Design and Construction of Shrimp Antiviral DNA Vaccines Expressing Long and Short Hairpins for Protection by RNA Interf
DNA vaccines present the aquaculture industry with an effective and economically viable method of controlling viral pathogens that drastically affect productivity. Since specific immune response is rudimentary in invertebrates, the presence of RNA interfe
- PDF / 381,984 Bytes
- 16 Pages / 504.57 x 720 pts Page_size
- 74 Downloads / 151 Views
1
Introduction DNA vaccines are essentially recombinant plasmid constructs capable of expressing pathogen-derived antigenic proteins that prime the host against future infection when administered intramuscularly or subcutaneously [1–3]. DNA vaccines present the aquaculture industry with an effective and economically viable method of checking the threat of various pathogens that drastically affect productivity. They are considered safer compared to live, attenuated, and whole inactivated vaccines and are more stable than protein/glycoprotein subunit vaccines. However, it is for the viral and parasitic diseases that they are particularly attractive options [4]. The DNA vaccine against infectious hematopoietic necrosis virus (IHNV) that affects salmonid fishes is most effective and the only one licensed for use in aquaculture since 2005 [5]. In invertebrates like shrimps, however, the specific immune response system is rudimentary [6] and although there are some reports on application of subunit vaccines [7, 8] and DNA vaccines expressing viral proteins [9], they are of limited efficacy. The discovery of RNA interference (RNAi) pathway in shrimps provided a promising new approach to vaccination, and in current times, the definition of DNA vaccines can be extended to include plasmid constructs that express short or long double stranded RNA (dsRNA) in the host and inhibit pathogen proliferation through RNA interference mechanism. In a dramatic discovery in 1998 it was found that dsRNA introduced into a eukaryotic cell results in silencing of the corresponding RNA transcript [10], a phenomenon that has been named
Sunil Thomas (ed.), Vaccine Design: Methods and Protocols, Volume 2: Vaccines for Veterinary Diseases, Methods in Molecular Biology, vol. 1404, DOI 10.1007/978-1-4939-3389-1_16, © Springer Science+Business Media New York 2016
225
226
Aparna Chaudhari et al.
“posttranscriptional gene silencing” (PTGS) or “RNA interference” (RNAi). The presence of dsRNA in the cytoplasm (whether it is transfected or synthesized within the cell) triggers the multidomain ribonuclease III enzyme Dicer [11]. This cleaves dsRNA into small interfering RNAs (siRNAs), which are 21–23 nucleotide fragments with characteristic 2-nucleotide 3′ overhangs. These siRNAs are recognized by the RNA-Induced Silencing Complex (RISC; [12]), a multienzyme unit that brings about separation of the two siRNA strands. The antisense siRNA strand remains bound to RISC, while the sense strand is released. In some organisms that have functional RNA-dependent RNA polymerase (RdRp) enzyme, the sense strand may be again converted into dsRNA [13]. The antisense strand guides RISC to bind the homologous (target) mRNA, and another RNase III Argonaute that is part of the complex cleaves it, silencing its expression [14]. The efficiency of siRNA depends on perfect complementarity of the seed sequence (positions 2–6) with the target mRNA. It has been reported that imperfect base pairing that creates a bulge in miRNA/siRNA marks the transcript for translational r
Data Loading...
