The 2'-5'-oligoadenylate synthetase in the lowest metazoa: isolation, cloning, expression and functional activity in the sponge Lubomirskia baicalensis.
The 2'-5'-oligoadenylate synthetase in the lowest metazoa: isolation, cloning, expression and functional activity in the sponge Lubomirskia baicalensis.
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DOI：
10.1016/j.molimm.2007.07.036
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发表时间：
2008-02
影响因子：
3.6
通讯作者：
H. Schröder;F. Natalio;M. Wiens;M. Tahir;M. Shukoor;W. Tremel;S. Belikov;A. Krasko;W. Müller
中科院分区：
文献类型：
--
作者：
H. Schröder;F. Natalio;M. Wiens;M. Tahir;M. Shukoor;W. Tremel;S. Belikov;A. Krasko;W. Müller
水生动物，特别是海绵等滤食性动物，比陆生物种接触到更高的病毒载量。到目前为止，海绵这种进化最古老的多细胞生物的抗病毒防御系统还不清楚。干扰素诱导的2‘-5’-寡腺苷合成酶[(2-5)A合成酶]系统是脊椎动物抵抗病毒感染的一种强大的保护机制。在本研究中，我们从淡水海绵中克隆了编码314个氨基酸的开放阅读框(ORF)的基因，计算大小为35748Da，推测是一个(2-5)A合成酶，并对该重组蛋白进行了抗体诱导。应用一种新的基于聚合物包被的铁磁性纳米颗粒的分离方法，从黄芩的粗提物中鉴定了天然酶。这些颗粒被合成的双链RNA[dsRNA]衍生，合成的聚(I：C)是一种已知的潜伏的(2-5)A合成酶的变构激活剂。这些颗粒被用来从黄卡尔氏乳杆菌的粗提物中分离出单一的35 kDa蛋白质，该粗提物与抗海绵酶的抗体发生交叉反应。原位杂交研究表明，该基因在含水运河周围的细胞中表达最高。最后，将黄加乳杆菌的体外细胞培养系统Primphs暴露于Poly(I：C)中；它们对这种dsRNA的反应是，在1天的孵育期后，(2-5)A合成酶基因的表达已经增加。我们得出结论，海绵含有(2-5)A合成酶抗病毒保护系统。
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.