Gene expression and alternative splicing reveal the co-regulation of host response mechanisms to avian leukosis virus subgroup J-infected in laying hens

Abstract

Avian leukosis in China has spread from broiler chickens to the local breeds and commercial laying hens. Studying resistance to avian leukosis is important for disease-resistant breeding programs. Gene expression and different transcripts may affect immune function. In this study, we compared five naturally infected Rhode Island Red (RIR) hens carrying tumor with five uninfected individuals to explore avian leukosis virus subgroup J (ALV-J) induced differences in gene expression and alternative splicing (AS) in the liver, spleen caused. Analyses revealed 847, 80 differentially expressed genes (DEGs), along with 207, 167 differential alternative splicing genes (DASGs) in the liver, spleen respectively. Most differential splicing events involved exon skipping. Although most genes showed no significant expression changes, their protein spatial structures were altered by AS. In the liver, microtubule cytoskeleton-related functions were co-regulated by both gene expression and splicing, with CCSER2 and MAPT exhibiting the highest splicing frequency. In the spleen, splicing predominantly affected RNA-processing genes, where PKLR and SRSF7 functioned as key regulators. Notably, PKLR-interacting genes (THRSP, ADH1C, AQP3) were significantly downregulated in infected groups, potentially promoting viral replication and tumor proliferation. These findings demonstrate that AS contributes to the host response to ALV-J infection through multiple mechanisms, including protein structural remodeling and dysregulation of coordinated interaction networks. This study provides new insights into the genetic basis of ALV-J resistance in laying hens.

Keywords: Alternative splicing; Avian leukosis virus subgroup J; Gene expression; Laying hen.

Fig. 1

Samples PCR result and tissues characterization. (A) ALV-J positive group with tumor tissue (left: control group; middle and right: ALV-J positive group). (B) Samples diagnosis analysis based on the PCR. (C) The relative expression of ALV-J in spleen was measured by qRT-PCR (n = 5). (D) The relative expression of ALV-J in liver was measured by qRT-PCR (n = 5).

Fig. 2

DEGs analysis of two tissues in ALV -J infected chicken. (A) The number of DEGs in the liver and spleen (n = 5). (B) Venn diagrams of DEGs in liver, spleen (n = 5). (C) Volcano plots presenting all DEGs between control and infection group in liver. FDR ≤ 0.05 and log₂FoldChange ≥ 1 or ≤ −1. (D) Volcano plots presenting all DEGs between control and infection group in spleen. (E). Functional enrichment analyses GO terms of DESs in the liver. (F) Functional enrichment analyses GO terms of DESs in the spleen.

Fig. 3

DASGs analysis in liver and spleen. (A) The number of DAS events in the liver and spleen (n = 5). (B) The number of DASGs in the liver and spleen (n = 5). (C) Venn diagrams of DASGs in liver, spleen (n = 5). (D) Functional enrichment analyses GO terms of DASGs in the liver. (E) Functional enrichment analyses GO terms of DASGs in the spleen.

Fig. 4

Liver-specific alternative splicing events associated with microtubule cytoskeleton regulation. (A) DASGs and DEGs functionally enriched in microtubule cytoskeleton-related pathways. (B) Exon skipping patterns of CCSER2 and MAPT, showing infection-induced alternative splicing events. (C) Predicted three-dimensional protein structures of CCSER2 isoforms generated through AS. (D) Predicted three-dimensional protein structures of MAPT isoforms generated through AS.

Fig. 5

Spleen-specific AS events associated with RNA regulatory proteins. (A) Heatmap of DASGs involved in mRNA binding and RNA splicing. (B) Protein-protein interaction network of DASGs related to mRNA binding and RNA splicing. (C) Exon skipping patterns of PKLR and SRSF7. (D) Predicted three-dimensional structures of PKLR isoforms. (E) Predicted three-dimensional structures of SRSF7 isoforms. (F) Protein-protein interaction network of DASGs and DEGs. (G) The relative expression of interaction genes of PKLR.