Comparative analysis of gene expression profiles in differentiated subcutaneous adipocytes between Jiaxing Black and Large White pigs

- A better understanding of the genetic regulation network of subcutaneous fat tissue is necessary for the rational selection of Chinese domestic pig breeds.
- In the present study, subcutaneous adipocytes were isolated from Jiaxing Black pigs a Chinese indigenous pig breed with redundant subcutaneous fat deposition and Large White pigs a lean-type pig breed with relatively low subcutaneous fat deposition.
- The expression profiles of mRNAs and lncRNAs were compared by RNA-seq analysis to identify biomarkers correlated with the differences of subcutaneous fat deposition between the two breeds..
- Gene Ontology and KEGG pathway enrichment analyses revealed that the differentially expressed genes and differentially expressed lncRNAs were mainly involved in the immune response, cell fate determination, PI3K-Akt signaling pathway and MAPK signaling pathway, which are known to be related to adipogenesis and lipid metabolism.
- The activity of the MAPK signaling pathway was found to be associated with subcutaneous adipogenesis.
- These results provide new targets for further investigation of the molecular mechanisms regulating subcutaneous fat deposition in pigs..
- The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material.
- Full list of author information is available at the end of the article.
- Fat deposition is one of the most important economic traits of pigs.
- Porcine SC fat deposition is largely determined by the proliferation and differentiation of adipocytes [15].
- [21] demonstrated that C1q/tumor necrosis factor-related protein 6 (C1QTNF6) regulates porcine SC fat deposition via the MAPK and p53 signaling pathways using RNA-seq analysis.
- Identifying the regulatory role of lncRNAs in porcine ad- ipogenesis is of great importance for understanding the molecular mechanisms that regulate SC fat deposition in pigs.
- Although several reports on lncRNAs in porcine adipose tissue were published in the past 2 years, our understanding how lncRNAs regulate fat deposition in pigs is still very limited..
- By contrast, LW pigs are the most widely distributed lean-type pig breed with relatively low SC fat deposition [10].
- For each sample and 85.76% were uniquely mapped to the current version of the pig genome (Sscrofa 11.1), representing and 11,310 genes, respect- ively (Additional file 1: Supplementary Table 1).
- b Quantitive comparison of the up- and down-regulated genes.
- “Rheumatoid arthritis”, “Chemokine signaling pathway”,.
- “Cytokine-cytokine receptor interaction”, “B cell receptor signaling pathway”, “Leukocyte transendothelial migra- tion”, and “Systemic lupus erythematosus”.
- “Osteoclast differentiation”, “MAPK signaling pathway”.
- and “PI3K-Akt signaling pathway” were significantly enriched, all of which are highly associated with adipo- cyte differentiation and lipid accumulation.
- The results of enrichment analysis illustrated the regulatory differ- ences of SC fat deposition between JX and LW pigs..
- These results reflected the distinct mRNA expres- sion profiles of the two breeds (Fig.
- The size of the bubble indicates the number of involved genes.
- a Unsupervised hierarchical clustering of the expression profile of significant genes of DEGs.
- c Unsupervised hierarchical clustering of the expression profile of hub genes in PPI network, and the arrows (red: up-regulated.
- GO enrichment (a) and KEGG pathway enrichment (b) of the target genes.
- lipid accumulation were identified, including “MAPK signaling pathway”, “Regulation of lipolysis in adipocyte”,.
- “Calcium signaling pathway”, “p53 signaling pathway”,.
- “PI3K-Akt signaling pathway”, “cGMP-PKG signaling pathway”, and “cAMP signaling pathway” (Fig.
- In addition, the target genes enriched in four categories, in- cluding “MAPK signaling pathway”, “PI3K-Akt signaling pathway”, “Immune response”, and “Cell proliferation, differentiation and migration” were shown in Fig.
- 4c, and they were also identified in the enrichment analysis of DEGs.
- Moreover, LPAR1 and AKAP9 were also re- ported to be associated with the PI3K/AKT signaling pathway or MAPK signaling pathway.
- Thus, the DELs might play an essential role in the distinct adipogenesis of JX pigs..
- a Unsupervised hierarchical clustering of the expression profile of twelve randomly selected DEGs.
- b Q-PCR validation of the expression level of twelve randomly selected DEGs.
- Unsupervised hierarchical clustering of the expression profile of ten randomly selected DELs.
- d Q-PCR validation of the expression level of ten randomly selected DELs.
- Validation of the DEGs and DELs.
- To validate the reliability of the RNA-seq results, 12 DEGs and 10 DELs were randomly chosen for quantitative PCR (qPCR) verification (Fig.
- Overall, 81.8% of the results were in agreement between the two techniques..
- Verification of the pathway analysis.
- Because the MAPK signaling pathway was identified in the functional enrichment analysis of both DEGs and DELs, its activity in SC fat tissues of the two breeds was examined.
- Expression levels and phosphorylation of two kinases in the MAPK pathway, ERK1/2 and p38, were determined by western blot analysis.
- Consequently, a better understanding of the regulation network controlling SC fat tissue deposition is necessary for the rational genetic improvement of Chinese domestic pig breeds.
- This result indicated that many genes are suppressed in the SC adipocytes of JX pigs, which may be related to vigorous adipogenesis inside the cells..
- DEGs and target genes of DELs were mainly enriched in three pathways related to lipid metabolism and adipocyte differentiation, namely “Calcium signaling pathway”, “PI3K-Akt signaling pathway” and “MAPK.
- 6 Verification of MAPK signaling pathway by western blot.
- Quantification of the protein levels is shown on the right, and the results are expressed as the means ± SE of three samples.
- signaling pathway”.
- The calcium signaling pathway can regulate lipolysis and the accumulation of adipose tissue by changing the concentration of calcium ions in adipo- cytes [29, 30].
- Plum et al.[33] re- ported that the PI3K signaling pathway is induced by leptin and participates in biological processes related to obesity.
- The MAPK signaling pathway is common to several cell types, and it is involved in many biological processes, including cell proliferation, differentiation, de- velopment, and apoptosis [35].
- The MAPK signaling pathway was among the top 20 most highly enriched pathways in KEGG analysis, and the GO annotation of DEGs uncovered the ERK1-ERK2 cascade, which indi- cates the activity of MAPK signaling pathway [36].
- The oncogenic form of Ras protein is a strong activator of the ERK pathway, and transfection of 3 T3-L1 preadipocyte cells with an activated Ras expression vector led to growth arrest and terminal adipocyte differentiation.
- The expression of the key adipogenic regulators C/EBPα, β, and δ, as well as PPARγ requires ERK activity, but the phosphorylation of PPARγ caused by ERK inhibits adipocyte differentiation [38].
- A study of the role of p38 MAPK signaling in regu- lating fat deposition in mice suggested that p38 MAPK could inhibit adipogenic differentiation by inhibiting the activity and expression of C/EBPβ and PPARγ during the whole process of lipogenesis in mice [39].
- Therefore, the results of this study indicate that the MAPK signaling pathway may be important for mediating SC adipogenesis in JX pigs, and the difference between the two breeds may be related to mitogenic activity in preadipocytes regulated by the MAPK pathway.
- Considering that ERK and p38 are two main kinases in the MAPK pathway, and.
- ERK in SC adipose tissue of the two pig breeds.
- Thus, a precise understanding of the regulatory mechanism of these two pathways will be crucial for revealing the reason why JX pigs have a lower lean carcass weight ratio, and will be helpful for effi- ciently treating obesity..
- Thus, the obtained differential expression pro- files of mRNAs and lncRNAs indicated that there are potential functions of cytokines and immune cells in the fat deposition and metabolism of pigs.
- The adipocyte expansion caused by a positive energy balance results in the expression of chemoattractant molecules and infiltration of inflamma- tory cells [51].
- This difference of immune-related signaling pathways may be associated with the excessive SC fat deposition in JX pigs.
- adipose tissue through adipocyte progenitor recruitment [53].
- In the protein-protein interaction network of the iden- tified DEGs, twelve hub genes were identified by the MCC topological algorithm.
- The transcription factor FOS is a member of the AP-1 complex, and knockdown of FOS abolished the ongoing differentiation process [61]..
- In the predicted protein-protein interaction network constructed in this study, MMP9 is located at a key central node.
- Thus, the target genes of the identified DELs were predicted and subjected to enrich- ment analysis.
- AKAP9 regulates activation-induced retention of T lymphocytes at sites of inflammation and is related to the activation of the MAPK pathway in thyroid cancer [64].
- The DEGs and DELs identified in this study possibly reflect differences of fat deposition or deeper genetic dif- ferences between the two breeds.
- According to GO and KEGG analysis, several pathways related to lipid metab- olism were enriched, including“positive regulation of ERK1 and ERK2 cascade”, “Calcium signaling pathway”,.
- “PI3K-Akt signaling pathway”, “Osteoclast differenti- ation”, and “MAPK signaling pathway”, which may ac- count for the difference of fat deposition between the two breeds.
- Our findings expand the knowledge of the regulatory network in porcine SC adipocytes, which is necessary for the rational breeding of pigs for im- proved fat-related traits..
- Elevated activity of the MAPK/p38 pathway was detected in the SC fat of JX pigs.
- Taken together, the results may help explain the excessive fat deposition of JX pigs and offer a clue for genetic improvement of Chinese domestic pig breeds..
- All experimental procedures involving animals were per- formed in accordance with the guidelines of the Animal Care and Use Committee at the Jiaxing University.
- The lncRNAs that appeared as hits in at least three of the four software tools were included in the final result.
- For the significance of the proximity of coregu- lated lncRNA and mRNAs, Spearman and Pearson cor- relation coefficients were calculated based on the expression values of each lncRNA and mRNA, and co- expressed lncRNA-mRNA pairs with correlation coeffi- cients ≥0.6 were selected.
- When these target genes were located 10 kb upstream or 20 kb downstream of the lncRNAs, they were considered as cis target genes..
- Enrichment analyses and construction of the protein- protein interaction network.
- GO annotation and KEGG pathway enrichment analysis were performed for DEGs and lncRNA target genes to explore the main biological functions of the differentially expressed mRNA and lncRNAs.
- A QuantStudio3 Real-time PCR Instrument (Thermo Fisher Scientific, USA) was used for the qRT-PCR assay, and the reaction system was set up according to the manufacturer’s instructions of the 2 × Plus SYBR real-time PCR kit (CWbiotech).
- All data were presented as the means ±standard error (SE), and the statistical analysis of the qRT-PCR assay was implemented using GraphPad Prism 8.
- All authors analyzed the results and approved the final version of the manuscript..
- The raw datasets generated in this study were deposited in the Sequence Read Archive under the accession number PRJNA631903..
- Comprehensive transcriptomic view of the role of the LGALS12 gene in porcine subcutaneous and intramuscular adipocytes.
- The novel gene pFAM134B positively regulates fat deposition in the subcutaneous fat of Sus scrofa .
- Comparison of the adipogenesis in intramuscular and subcutaneous adipocytes from Bamei and Landrace pigs.
- Evaluation of the four breeds in synthetic line of Jiaxing Black Pigs and Berkshire for meat quality traits, carcass characteristics, and flavor substances.
- The development and endocrine functions of adipose tissue.
- Identification of the tuberous sclerosis complex-2 tumor suppressor gene product tuberin as a target of the phosphoinositide 3-kinase/akt pathway.
- Enhanced leptin-stimulated Pi3k activation in the CNS promotes white adipose tissue transdifferentiation..
- Caulerpa lentillifera extract ameliorates insulin resistance and regulates glucose metabolism in C57BL/KsJ-db/db mice via PI3K/AKT signaling pathway in myocytes.
- Signaling pathway of MAPK/ERK in cell proliferation, differentiation, migration, senescence and apoptosis.
- Adiponectin decreases lipids deposition by p38 MAPK/ATF2 signaling pathway in muscle of broilers.
- The role of adipose tissue immune cells in obesity and low-grade inflammation.
- Role of antigen presentation in the production of pro-inflammatory cytokines in obese adipose tissue.
- The role of infiltrating immune cells in dysfunctional adipose tissue.
- A mutation in the c-fos gene associated with congenital generalized lipodystrophy..
- Genome-Wide Profiling of the Microrna Transcriptome Regulatory Network to Identify Putative Candidate Genes Associated with Backfat Deposition in Pigs

Xem thử không khả dụng, vui lòng xem tại trang nguồn
hoặc xem Tóm tắt