PROTEOLYTIC DIGESTION OF REOVIRUS IN THE INTESTINAL LUMENS OF NEONATAL MICE
PROTEOLYTIC DIGESTION OF REOVIRUS IN THE INTESTINAL LUMENS OF NEONATAL MICE
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DOI：
10.1128/jvi.63.11.4676-4681.1989
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发表时间：
1989-11-01
影响因子：
5.4
通讯作者：
FIELDS, BN
中科院分区：
文献类型：
--
作者：
BODKIN, DK;NIBERT, ML;FIELDS, BN
用两种方法证明了呼肠孤病毒在新生小鼠肠道中的蛋白分解。第一种方法是将放射性标记病毒经口接种到新生小鼠体内；在接种后0-30分钟采集肠道冲洗液。从肠道洗涤中回收的病毒在聚丙烯酰胺中进行电泳，以确定病毒蛋白是否发生了蛋白水解性消化。Sigma 3的完全丧失和1c裂解产物Delta的产生表明，在接种后10-30分钟内发生消化，导致中间亚病毒颗粒(ISVP)的快速产生。形成的产物类似于用胰凝乳酶在体外消化病毒时看到的产物。第二种方法利用了ISVP在NH4Cl处理的细胞中生长的事实，而完整病毒在这些条件下不生长(L.J.Sturzenbecker，M.Nibert，D.Furlong和B.N.Fields，J.Virol)。61：2351-2361,1987)。因此，检测病毒在NH4Cl处理的细胞中的生长能力是确定样本中是否含有ISVP的一种手段。使用这种方法，我们证明了在接种后8h，ISVPs在肠道组织和肠腔中占主导地位。在接种后8~15h，组织中ISVPs的比例下降，因此在接种后15h，在洗去腔内容物和病毒的肠道组织中不再检测到ISVPs。相反，肠腔在接种后的任何时候都含有一些ISVP。因此，经口接种后，哺乳动物呼肠孤病毒转化为蛋白水解性裂解病毒，这表明蛋白水解性在胃肠道感染的起始中起着重要作用。
Two approaches were used to demonstrate proteolysis of reovirus in the intestine of the neonatal mouse. The first approach utilized peroral inoculation of radiolabeled virus into neonatal mice; the intestinal washings were harvested at 0 to 30 min postinoculation. The virus recovered from the intestinal washings was electrophoresed in polyacrylamide to determine whether proteolytic digestion of viral proteins had occurred. Complete loss of sigma 3 and generation of the .mu.1c cleavage product delta demonstrated that digestion occurred within 10 to 30 min after the inoculation, resulting in the rapid generation of intermediate subviral particles (ISVPs). The products formed resembled those seen when the virus is digested in vitro with chymotrypsin. The second approach took advantage of the fact that ISVPs grow in cells treated with NH4Cl, whereas intact virus does not grow under these conditions (L. J. Sturzenbecker, M. Nibert, D. Furlong, and B. N. Fields, J. Virol. 61:2351-2361, 1987). Thus, assaying virus for its ability to grow in NH4Cl-treated cells represents a means of ascertaining whether the samples contain ISVPs. Using this approach, we demonstrated that up to 8 h postinoculation ISVPs predominate in the intestinal tissue and in the intestinal lumen. Between 8 and 15 h postinoculation, there is a loss in the proportion of ISVPs in the tissue so that by 15 h postinoculation ISVPs are no longer detectable in intestinal tissue washed of lumen contents and virus. In contrast, the lumen of the intestine contains some ISVPs at all times postinoculation. Thus, after peroral inoculation, the mammalian reoviruses are converted to proteolytically cleaved virus, suggesting that proteolysis plays an important role in initiation of infection in the gastrointestinal tract.