Consensus module analysis of abdominal fat deposition across multiple broiler lines

- Consensus module analysis of abdominal fat deposition across multiple broiler lines.
- Background: Despite several RNA-Seq and microarray studies on differentially expressed genes (DEGs) between high- and low-abdominal fat deposition in different broiler lines, to our knowledge, gene coexpression analysis across multiple broiler lines has rarely been reported.
- Here, we constructed a consensus gene coexpression network focused on identifying consensus gene coexpression modules associated with abdominal fat deposition across multiple broiler lines using two public RNA-Seq datasets (GSE42980 and GSE49121)..
- Results: In the consensus gene coexpression network, we identified eight consensus modules significantly correlated with abdominal fat deposition across four broiler lines using the consensus module analysis function in the weighted gene coexpression network analysis (WGCNA) package.
- The eight consensus modules were.
- moderately to strongly preserved in the abdominal fat RNA-Seq dataset of another broiler line (SRP058295)..
- Furthermore, we identified 5462 DEGs between high- and low-abdominal fat lines (FL and LL) (GSE42980) and 6904 DEGs between high- and low-growth (HG and LG) (GSE49121), including 1828 overlapping DEGs with similar expression profiles in both datasets, which were clustered into eight consensus modules.
- Pyruvate metabolism, fatty acid metabolism, and steroid biosynthesis were significantly enriched in the green, yellow, and medium purple 3 consensus modules.
- The PPAR signaling pathway and adipocytokine signaling pathway were significantly enriched in the green and purple consensus modules.
- Autophagy, mitophagy, and lysosome were significantly enriched in the medium purple 3 and yellow consensus modules..
- Conclusion: Based on lipid metabolism pathways enriched in eight consensus modules and the overexpression of numerous lipogenic genes in both FL vs.
- LG, we hypothesize that more fatty acids, triacylglycerols (TAGs), and cholesterol might be synthesized in broilers with high abdominal fat than in broilers with low abdominal fat.
- According to autophagy, mitophagy, and lysosome enrichment in eight consensus modules, we inferred that autophagy might participate in broiler abdominal fat deposition.
- Altogether, these studies suggest eight consensus modules associated with abdominal fat deposition in broilers.
- Our study also provides an idea for investigating the molecular mechanism of abdominal fat deposition across multiple broiler lines..
- Keywords: WGCNA, Broiler, Abdominal fat deposition, Lipid metabolism, Autophagy.
- abdominal fat deposition [3].
- Excessive abdominal fat deposition not only wastes feed to producers but also in- creases risk to human health [4, 5].
- Therefore, it is com- mercially relevant to investigate the molecular mechanism of abdominal fat deposition and to reduce this deposition in broilers..
- At present, several studies have reported abdominal fat differentially expressed genes (DEGs) between high- and.
- low-abdominal fat deposition in different broiler lines using RNA-Seq or microarray technology.
- There were 1687 DEGs identified between high- and low-abdominal fat line (FL and LL) broilers with a 2.8-fold divergence in abdominal fat percentage (AFP) at 7 weeks of age [6]..
- A total of 2410 DEGs were identified in abdominal fat tissue between high- and low-growth-rate (HG and LG) broilers, with a 19.2-fold divergence in AFP at 7 weeks of age [7].
- A total of 230 DEGs were identified between the North- east Agricultural University (NEAU) broiler lines diver- gently selected for abdominal fat content with a 1.9-fold divergence in AFP at 7 weeks of age [9].
- In each study, AFP was significantly different between broilers with high and low abdominal fat deposition, but the DEGs were not the same.
- The differences among these studies may disturb further research on abdominal fat deposition in broilers.
- Therefore, it is important to con- struct a consensus gene coexpression network across multiple broiler lines and to detect consensus modules correlated with abdominal fat deposition.
- In the current study, to explore the molecular mech- anism of abdominal fat deposition across multiple broiler lines, we constructed a consensus gene coexpres- sion network across GSE42980 (12 FL and 12 LL sam- ples) and GSE49121 (8 HG and 8 LG samples) and detected consensus modules associated with abdominal fat deposition.
- Then, we tested the preservation of con- sensus modules in abdominal fat RNA-Seq data of an- other broiler line (SRP058295).
- Our study provides an idea for investigating abdominal fat deposition across multiple broiler lines..
- To reduce the influence of nongenetic factors, we collected public abdominal fat gene expression profile datasets of broilers under the same age, sex, and normal feed conditions.
- low (LL) abdominal fat at similar feed intake and body weight, and FL and LL broilers exhibited a 2.8-fold divergence in AFP at 7 weeks of age [6, 12].
- GSE42980 is an abdominal fat RNA-Seq dataset of male broilers at 7 weeks of age that consists of 12 FL and 12 LL samples [6].
- GSE49121 is an abdominal fat RNA-Seq dataset of male broilers that consists of eight HG and eight LG samples..
- Because two pairwise broiler lines (FL and LL, HG and LG) are divergently selected over several generations and exhibit significant divergence in AFP at 7 weeks of age, the four broiler lines are good genetic models for abdominal fat deposition research.
- SRP058295 is an abdominal fat RNA-Seq dataset of male broilers [8]..
- Therefore, the SRP058295 data- set was used to validate the preservation of consensus modules across two datasets.
- To investigate abdominal fat gene expression differences across two pairwise comparisons, 5462 and 6904.
- Clustering dendrograms of GSE42980 and GSE49121 revealed a degree of preservation of consensus modules across the two datasets (Fig.
- The density value D = 0.66 reflected an overall preservation of consensus modules across the two networks (Fig.
- Consensus modules correlated with abdominal fat deposition.
- To detect consensus modules associated with abdominal fat deposition, we calculated the correlation between each of the consensus modules and abdominal fat depos- ition across the two datasets.
- Eight consensus modules (dark green, green, medium purple 3, purple, saddle brown, sky blue, turquoise, and yellow) were significantly correlated with abdominal fat deposition across the two datasets, and the correlation of the yellow consensus module was the highest (r = 0.93, p Fig.
- There were 13 consensus modules significantly correlated with fat deposition within GSE42980, and the correlation of the blue con- sensus module was the highest (r = 0.93, p Additional file 5: Figure S3, a).
- A total of 27 consensus modules were significantly correlated with abdominal fat deposition within GSE49121, and the correlation of the steel blue consensus module was the highest (r = 0.99, p Additional file 5: Figure S3, b)..
- Preservation of interested consensus modules.
- To verify the preservation of eight consensus modules across two datasets, we detected how many genes among.
- the 2020 common DEGs were in the eight consensus modules.
- The results showed that a total of 1828 genes (90.5% of common DEGs) were detected in the eight consensus modules (Fig.
- To test whether the eight consensus modules were stable in other broiler lines, preservation of eight consensus modules was tested in another broiler abdominal fat RNA-Seq dataset (SRP058295).
- and yellow consensus modules, and the relevant genes of the three pathways, ULK2, BECN1, MAP1 LC3A, MAP1 LC3B, ATG3, ATG4B, ATG12, and ATG9A, were identi- fied in the consensus modules (Additional file 6: Table S3)..
- To detect gene coexpression modules associated with abdominal fat deposition, we performed the enrichment of KEGG pathways and GO terms of modules within GSE42980 and GSE49121, respectively.
- There were 116 pathways and 1427 GO terms significantly enriched in the genes of modules significantly correlated with abdominal fat deposition within the GSE42980 dataset (Additional file 6: Table S3).
- Broiler chickens have been selected to increase produc- tion performance, but these gains are also accompanied by excessive abdominal fat deposition [4].
- Therefore, to iden- tify the genes associated with abdominal fat deposition across multiple broiler lines, we constructed a consensus gene coexpression network and identified eight consen- sus modules significantly correlated with abdominal fat deposition across the GSE42980 and GSE49121 datasets..
- Then, the preservation of eight consensus modules was verified in another broiler abdominal fat RNA-Seq data- set (SRP058295).
- The expression level of ACSBG2 mRNA is significantly upregulated during abdominal fat-derived preadipocyte differentiation of chickens [14, 15].
- The fatty acid metab- olism genes FADS2 and SCD are significantly upregu- lated in chickens with high abdominal fat content and associated with the PPAR signaling pathway [16].
- Therefore, we inferred that the synthesis of fatty acids, TAG, and cholesterol in broilers with high abdominal fat deposition was greater than that in broilers with low abdominal fat deposition..
- Abdominal fat deposition is a complex biological process that is controlled by a series of critical genes in signaling pathways.
- In chickens, PPARG is a well-known critical gene that is reported to positively regulate adipo- genesis, lipid metabolism, adipocyte differentiation, and abdominal fat deposition [22–24].
- chicken abdominal fat.
- In the current study, autophagy, mitophagy, and lysosome pathways were significantly enriched in the medium purple 3 and yellow consensus modules.
- 3 Consensus module-abdominal fat deposition relationship.
- Therefore, we inferred that autophagy initi- ation might participate in broiler abdominal fat deposition..
- In our current study, we predicted eight consensus modules across four broiler lines (FL and LL, HG and LG), the preservations of which were verified in an- other broiler line dataset (SRP058295).
- Pyruvate me- tabolism, fatty acid metabolism, glycerolipid metabolism, steroid biosynthesis, the PPAR signaling pathway, and the adipocytokine signaling pathway were significantly enriched in the eight consensus modules.
- Relevant genes of these lipid metabolism pathways included PDHA1, FASN, ACSBG2, FADS2, SCD, GPAT3, PNPLA7, DHCR7, PPARG, RXRG, FABP3, and CD36, all of which were identified in eight consensus modules and significantly upregulated in both the FL vs.
- Based on lipid metabolism pathways enriched in eight consensus modules and overexpression of numerous lipogenic genes, we hypothesize that more fatty acids, TAGs, and cholesterol might be synthesized in broilers with high abdominal fat than in broilers with low abdom- inal fat.
- Therefore, we inferred that autophagy initiation might participate in broiler abdominal fat deposition..
- These genes identified in eight consensus modules can provide targets for further research into abdom- inal fat deposition in multiple broiler lines.
- To reduce the influence of nongenetic factors, such as analytical methods, versions of the genome and annotated files, experimental technologies (RNA-Seq or microarray), sex, diet condition, and weeks of age, we only collected abdominal fat RNA-Seq data of male broilers at 7 weeks of age with normal feed.
- In the current study, we downloaded GSE42980 [6], GSE49121 [7], and SRP058295 [8] from the GEO (Gene Expression Omnibus) and SRA (Sequence Read Archive) databases of NCBI (National Center for Bio- technology Information)..
- 4 Venn diagram of genes within eight consensus modules and DEGs within GSE42980 and GSE49121.
- and the circle of eight consensus modules represents the number of genes within eight consensus modules.
- 5 Preservation analyses of eight consensus modules.
- Preservation of eight consensus modules tested in the SRP058295 dataset using Zsummary.
- Table 2 KEGG enrichment in eight consensus modules.
- Consensus modules Pathway ID Description p -value Gene count.
- LG (GSE49121) were identified by testing the two factor var- iables of high- and low-abdominal fat deposition and batch using DEseq2 (1.22.2) with default parameters [41].
- Then, gene coexpression networks for Table 2 KEGG enrichment in eight consensus modules (Continued).
- Correlation analysis between consensus modules and abdominal fat deposition.
- To identify mod- ules of interest, the correlation between each module eigengene of consensus modules and abdominal fat deposition traits was calculated using the correlation analysis function in the WGCNA package [10], and significant consensus modules were identified with the threshold (p-value ≤0.05)..
- Preservation test of consensus modules.
- To test the preservation of consensus modules signifi- cantly correlated with abdominal fat deposition in other broiler lines, abdominal fat RNA-Seq data of male broilers (SRP058295) were downloaded to construct a test network for preservation analysis.
- The preservation analysis of consensus modules was performed using the modulePreservation function in the WGCNA package with the parameters nPermutations = 200, randomSeed = 1, quickCor = 0, verbose = 3 [11].
- Function enrichment of consensus modules.
- The significance threshold for the pathways and GO terms associated with the consensus modules was set at a p-value ≤0.05..
- Summary of genes within consensus modules and DEGs within FL vs.
- Heatmaps of consensus modules correlated with abdominal fat deposition within GSE42980 or GSE49121..
- Functional enrichment correlated with genes within eight consensus modules..
- AFP: Abdominal fat percentage.
- Chicken lines divergent for low or high abdominal fat deposition: a relevant model to study the regulation of energy metabolism.
- Genomic insights into the multiple factors controlling abdominal fat deposition in a chicken model.
- RNA-Seq analysis of abdominal fat in genetically fat and lean chickens highlights a divergence in expression of genes controlling adiposity, hemostasis, and lipid metabolism.
- Transcriptional analysis of abdominal fat in chickens divergently selected on bodyweight at two ages reveals novel mechanisms controlling adiposity: validating visceral adipose tissue as a dynamic endocrine and metabolic organ.
- RNA-Seq analysis of abdominal fat reveals differences between modern commercial broiler chickens with high and low feed efficiencies.
- Transcriptional analysis of abdominal fat in genetically fat and lean chickens reveals adipokines, lipogenic genes and a link between hemostasis and leanness.
- Identification of differentially expressed genes and pathways between intramuscular and abdominal fat-derived preadipocyte differentiation of chickens in vitro.
- Decreased testosterone levels after caponization leads to abdominal fat deposition in chickens

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