title: Anti-lipopolysaccharide factor isoform 3 from Penaeus monodon (ALFPm3) exhibits antiviral activity by interacting with WSSV structural proteins.
abstract: In innate immunity, antimicrobial peptides (AMPs) play a vital role in combating microbial pathogens. Among the AMPs identified in Penaeus monodon, only anti-lipopolysaccharide factor isoform 3 (ALFPm3) has been reported to exhibit activity against white spot syndrome virus (WSSV). However, the mechanism(s) involved are still not clear. In the present study, ALFPm3-interacting proteins were screened for from a WSSV library using the yeast two-hybrid screening system, revealing the five potential ALFPm3-interacting proteins of WSSV186, WSSV189, WSSV395, WSSV458 and WSSV471. Temporal transcriptional analysis in WSSV-infected P. monodon revealed that all five of these WSSV gene transcripts were expressed in the late phase of infection (24h and 48h post-infection). Of these, WSSV189 that was previously identified as a structural protein, was selected for further analysis and was shown to be an enveloped protein by Western blot and immunoelectron microscopy analyses. The in vitro pull-down assay using recombinant WSSV189 (rWSSV189) protein as bait confirmed the interaction between ALFPm3 and WSSV189 proteins. Moreover, pre-incubation of rWSSV189 protein with rALFPm3 protein interfered with the latter's neutralization effect on WSSV in vivo, as shown by the increased cumulative mortality of shrimp injected with WSSV following prior treatment with pre-incubated rWSSV189 and rALFPm3 proteins compared to that in shrimp pre-treated with rALFPm3 protein. Thus, ALFPm3 likely performs its anti-WSSV action by binding to the envelope protein WSSV189 and possibly other WSSV structural proteins.
authors.affiliation: Center of Excellence for Molecular Biology and Genomics of Shrimp, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand.
authors.forename: Sivalee
authors.initials: S
authors.lastname: Suraprasit
authors.affiliation: Center of Excellence for Molecular Biology and Genomics of Shrimp, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand.
authors.forename: Thanachai
authors.initials: T
authors.lastname: Methatham
authors.affiliation: Center of Excellence for Molecular Biology and Genomics of Shrimp, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand.
authors.forename: Phattarunda
authors.initials: P
authors.lastname: Jaree
authors.affiliation: Center of Excellence for Shrimp Molecular Biology and Biotechnology, Mahidol University, Rama VI Rd., Bangkok 10400, Thailand; National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Pathumthani 12120, Thailand.
authors.forename: Kornsunee
authors.initials: K
authors.lastname: Phiwsaiya
authors.affiliation: Center of Excellence for Shrimp Molecular Biology and Biotechnology, Mahidol University, Rama VI Rd., Bangkok 10400, Thailand; National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency, Pathumthani 12120, Thailand.
authors.forename: Saengchan
authors.initials: S
authors.lastname: Senapin
authors.affiliation: Laboratory of Genome Science, Graduate School of Marine Science and Technology, Tokyo University of Marine Science and Technology, Tokyo, Japan.
authors.forename: Ikuo
authors.initials: I
authors.lastname: Hirono
authors.affiliation: Institute of Zoology, National Taiwan University, Taipei, Taiwan, Republic of China; Institute of Bioinformatics and Biosignal Transduction, College of Bioscience and Biotechnology, National Cheng Kung University, Tainan, Taiwan, Republic of China.
authors.forename: Chu Fang
authors.initials: CF
authors.lastname: Lo
authors.affiliation: Center of Excellence for Molecular Biology and Genomics of Shrimp, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand.
authors.forename: Anchalee
authors.initials: A
authors.lastname: Tassanakajon
authors.affiliation: Center of Excellence for Molecular Biology and Genomics of Shrimp, Department of Biochemistry, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand. Electronic address: kunlaya.s@chula.ac.th.
authors.forename: Kunlaya
authors.initials: K
authors.lastname: Somboonwiwat
mesh_terms: D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000494:pharmacology*; D000998:Antiviral Agents / Q000494:pharmacology; D060829:Arthropod Proteins / Q000494:pharmacology*; D033561:Penaeidae / Q000821:virology*; D011485:Protein Binding; D020033:Protein Isoforms / Q000494:pharmacology; D020798:Two-Hybrid System Techniques; D014759:Viral Envelope Proteins / Q000378:metabolism*; D046848:White spot syndrome virus 1 / Q000187:drug effects*
chemical_list: D023181:Antimicrobial Cationic Peptides; D000998:Antiviral Agents; D060829:Arthropod Proteins; D020033:Protein Isoforms; D014759:Viral Envelope Proteins; C034705:antilipopolysaccharide factor (Limulus)
keywords: Anti-lipopolysaccharide factor; Antiviral activity; Penaeus monodon; WSSV; Yeast two-hybrid screening