Non-specific activation of antiviral immunity and induction of RNA interference may engage the same pathway in the Pacific white leg shrimp Litopenaeus vannamei

Abstract

Many questions remain unanswered regarding RNAi-based mechanisms and dsRNA-induced antiviral immune responses in penaeid shrimp. In this study, we report the characterization in the white leg shrimp Litopenaeus vannamei of RNAi pathway associated proteins Lv-Ago 1 and Lv-Ago 2, two members of the Argonaute family of proteins, as well as Lv-sid 1, the first shrimp homologue of Sid-1, a membrane channel-forming protein implicated in the cellular import of dsRNA. To decipher their functional implication in RNAi-related phenomena, we monitored their relative expression following stimulation by specific and non-specific RNA duplexes of diverse length. The findings show that the length of small RNA duplexes plays a critical role in the activation of both RNAi-related and innate antiviral responses. They also suggest that these two mechanisms of antiviral response may activate the same pathway, requiring Lv-Sid 1 and Lv-Ago 2 induction.

Isolation and characterization of cDNAs encoding Ars2 and Pasha homologues, two components of the RNA interference pathway in Litopenaeus vannamei

“…For shrimp, the siRNA pathway participates in special antiviral immunity as well as non-special antiviral immunity [29,30,44]. It also appears that shrimp can exploit the miRNA pathway to evade antiviral responses [30,33,34]. In shrimp, the gill is an important immune tissue and has been considered as the target tissue in some studies about shrimp RNAi pathway [29,30,33,34,44].…”

“…On one hand, the miRNA pathway could participate in antiviral immunity by disrupting foreign nucleic acids, on the other hand, the host miRNA pathway is also used by some viruses to evade the host immune system [20,41e43]. For shrimp, the siRNA pathway participates in special antiviral immunity as well as non-special antiviral immunity [29,30,44]. It also appears that shrimp can exploit the miRNA pathway to evade antiviral responses [30,33,34].…”

“…For the Poly(CeG) (SigmaeAldrich) stimulation experiment, Poly(CeG) was diluted to 2.5 mg/uL in 400 mM NaCl and 10 mM Tris-HCL (pH7.4) [31], and each healthy L. vannamei was injected at the second abdominal segment with 30 mL diluted Poly(CeG). Total RNA of the gill was isolated immediately or at 3, 6,9,12,18,24,30,36,48, or 72 h after injection. All total RNA samples were extracted using the RNeasy Mini Kit (Qiagen, Germany) and subsequently reverse transcribed to cDNA using PrimeScript RT Reagent Kit (TaKaRa, Japan) according to the manufacturer's instructions.…”

“…Knockdown of Penaeus monodon Dcr1 (PmDcr1) resulted in more rapid mortality and higher viral loads after gill-associated virus (GAV) infection [33]; L. vannamei Dcr1 (LvDcr1) mRNA was upregulated in hemocytes and gills after TSV injection [34]; Marsupenaeus japonicus Dcr1 (MjDcr1) has not yet been studied in detail. Other components of the shrimp RNAi pathway, such as Fenneropenaeus chinensis transactivating response RNA-binding protein 1e3 (FcTRBP1e3), L. vannamei Argonaute 2 (LvAgo2), L. vannamei systemic RNA interference-deficient (LvSID-1) and P. monodon Argonaute 1 (PmAgo1), were also shown to participate in antiviral immunity [30,35,36].…”

Enhancement of shrimp immunity against white spot syndrome virus by Macrobrachium rosenbergii nodavirus-like particle encapsulated VP28 double-stranded RNA

“…These findings indicate the natural existence of intact RNAi machinery within shrimp tissues. To date, two key protein components of RNAi machinery -dicer and argonaute -have been identified in Penaeus shrimp and experimentally proven to play an important role in RNA-induced silencing complex (RISC) (Chen et al, 2011;Labreuche et al, 2010;Yao et al, 2010).…”