The 2'-5'-oligoadenylate synthetase in the lowest metazoa: isolation, cloning, expression and functional activity in the sponge Lubomirskia baicalensis.
Aquatic animals, especially filter feeders such as sponges [phylum Porifera], are exposed to a higher viral load than terrestrial species. Until now, the antiviral defense system in the evolutionary oldest multicellular organisms, sponges, is not understood. One powerful protection of vertebrates against virus infection is mediated by the interferon (IFN)-inducible 2'-5'-oligoadenylate synthetase [(2-5)A synthetase] system. In the present study we cloned from the freshwater sponge Lubomirskia baicalensis a cDNA encoding a 314 aa long ORF with a calculated size of 35748Da, a putative (2-5)A synthetase, and raised antibodies against the recombinant protein. The native enzyme was identified in a crude extract from L. baicalensis by application of a novel separation procedure based on polymer coated ferromagnetic nanoparticles. The particles were derivatized with a synthetic double-stranded RNA [dsRNA], synthetic poly(I:C), a known allosteric activator of the latent (2-5)A synthetase. These particles were used to separate a single 35kDa protein from a crude extract of L. baicalensis, which cross-reacted with antibodies raised against the sponge enzyme. In situ hybridization studies revealed that highest expression of the gene is seen in cells surrounding the aquiferous canals. Finally primmorphs, an in vitro cell culture system, from L. baicalensis were exposed to poly(I:C); they responded to this dsRNA with an increased expression of the (2-5)A synthetase gene already after a 1-day incubation period. We conclude that sponges contain the (2-5)A synthetase antiviral protection system.
Schröder
Heinz C
HC
Institut für Physiologische Chemie, Abteilung Angewandte Molekularbiologie, Universität, Duesbergweg 6, D-55099 Mainz, Germany.
Natalio
Filipe
F
Wiens
Matthias
M
Tahir
Muhammad Nawaz
MN
Shukoor
Mohammed Ibrahim
MI
Tremel
Wolfgang
W
Belikov
Sergey I
SI
Krasko
Anatoli
A
Müller
Werner E G
WE
D015088:2',5'-Oligoadenylate Synthetase / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D004789:Enzyme Activation; D017403:In Situ Hybridization; D008969:Molecular Sequence Data; D011070:Poly I-C / Q000737:chemistry / Q000494:pharmacology; D011161:Porifera / Q000166:cytology / Q000201:enzymology*; D012330:RNA, Double-Stranded / Q000737:chemistry / Q000494:pharmacology
D018076:DNA, Complementary; D012330:RNA, Double-Stranded; D015088:2',5'-Oligoadenylate Synthetase; D011070:Poly I-C