The Molecular Mechanism For Increased Pathogenicity Of Avian Leukosis Virus Subgroup J

The diseases of the avian leukosis subgroup J with neoplasia in chickens was induced byavian leukosis virus subgroup J (ALV-J). ALV-J was first isolated from meat-type chickens in1988and then became widespread around the world. Field cases of ALV-J infection and tumors incommercial layer chickens were not found worldwide until2004. However, in China, field casesof ALV-J infection and tumors in commercial layer chickens emerged in2004and then layerflocks have experienced outbreaks of ALV-J from2008. Therefore, to explore the mechanism ofthe enhanced ALV-J pathogenicity has great scientific significance based on the ALV-J has beengaining a stronger foot-hold in China.A total of16ALV-J strains were isolated from clinical samples from different layer flocksand then the molecular epidemiology of ALV-J strains isolated was investigated. The env gene and3’UTR of ALV-J layer strains were different from those of the ALV-J broiler isolates. Furtheranalysis of3’UTR by chronologically showed that the205-nucleotide deletion in the3’UTR ofALV-J is the result of the natural evolution of the ALV-J genome.To determine the role of the205-nucleotide deletion in the stronger pathogenicity of ALV-J inChina, a pair of viruses were constructed and rescued. The first virus was an ALV-J Chineseisolate (designated HLJ09SH01) containing the205-nucleotide deletion in its3’UTR. The secondvirus was a chimeric clone in which the3’UTR contains a205-nucleotide sequence correspondingto a region of the ALV-J prototype virus. The occurring205-nucleotide deletion in the3’UTRfacilitates the transport of unspliced RNA and contributes to higher viral replication in vascularendothelial cells. Compared to rHLJ09SH01A205, rHLJ09SH01showed a moderate growthadvantage in vivo, in addition to exhibiting a higher oncogenicity rate and lethality rate in layersand broilers. All of these not only indicate that the unique205-nucleotide deletion in the ALV-Jgenome occurred naturally in China and contributes to increased pathogenicity in layers andbroilers under experimental conditions.To explore the molecular mechanism underlying the stronger pathogenicity of ALV-J, Chickembryos, which possess a mechanism for vasculature formation that is similar to the angiogenesisthat plays a vital role in tumorigenesis, were used to confirm that the increased replication ofALV-J increased the expression of VEGF-A and VEGFR-2and then increased oncogenicity andpathogenicity; The ALV-J persistent infection induces TP53gene mutation, producing a p53mutants. Compared to the wild type p53in DF-1cells, p53mutants lost the ability to inhibitALV-J promoter activity and relieved the inhibition of viral replication by wild type p53in DF-1cells. This indicates that ALV-J overcame the host factor restriction and facilitated viralreplication, which was an important mechanism for pathogenicity.The rigorous eradication programmes were used to minimize the harm of ALV-J, howevermiRNAs may be available to be against the virus, which could not be prevented by vaccine.Although viral miRNAs predicted by bioinformatics were not identified by experimentalidentification, the host miRNA gga-miR-1650was screened for its potential interaction with the 5’UTR of ALV-J and the ability to suppress luciferase-reporter activity. A mutational analysis ofpredicted gga-miR-1650-binding sites showed that the5’ and3’ ends of gga-miR-1650contributed to the interaction between gga-miR-1650and its target located at the5’UTR.Overexpression of gga-miR-1650was shown to down-regulate the expression of the viral proteinand influence the replication of ALV-J through binding to the5’UTR.Overall, based on the ALV-J induced serious harm in the Chinese poultry industry, we firsttook molecular epidemiological research on ALV-J isolates and found that the205-nucleotidedeletion in the3’UTR was the natural evolution of the ALV-J genome and contributes to theincreased pathogenicity of ALV-J not only in layers but also in broilers. Then, we found that theincreased replication of ALV-J increased the expression of VEGF-A and VEGFR-2for promotingtumorigenesis; ALV-J overcame the host p53restriction and facilitated viral replication, whichwas an important mechanism for pathogenicity. Finally, gga-miR-1650was identified to inhibitthe ALV-J replication. All these works contribute to clarify the stronger pathogenicity of ALV-J inChina and provides the basis for the development of new strategies for anti-ALV-J intervention.