Transcriptome sequencing of and microarray development for a Helicoverpa zea cell line to investigate in vitro insect cell-baculovirus interactions

• Published in: PloS one Vol. 7; no. 5; p. e36324
• Main Authors: Nguyen, Quan, Palfreyman, Robin W, Chan, Leslie C L, Reid, Steven, Nielsen, Lars K
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 18.05.2012; Public Library of Science (PLoS)
• Subjects: Acids; Agriculture; Analysis; Animals; Apoptosis; Baculoviridae - genetics; Baculovirus; Bayesian analysis; Bioengineering; Biological control; Biological pest control; Biology; Cell culture; Cell Line; DNA microarrays; Ecology; Gene expression; Gene regulation; Gene sequencing; Genes; Genomes; Genomics; Health aspects; Helicoverpa zea; Host range; Immune response; Immune system; Immunology; Infections; Infectivity; Informatics; Insects; Integrated software; Investigations; JNK protein; Metabolism; Microarray Analysis; Molecular Sequence Data; Moths - genetics; Moths - virology; Nanotechnology; Pathways; Pests; Proteins; Ribonucleic acid; RNA; Sequence Analysis, DNA; Signal transduction; Signaling; Studies; Transcription; Transcriptome; Up-Regulation; Virulence; Viruses; St Lucia
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0036324

The Heliothine insect complex contains some of the most destructive pests of agricultural crops worldwide, including the closely related Helicoverpa zea and H. armigera. Biological control using baculoviruses is practiced at a moderate level worldwide. In order to enable more wide spread use, a better understanding of cell-virus interactions is required. While many baculoviruses have been sequenced, none of the Heliothine insect genomes have been available. In this study, we sequenced, assembled and functionally annotated 29,586 transcripts from cultured H. zea cells using Illumina 100 bps and paired-end transcriptome sequencing (RNA-seq). The transcript sequences had high assembly coverage (64.5 times). 23,401 sequences had putative protein functions, and over 13,000 sequences had high similarities to available sequences in other insect species. The sequence database was estimated to cover at least 85% of all H. zea genes. The sequences were used to construct a microarray, which was evaluated on the infection of H. zea cells with H. Armigera single-capsid nucleopolyhedrovirus (HearNPV). The analysis revealed that up-regulation of apoptosis genes is the main cellular response in the early infection phase (18 hours post infection), while genes linked to four major immunological signalling pathways (Toll, IMD, Jak-STAT and JNK) were down-regulated. Only small changes (generally downwards) were observed for central carbon metabolism. The transcriptome and microarray platform developed in this study represent a greatly expanded resource base for H. zea insect-HearNPV interaction studies, in which key cellular pathways such as those for metabolism, immune response, transcription and replication have been identified. This resource will be used to develop better cell culture-based virus production processes, and more generally to investigate the molecular basis of host range and susceptibility, virus infectivity and virulence, and the ecology and evolution of baculoviruses.