Identification and functional characterization of Dicer2 and five single VWC domain proteins of Litopenaeus vannamei

Dev Comp Immunol. 2011 Jun;35(6):661-71. doi: 10.1016/j.dci.2011.01.010. Epub 2011 Jan 20.

Abstract

Dicer (Dcr) is the key protein of the RNA interference (RNAi) pathway. To investigate the role of the RNAi pathway in shrimp anti-viral immunity, Litopenaeus vannamei Dcr2 (designated as LvDcr2) was identified and characterized. The full-length cDNA of LvDcr2 was 5513bp long, with an open reading frame encoding a putative protein of 1502 amino acids. In addition, five proteins homologous to the single von Willebrand factor type C (VWC) domain protein (SVC) were also identified in L. vannamei and named LvSVC1-5. These LvSVCs were between 102 and 190 amino acids in length and all contained a motif similar to Drosophila melanogaster SVC proteins (DmSVCs). By co-immunoprecipitation assays and pull-down assays, we demonstrated that LvDcr2, L. vannamei Argonaute 2 (LvAgo2), and L. vannamei transactivating response RNA-binding protein isoform 1 (LvTRBP1) interacted with each other. A luciferase reporter assay indicated that the promoters of LvSVC1, LvSVC4, LvSVC5, and DmSVC Vago (DmVago) were activated by LvDcr2 as well as by Drosophila Dcr2 (DmDcr2). Real-time RT-PCR showed that LvDcr2 and LvSVCs were up-regulated in immune responses against Poly(C-G) or WSSV challenge. These results suggested that LvDcr2 formed complexes with LvAgo2 and LvTRBP1 to act as the cores of shrimp small interfering RNA (siRNA)-induced silencing complex (siRISC)/siRISC-loading complex (siRLC), role in shrimp siRNA pathway. Furthermore, these results also suggested that LvDcr2 may engage in non-specific activation of anti-viral immunity.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs
  • Amino Acid Sequence
  • Animals
  • Biological Factors / genetics*
  • Biological Factors / immunology
  • Biological Factors / metabolism
  • Cloning, Molecular
  • Genes, Reporter
  • Immunoprecipitation
  • Luciferases, Renilla / biosynthesis
  • Luciferases, Renilla / genetics
  • Molecular Sequence Data
  • Organ Specificity
  • Penaeidae / enzymology*
  • Penaeidae / immunology
  • Penaeidae / virology
  • Phylogeny
  • Promoter Regions, Genetic
  • Protein Binding
  • Protein Sorting Signals
  • Protein Structure, Tertiary*
  • RNA-Binding Proteins / metabolism
  • Ribonuclease III / chemistry
  • Ribonuclease III / genetics*
  • Ribonuclease III / metabolism
  • Transcription, Genetic
  • Up-Regulation
  • White spot syndrome virus 1
  • von Willebrand Factor / chemistry
  • von Willebrand Factor / metabolism

Substances

  • Biological Factors
  • Protein Sorting Signals
  • RNA-Binding Proteins
  • von Willebrand Factor
  • trans-activation responsive RNA-binding protein
  • Luciferases, Renilla
  • Ribonuclease III

Associated data

  • GENBANK/HQ541157
  • GENBANK/HQ541158
  • GENBANK/HQ541159
  • GENBANK/HQ541160
  • GENBANK/HQ541161
  • GENBANK/HQ541162
  • GENBANK/HQ541163