RESUMO
The LaSota strain of Newcastle disease virus (NDV) is a commonly used vaccine to control Newcastle disease. However, improper immunization is a common reason for vaccine failure. Hence, it is imperative to thoroughly explore innate immunity-related molecular regulatory responses to the LaSota vaccine. In this text, 140 long non-coding RNAs (lncRNAs), 8 microRNAs (miRNAs), and 1514 mRNAs were identified to be differentially expressed by RNA sequencing analysis in the thymic tissues of Chinese Partridge Shank chickens after LaSota vaccine inoculation. Moreover, 70 dysregulated genes related to innate immunity were identified based on GO, Reactome pathway, and InnateDB annotations and differential expression analysis. Additionally, dysregulated lncRNAs and innate immunity-related mRNAs that could interact with dysregulated miRNAs were identified based on bioinformatics prediction analysis via the miRanda software and differential expression analysis. Among these transcripts, expression patterns of five lncRNAs, seven miRNAs, and six mRNAs were further examined by RT-qPCR assay. Both RNA-seq and RT-qPCR outcomes showed that 10 transcripts (MSTRG.22689.1, ENSGALT00000065826, ENSGALT00000059336, ENSGALT00000060887, gga-miR-6575-5p, gga-miR-6631-5p, gga-miR-1727, paraoxonase 2 (PON2), mitogen-activated protein kinase 10, and cystic fibrosis transmembrane conductance regulator (CFTR) were highly expressed, and 4 transcripts (MSTRG.188121.10, gga-miR-6655-5p, gga-miR-6548-3p, and matrix metallopeptidase 9 (MMP9) were low expressed after NDV infection. Additionally, two potential competing endogenous RNA networks (ENSGALT00000060887/gga-miR-6575-5p/PON2 or MSTRG.188121.10/gga-miR-6631-5p/MMP9) and some co-expression axes (ENSGALT00000065826/gga-miR-6631-5p, MSTRG.188121.10/gga-miR-6575-5p, MSTRG.188121.10/CFTR, ENSGALT00000060887/MMP9) were identified based on RT-qPCR and co-expression analyses. In conclusion, we identified multiple dysregulated lncRNAs, miRNAs, and mRNAs after LaSota infection and some potential regulatory networks for these dysregulated transcripts.
Assuntos
MicroRNAs , RNA Longo não Codificante , Vacinas , Animais , Galinhas/genética , Galinhas/metabolismo , China , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Metaloproteinase 9 da Matriz/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Vírus da Doença de Newcastle/genética , Vírus da Doença de Newcastle/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transcriptoma/genética , Vacinas/metabolismoRESUMO
Newcastle disease (ND), caused by virulent Newcastle disease virus (NDV), is a contagious viral disease affecting various birds and poultry worldwide. In this project, differentially expressed (DE) circRNAs, miRNAs and mRNAs were identified by high-throughput RNA sequencing (RNA-Seq) in chicken thymus at 24, 48, 72 or 96 h post LaSota NDV vaccine injection versus pre-inoculation group. The vital terms or pathways enriched by vaccine-influenced genes were tested through KEGG and GO analysis. DE genes implicated in innate immunity were preliminarily screened out through GO, InnateDB and Reactome Pathway databases. The interaction networks of DE innate immune genes were established by STRING website. Considering the high expression of gga-miR-6631-5p across all the four time points, DE circRNAs or mRNAs with the possibility to bind to gga-miR-6631-5p were screened out. Among DE genes that had the probability to interact with gga-miR-6631-5p, 7 genes were found to be related to innate immunity. Furthermore, gga-miR-6631-5p promoted LaSota NDV replication by targeting insulin induced gene 1 (INSIG1) in DF-1 chicken fibroblast cells. Taken together, our data provided the comprehensive information about molecular responses to NDV LaSota vaccine in Chinese Partridge Shank Chickens and elucidated the vital roles of gga-miR-6631-5p/INSIG1 axis in LaSota NDV replication.
Assuntos
Doença de Newcastle/genética , Pequeno RNA não Traduzido/genética , Replicação Viral/genética , Animais , Galinhas/genética , Galinhas/virologia , Expressão Gênica/genética , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica/genética , Imunidade Inata , MicroRNAs/genética , Doença de Newcastle/virologia , Vírus da Doença de Newcastle/genética , Vírus da Doença de Newcastle/patogenicidade , Doenças das Aves Domésticas/virologia , RNA Circular/genética , RNA Mensageiro/genética , Timo/metabolismo , Timo/virologia , Transcriptoma/genética , VacinaçãoRESUMO
Newcastle disease virus (NDV) is a contagious poultry paramyxovirus, leading to substantial economic losses to the poultry industry. Here, RNA-seq was carried out to investigate the altered expression of immune-related genes in chicken thymus within 96 h in response to NDV infection. In NDV-infected chicken thymus tissues, comparative transcriptome analysis revealed 1386 differentially expressed genes (DEGs) at 24 h with 989 up- and 397 down-regulated genes, 728 DEGs at 48 h with 567 up- and 161 down-regulated genes, 1514 DEGs at 72 h with 1016 up- and 498 down-regulated genes, and 1196 DEGs at 96 h with 522 up- and 674 down-regulated genes, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that these candidate targets mainly participate in biological processes or biochemical, metabolic and signal transduction processes. Notably, there is large enrichment in biological processes, cell components and metabolic processes, which may be related to NDV pathogenicity. In addition, the expression of five immune-related DEGs identified by RNA-seq was validated by quantitative real-time polymerase chain reaction (qRT-PCR). Our results indicated that the expression levels of AvBD5, IL16, IL22 and IL18R1 were obviously up-regulated, and Il-18 expression was also changed, but not significantly, which play key roles in the defense against NDV. Overall, we identified several candidate targets that may be involved in the regulation of NDV infection, which provide new insights into the complicated regulatory mechanisms of virus-host interactions, and explore new strategies for protecting chickens against the virus.
