measurement of lipid droplets and TG in differentiated bovine adipocytes. Furthermore, the overexpression of the <i>FGF10</i> gene down regulated the mRNA levels of adipogenic marker genes such as CCAAT enhancer binding protein alpha (<i>C/EBPα</i>), fatty acid binding protein (<i>FABP4</i>), peroxisome proliferator-activated receptor-γ (<i>PPARγ</i>), lipoprotein lipase (<i>LPL</i>), and Fas cell surface death receptor (<i>FAS</i>), similarly, down-regulation of the <i>FGF10</i> gene enriched the mRNA levels of <i>C/EBPα, PPARγ, FABP4</i>, and <i>LPL</i> genes (p<0.01). Additionally, the protein levels of PPARγ and FABP4 were reduced (p<0.05) in adipocytes infected with Ad-<i>FGF10</i> gene and enriched in adipocytes transfected with siFGF10. Moreover, a total of 1,774 differentially expressed genes (DEGs) including 157 up regulated and 1,617 down regulated genes were explored in adipocytes infected with Ad-FGF10 or Ad-NC through deep RNA-sequencing. The top Kyoto encyclopedia of genes and genomes pathways regulated through DEGs were the PPAR signaling pathway, cell cycle, base excision repair, DNA replication, apoptosis, and regulation of lipolysis in adipocytes.Conclusion: Therefore, we can conclude that the <i>FGF10</i> gene is a negative regulator of bovine adipogenesis and could be used as a candidate gene in marker-assisted selection.
Abstract
Objective: The present study was executed to explore the molecular mechanism of fibroblast growth factor 10 (FGF10) gene in bovine adipogenesis. Methods: The bovine FGF10 gene was overexpressed through Ad-FGF10 or inhibited through siFGF10 and their negative control (NC) in bovine adipocytes, and the multiplicity of infection, transfection efficiency, interference efficiency were evaluated through quantitative real-time polymerase chain reaction, western blotting and fluorescence microscopy. The lipid droplets, triglycerides (TG) content and the expression levels of adipogenic marker genes were measured during preadipocytes differentiation. The differentially expressed genes were explored through deep RNA sequencing. Results: The highest mRNA level was found in omasum, subcutaneous fat, and intramuscular fat. Moreover, the highest mRNA level was found in adipocytes at day 4 of differentiation. The results of red-oil o staining showed that overexpression (Ad-FGF10) of the FGF10 gene significantly (p<0.05) reduced the lipid droplets and TG content, and their downregulation (siFGF10) increased the measurement of lipid droplets and TG in differentiated bovine adipocytes. Furthermore, the overexpression of the FGF10 gene down regulated the mRNA levels of adipogenic marker genes such as CCAAT enhancer binding protein alpha (C/EBPα), fatty acid binding protein (FABP4), peroxisome proliferator-activated receptor-γ (PPARγ), lipoprotein lipase (LPL), and Fas cell surface death receptor (FAS), similarly, down-regulation of the FGF10 gene enriched the mRNA levels of C/EBPα, PPARγ, FABP4, and LPL genes (p<0.01). Additionally, the protein levels of PPARγ and FABP4 were reduced (p<0.05) in adipocytes infected with Ad-FGF10 gene and enriched in adipocytes transfected with siFGF10. Moreover, a total of 1,774 differentially expressed genes (DEGs) including 157 up regulated and 1,617 down regulated genes were explored in adipocytes infected with Ad-FGF10 or Ad-NC through deep RNA-sequencing. The top Kyoto encyclopedia of genes and genomes pathways regulated through DEGs were the PPAR signaling pathway, cell cycle, base excision repair, DNA replication, apoptosis, and regulation of lipolysis in adipocytes. Conclusion: Therefore, we can conclude that the FGF10 gene is a negative regulator of bovine adipogenesis and could be used as a candidate gene in marker-assisted selection.
참고문헌 (39)
- Mei C, "Genome-wide analysis reveals the effects of artificial selection on production and meat quality traits in Qinchuan cattle", Genomics, 111, pp.1201~1208 (2019)[DOI]
- Guo H, "Genetic variants in the promoter region of the KLF3 gene associated with fat deposition in Qinchuan cattle", Gene, 672, pp.50~55 (2018)[DOI]
- Raza SHA, "Association between FASN gene polymorphisms ultrasound carcass traits and intramuscular fat in Qinchuan cattle", Gene, 645, pp.55~59 (2018)[DOI]
- Wei D, "Polymorphism in promoter of SIX4 gene shows association with its transcription and body measurement traits in Qinchuan cattle", Gene, 656, pp.9~16 (2018)[DOI]
- Chen N, "Population structure and ancestry of Qinchuan cattle", Anim Genetics, 49, pp.246~248 (2018)[DOI]
- Martins TS, "Molecular factors underlying the deposition of intramuscular fat and collagen in skeletal muscle of nellore and angus cattle", PLoS ONE, 10, pp.e0139943~ (2015)[DOI]
- Raza SHA, "Expression of the bovine KLF6 gene polymorphisms and their association with carcass and body measures in Qinchuan cattle(Bos Taurus)", Genomics, 112, pp.423~431 (2020)[DOI]
- Fu YY, "The adipokine Chemerin induces lipolysis and adipogenesis in bovine intramuscular adipocytes", Mol Cell Biochem, 418, pp.39~48 (2016)[DOI]
- Cai H, "Global Transcriptome analysis during adipogenic differentiation and involvement of transthyretin gene in adipogenesis in cattle", Front Genet, 9, pp.463~ (2018)[DOI]
- Frank D, "Consumer acceptability of intramuscular fat", Food Sci Anim Resour, 36, pp.699~708 (2016)[DOI]
- Sarantopoulos CN, "Elucidating the preadipocyte and its role in adipocyte formation : a comprehensive review", Stem Cell Rev Rep, 14, pp.27~42 (2018)[DOI]
- Sakaue H, "Requirement of fibroblast growth factor 10 in development of white adipose tissue", Genes Dev, 16, pp.908~912 (2002)[DOI]
- Ohta H, "FGF10 and FGF21 as regulators in adipocyte development and metabolism", Endocr Metab Immune Disord Drug Targets, 11, pp.302~309 (2011)[DOI]
- Ahn J, "MicroRNA-146b promotes adipogenesis by suppressing the SIRT1-FOXO1cascade", EMBO Mol Med, 5, pp.1602~1612 (2013)[DOI]
- Nurgulsim K, "Bioinformatics and genetic variants analysis of FGF10 gene promoter with their association at carcass quality and body measurement traits in Qinchuan beef cattle", Anim Biotechnol, 34, pp.1950~1959 (2023)[DOI]
- Khan R, "Genetic variants in the TORC2 gene promoter and their association with body measurement and carcass quality traits in Qinchuan cattle", PloS one, 15, pp.e0227254~ (2020)[DOI]
- Khan R, "Bioinformatics analysis and transcriptional regulation of TORC1 gene through transcription factors NRF1 and Smad3 in bovine preadipocytes", Genomics, 112, pp.1575~1587 (2020)[DOI]
- Khan R, "RNA-seq reveal role of bovine TORC2 in the regulation of adipogenesis", Arch Biochem Biophys, 680, pp.108236~ (2020)[DOI]
- Chen S, "fastp : an ultra-fast all-in-one FASTQ preprocessor", Bioinformatics(Oxford, England), 34, pp.i884~90 (2018)[DOI]
- Langmead B, "Fast gapped-read alignment with Bowtie 2", Nat Methods, 9, pp.357~359 (2012)[DOI]
- Kim D, "HISAT : a fast spliced aligner with low memory requirements", Nat Methods, 12, pp.357~360 (2015)[DOI]
- Pertea M, "Transcriptlevel expression analysis of RNA-seq experiments with HISAT, StringTie and Ballgown", Nat Protoc, 11, pp.1650~1667 (2016)[DOI]
- Love MI, "Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2", Genome Biol, 15, pp.550~ (2014)[DOI]
- Robinson MD, "edgeR : a Bioconductor package for differential expression analysis of digital gene expression data", Bioinformatics(Oxford, England), 26, pp.139~140 (2010)[DOI]
- Zhang R, "Expression profiles and associations of FGF1 and FGF10 with intramuscular fat in Tibetan chicken", Br Poult Sci, 59, pp.613~617 (2018)[DOI]
- Sakaue H, "Requirement of fibroblast growth factor 10 in development of white adipose tissue", Genes Dev, 16, pp.908~912 (2002)[DOI]
- Matsubara Y, "Characterization of the expression profiles of adipogenesis-related factors, ZNF423, KLFs and FGF10, during preadipocyte differentiation and abdominal adipose tissue development in chickens", Comp Biochem Physiol B Biochem Mol Biol, 165, pp.189~195 (2013)[DOI]
- Khan R, "Function and transcriptional regulation of bovine TORC2 gene in adipocytes : roles of C", EBPγ, XBP1, INSM1 and ZNF263, 2019, pp.20, 4338~
- Asaki T, "Roles of fibroblast growth factor 10(Fgf10)in adipogenesis in vivo", Mol Cell Endocrinol, 218, pp.119~128 (2004)[DOI]
- Xu Q, "Fibroblast growth factor 10(FGF10)promotes the adipogenesis of intramuscular preadipocytes in goat", Mol Biol Rep, 45, pp.1881~1888 (2018)[DOI]
- Huang R, "The role of peroxisome proliferator-activated receptors(PPARs)in pan-cancer", PPAR Res, 2020, pp.6527564~ (2020)[DOI]
- Spiegelman B, "Regulation of adipogenesis and energy balance by PPARγ and PGC-1", Int J Obes, 24, pp.S8~S10 (2000)[DOI]
- Liang C, "The molecular characteristics of the FAM13A gene and the role of transcription factors ACSL1 and ASCL2 in its core promoter region", Genes, 10, pp.981~ (2019)[DOI]
- Samulin J, "Differential gene expression of fatty acid binding proteins during porcine adipogenesis", Comp Biochem Physiol B Biochem Mol Biol, 151, pp.147~152 (2008)[DOI]
- Shijun L, "Function and characterization of the promoter region of perilipin 1(PLIN1) : Roles of E2F1, PLAG1, C", EBPβ, and SMAD3 in bovine adipocytes, 2020, pp.112, 2400~2409
- Powers CJ, "Fibroblast growth factors, their receptors and signaling", Endocr Relat Cancer, 7, pp.165~197 (2000)
- Patel NG, "Essential role of fibroblast growth factor signaling in preadipoctye differentiation", J Clin Endocrinol Metab, 90, pp.1226~1232 (2005)[DOI]
- Xu Q, "Fibroblast growth factor 21 regulates lipid accumulation and adipogenesis in goat intramuscular adipocyte", Anim Biotechnol, 32, pp.318~326 (2021)[DOI]
- Widberg CH, "Fibroblast growth factor receptor 1 is a key regulator of early adipogenic events in human preadipocytes", Am J Physiol Endocrinol Metab, 296, pp.E121~E131 (2009)[DOI]