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Using scale and feather traits for module construction provides a functional approach to chicken epidermal development

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Abstract

Gene co-expression network analysis has been a research method widely used in systematically exploring gene function and interaction. Using the Weighted Gene Co-expression Network Analysis (WGCNA) approach to construct a gene co-expression network using data from a customized 44K microarray transcriptome of chicken epidermal embryogenesis, we have identified two distinct modules that are highly correlated with scale or feather development traits. Signaling pathways related to feather development were enriched in the traditional KEGG pathway analysis and functional terms relating specifically to embryonic epidermal development were also enriched in the Gene Ontology analysis. Significant enrichment annotations were discovered from customized enrichment tools such as Modular Single-Set Enrichment Test (MSET) and Medical Subject Headings (MeSH). Hub genes in both trait-correlated modules showed strong specific functional enrichment toward epidermal development. Also, regulatory elements, such as transcription factors and miRNAs, were targeted in the significant enrichment result. This work highlights the advantage of this methodology for functional prediction of genes not previously associated with scale- and feather trait-related modules.

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Acknowledgements

Authors are thankful to Dr. Richard L. Goodwin (University of South Carolina, School of Medicine) for kindly providing chicken embryos and Dr. Diego Altomare for performing microarray experiments. The microarray work was supported by the National Institute of General Medical Sciences (grant number 8 P20 GM103499) and the National Center for Research Resources (grant number 5 P20 RR016461).

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Authors and Affiliations

  1. Department of Biological Sciences, University of South Carolina, Columbia, SC, 29208, USA

    Weier Bao, Matthew J. Greenwold & Roger H. Sawyer

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  1. Weier Bao
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  2. Matthew J. Greenwold
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  3. Roger H. Sawyer
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Correspondence to Weier Bao.

Electronic supplementary material

Supplemental Table 1

We apply a binary system (1 for positive and 0 for negative) to define and quantify the qualitative traits to measure the external trait based on the microarray expression profiles. This is the spreadsheet of this particular trait-defining strategy for 31 samples (XLSX 10 kb)

Supplemental Table 2

For the scale and feather traits, we list all modules having moderate Module-Trait correlations (r > 0.5, p < 0.01) individually, we also highlighted the modules also having moderate MM-GS correlations (r > 0.5, p < 0.01) (XLSX 10 kb)

Supplemental Table 3

Demonstrates all epidermal development-related genes (EDRGs) in brown and black modules. We also list the number of the edges (connectivity) for these genes. The rank of these EDRGs based on the connectivity number among all genes having gene symbol annotation and the top percentage were listed (XLSX 10 kb)

Supplemental Fig. 1

Illustrates the sample clustering based on individual sample expression profiles. Each array sample was named as the tissue type with the days of embryonic stages. The y-axis (height) is the distance metric calculated by average linkage hierarchical clustering method. There are no outliers and most of the samples with the same tissue type and embryonic stage cluster together (PNG 72 kb)

Supplemental Fig. 2

We performed the analysis of network topology to facilitate choosing soft threshold power β for constructing the WGCNA network. The left panel demonstrates the scale-free topology fit index (vertical axis) as the soft threshold power β (horizontal axis) varies. The right panel displays the mean connectivity (degree, vertical axis) as the soft threshold power β (horizontal axis) varies. The soft threshold power was selected based on the criterion of approximate scale-free topology which has high scale-free topology fit index (normally above 0.8) and the saturation of the mean connectivity was reached by the lowest power β (PNG 66 kb)

Supplemental File 1

To incorporate the external traits, gene significance (GS) of the genes having Entrez Gene ID annotations for Feather (“Feather trim” worksheet) and Scale (Scale trim worksheet) traits were sorted based on the absolute GS value. We also list the corresponding module color and the p value of GS (XLSX 4447 kb)

Supplemental File 2

We performed the KEGG pathway and Gene Ontology enrichment analysis individually for 11 modules having both correlated Module-Trait and MM-GS relationships (red color modules in Supplemental Table 2). The 2 worksheets listed all significantly enriched KEGG pathways and GO annotations with details based on the corrected p value (XLSX 10 kb) (XLSX 337 kb)

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Bao, W., Greenwold, M.J. & Sawyer, R.H. Using scale and feather traits for module construction provides a functional approach to chicken epidermal development. Funct Integr Genomics 17, 641–651 (2017). https://doi.org/10.1007/s10142-017-0561-0

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