Systems analysis of immune responses in Marek's disease virus-infected chickens identifies a gene involved in susceptibility and highlights a possible novel pathogenicity mechanism

Abstract

Marek's disease virus (MDV) is a highly contagious oncogenic alphaherpesvirus that causes disease that is both a cancer model and a continuing threat to the world's poultry industry. This comprehensive gene expression study analyzes the host response to infection in both resistant and susceptible lines of chickens and inherent expression differences between the two lines following the infection of the host. A novel pathogenicity mechanism, involving the downregulation of genes containing HIC1 transcription factor binding sites as early as 4 days postinfection, was suggested from this analysis. HIC1 drives antitumor mechanisms, suggesting that MDV infection switches off genes involved in antitumor regulation several days before the expression of the MDV oncogene meq. The comparison of the gene expression data to previous QTL data identified several genes as candidates for involvement in resistance to MD. One of these genes, IRG1, was confirmed by single nucleotide polymorphism analysis to be involved in susceptibility. Its precise mechanism remains to be elucidated, although the analysis of gene expression data suggests it has a role in apoptosis. Understanding which genes are involved in susceptibility/resistance to MD and defining the pathological mechanisms of the disease gives us a much greater ability to try to reduce the incidence of this virus, which is costly to the poultry industry in terms of both animal welfare and economics.

Fig. 1.

Gene expression clusters generated using the Expander program (http://acgt.cs.tau.ac.il/expander/expander.html). Shown are the expression profiles of genes upregulated during the response to virus (A) and those downregulated (B). When genes expressed in the two different lines were examined, similar profiles were noted: gene expression was inherently higher in the resistant line (C), and genes were more highly expressed in the susceptible line (D).

Fig. 2.

Overrepresentation analysis using the Expander program. (A) The GO biological processes which are significantly enriched during the host response to infection. The frequency of genes of a functional class within the examined set is described as a percentage of the total. (B) Transcription factor binding sites present in differentially expressed genes which are significantly overrepresented in up- and downregulated genes during the host response to MDV infection. The frequency ratio (frequency of the set divided by the frequency of the background) is shown.

Fig. 3.

Ingenuity pathway analysis of the host response to MDV infection. Shown are the most highly represented canonical pathways as revealed after IPA of genes differentially expressed during the host response to MDV (in the spleen). The line represents the ratio of the number of genes represented within each pathway to the total number of genes in the pathway.

Fig. 4.

Network analysis using the IPA program. (A) Interaction network representing genes involved in the inflammatory response, showing genes upregulated in response to MDV infection (red) and genes downregulated (green). IRG1 is seen to be highly upregulated in this analysis. (B) Genes currently known to interact with IRG1.

Fig. 5.

Analysis of the IRG1-containing cluster of genes identified using Biolayout Express 3D software. (A) EasyGO analysis shows a significant enrichment of genes involved in apoptosis and the regulation of apoptosis. (B) IPA analysis also reveals cell death to be a highly featured function of the genes in this cluster. (C) A subcluster identified using the Expander program is seen to be highly overrepresented (more than 4-fold) by genes which contain a binding site for the transcription factor OCT1 (octamer-binding transcription factor 1). Genes with binding sites for DBP (albumin D box-binding protein) also are seen to be enriched. DBP is thought to be involved in the modulation of circadian rhythms.