ABSTRACT
The open reading frame 117 (3h-117) of Heliothis virescens ascovirus 3h (HvAV-3h), which is a conserved coding region present in all completely sequenced ascovirus members, was characterized in this study. By RT-PCR detection, 3h-117 transcription began at 6-h post-infection (hpi) and remained stable until 168 hpi in HvAV-3h-infected Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) larvae. In addition, 3h-117 putatively encodes a 21.5-kDa protein (3H-117) predicted to be a CTD-like phosphatase. Western blot analysis using a prepared rabbit polyclonal antibody specific to 3H-117 showed that the product could be detected at 24 hpi, which remained stably detectable until 168 hpi. The same analysis also demonstrated that the 3H-117 protein localized in the virions of HvAV-3h. Immunofluorescence analysis showed that at 24 hpi, 3H-117 was mainly located in the nuclei of H. armigera larval fat body cells and later spread into the cytoplasm. In summary, our results indicate that 3H-117 is a structural protein of HvAV-3h.
Subject(s)
Ascoviridae/growth & development , Lepidoptera/virology , Transcription, Genetic , Viral Structural Proteins/biosynthesis , Animals , Ascoviridae/chemistry , Ascoviridae/genetics , Blotting, Western , Gene Expression Profiling , Larva/virology , Rabbits , Real-Time Polymerase Chain Reaction , Viral Structural Proteins/genetics , Virion/chemistryABSTRACT
Ascoviruses are enveloped, circular, double-stranded DNA viruses that can effectively control the appetite of lepidopteran larvae, thereby reducing the consequent damage and economic losses to crops. In this study, the virion of a sequenced Heliothis virescens ascovirus 3i (HvAV-3i) strain was used to perform proteomic analysis using both in-gel and in-solution digestion. A total of 81 viral proteins, of which 67 were associated with the virions, were identified in the proteome of HvAV-3i virions. Among these proteins, 23 with annotated functions were associated with DNA/RNA metabolism/transcription, virion assembly, sugar and lipid metabolism, signalling, cellular homoeostasis and cell lysis. Twenty-one viral membrane proteins were also identified. Some of the minor 'virion' proteins identified may be non-virion contaminants of viral proteins synthesized during replication, identified by more recent and highly sensitive methods. The extensive identification of the ascoviral proteome will establish a foundation for further investigation of ascoviral replication and infection.
Subject(s)
Ascoviridae/chemistry , Proteome/analysis , Viral Proteins/analysis , Virion/chemistry , Computational Biology , ProteomicsABSTRACT
Ascoviruses are a family of insect viruses with circular, double-stranded DNA genomes. With the sequencing of the Trichoplusia ni ascovirus 2c (TnAV-2c) genome, the virion structural proteins were identified by using tandem mass spectrometry. From at least eight protein bands visible on a Coomassie blue-stained gel of TnAV-2c virion proteins, seven bands generated protein sequences that matched predicted open reading frames (ORFs) in the genome, i.e. ORFs 2, 43, 115, 141, 142, 147 and 153. Among these ORFs, only ORF153, encoding the major capsid protein, has been characterized previously.
