Identification and characterization of an interleukin-16-like gene from pacific white shrimp Litopenaeus vannamei.

Interleukins are a group of cytokines that play essential roles in immune regulation. Almost all interleukin genes are only found in vertebrates. In this study, an interleukin-16-like gene (LvIL-16L) was identified from Pacific white shrimp, Litopenaeus vannamei. LvIL-16L was predicted to encode a precursor (pro-LvIL-16L) with 1378 amino acids, sharing similarities with predicted pro-IL-16-like proteins from insects. The C-terminus of pro-LvIL-16L protein contained two PDZ domains homologous to the mature IL-16 cytokine of vertebrates. In tissues, LvIL-16L could be processed into a ∼36 kDa mature peptide through a caspase-3 cleavage site, which was verified by in vitro site mutation analysis and in vivo RNA interference (RNAi) experiments. The LvIL-16L mRNA could be detected in all the analyzed tissues and the expression of LvIL-16L was significantly up-regulated after immune stimulation. Using RNAi strategy, the role of LvIL-16L in immune responses was initially investigated. Interestingly, knockdown of LvIL-16L could significantly increase the mortality of the Vibro parahaemolyticus infected shrimps but reduce that of the WSSV infected shrimps, suggesting that LvIL-16L could have opposite effects on the antiviral and antibacterial immune responses in shrimp. To our knowledge, this is the first study of an IL-16-like gene in invertebrates, which could help to elucidate interleukin evolution and regulatory mechanisms of shrimp immune responses.

Do shallow soil, low water availability, or their combination increase the competition between grasses with different root systems in karst soil?

Uneven soil depth and low water availability are the key limiting factors to vegetation restoration and reconstruction in limestone soils such as in vulnerable karst regions. Belowground competition will possibly increase under limited soil resources. Here, we investigate whether low resource availability (including shallow soil, low water availability, and shallow soil and low water availability combined) stimulates the competition between grasses with different root systems in karst soil, by assessing their growth response, biomass allocation, and morphological plasticity. In a full three-way factorial blocked design of soil depth by water availability by neighbor identity, we grew Festuca arundinacea (deep-rooted) and Lolium perenne (shallow-rooted) under normal versus shallow soil depth, high versus low water availability, and in monoculture (conspecific neighbor) versus mixture (neighbor of the other species). The key results were as follows: (1) total biomass and aboveground biomass in either of the species decreased with reduction of resources but were not affected by planting patterns (monoculture or mixture) even at low resource levels. (2) For F. arundinacea, root biomass, root mass fraction, total root length, and root volume were higher in mixture than in monoculture at high resource level (consistent with resource use complementarity), but lower in mixture than in monoculture at low resource levels (consistent with interspecific competition). In contrast for L. perenne, either at high or low resource level, these root traits had mostly similar values at both planting patterns. These results suggest that deep-rooted and shallow-rooted plant species can coexist in karst regions under current climatic regimes. Declining resources, due to shallow soil, a decrease in precipitation, or combined shallow soil and karst drought, increased the root competition between plants of deep-rooted and shallow-rooted species. The root systems of deep-rooted plants may be too small to get sufficient water and nutrients from dry, shallow soil, while shallow-rooted plants will maintain a dominant position with their already adaptive strategy in respect of root biomass allocation and root growth.

A mitochondrial outer membrane-localized protein encoded by White spot syndrome virus.

White spot syndrome virus (WSSV) is a devastating pathogen of crustaceans that causes huge losses to global shrimp farming. In this study, a mitochondrial outer membrane protein encoded by WSSV, termed as wsv152, was identified. Bioinformatics analysis showed that wsv152 shared no similarity with any known proteins. Real-time RT-PCR revealed that wsv152 was initially transcribed at 6 h post WSSV infection. Fluorescent microscopy demonstrated that the wsv152-GFP fusion protein was present in the mitochondrion. Western blot further suggested that wsv152 was specifically localized on the outer membrane of mitochondria. To our knowledge, this is the first report of a mitochondrion-localized protein encoded by WSSV.

Daxx from Pacific white shrimp Litopenaeus vannamei is involved in activation of NF-κB pathway.

Death domain-associated factor 6 (Daxx) is a Fas-binding protein that mediates the activation of Jun amino-terminal kinase (JNK) pathway and Fas-induced apoptosis. In this study, a crustacean Daxx (LvDaxx) was firstly cloned and identified from Pacific white shrimp Litopenaeus vannamei. The LvDaxx cDNA was 2644 bp in length with an Open Reading Frame (ORF) of 2217 bp. Sequence analysis indicated that LvDaxx contained a single Daxx domain and two nuclear localization signals (NLSs) and shared a similarity with Drosophila melanogaster Daxx. LvDaxx was a nuclear-localized protein that was expressed highest in hemocytes and could be up-regulated in pathogen- and stimulant-challenge shrimps. LvDaxx could activate the artificial promoter containing an NF-κB binding site and the promoters of white spot syndrome virus (WSSV) ie1 gene and arthropod antimicrobial peptides (AMPs), suggesting LvDaxx could be involved in the activation of the NF-κB pathway. Knock-down of LvDaxx in vivo resulted in down-regulation of shrimp AMPs and reduction of WSSV copies in tissues. Furthermore, suppression of LvDaxx significantly decreased the mortality of WSSV-infected shrimps, but increased the mortality of Vibrio Parahaemolyticus-infected shrimps. Thus, these suggested that LvDaxx could play a role in the innate immunity against Vibrio parahaemolyticus in L. vannamei, while in the antiviral response, LvDaxx may be hijacked by WSSV and play a complex role in WSSV pathogenesis.

Identification of a JAK/STAT pathway receptor domeless from Pacific white shrimp Litopenaeus vannamei.

The Janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling pathway was known to participate in dozens of immune responses in organisms. Domeless, first identified in Drosophila melanogaster, is a unique receptor involved in invertebrate JAK/STAT pathway. In this study, a cytokine receptor (LvDOME) was identified in Litopenaeus vannamei. The LvDOME cDNA was 5178bp in length with an Open Reading Frame (ORF) of 4191bp. LvDOME contained two cytokine binding modules (CBMs) and three fibronectin-type-III-like (FNIII) domains, similar to most vertebrate IL-6 receptors. LvDOME was expressed highest in shrimp muscle and could be up-regulated in the late stage of white spot syndrome virus (WSSV) infection. LvDOME could significantly enhance the activity of the WSSV wsv069 gene promoter through acting on the STAT-binding motif, suggesting LvDOME could activate the JAK/STAT pathway. Moreover, knockdown of LvDOME resulted in lower cumulative mortality of shrimps and less WSSV copies, suggesting LvDOME may be hijacked by WSSV to benefit virus replication. To our knowledge, this is the first report on the receptor of JAK/STAT pathway in shrimp.