Pneumopathogenicity of a Sendai virus protease-activation mutant, TCs, which is sensitive to trypsin and chymotrypsin

Abstract

A protease-activation mutant of Sendai virus, TCs, was isolated from a trypsin-resistant mutant, TR-5. TCs was activated in vitro by both trypsin and chymotrypsin. TCs was, however, less sensitive to trypsin and chymotrypsin than were the wild-type virus and TR-5, respectively. F protein of TCs had a single amino acid substitution at residue 114 from glutamine to arginine, resulting in the appearance of the new cleavage site for trypsin and the shift of the cleavage site for chymotrypsin. Activation of TCs in the lungs of mice occurred less efficiently than that of the wild type, and TCs caused a less severe pneumopathogenicity than did the wild-type virus, which supports our previous view that the in vitro trypsin sensitivity of Sendai virus can be a good indication of pneumopathogenicity in mice.Sendai virus penetrates host cells through adsorption of HANA protein to the cellular receptors (19,28) and fusion of the viral envelope with the plasma membrane which is mediated by F protein (5,19). Since envelope fusion activity of F protein is acquired by posttranslational cleavage by certain proteases inherent to host cells (3,14,15,(20)(21)(22)(23), the structure of F protein around the cleavage site is thought to be important for the virus to express infectivity and to determine host range and organ tropism. This view was verified by the fact that the wild-type virus whose F protein accepted the cleavage by trypsin next to arginine at residue 116 could multiply in the lungs of mice (24) in which trypsin like protease was present (23), while trypsin-resistant mutants TR-2 and TR-5 could not multiply because the arginine residue mutated to isoleucine and were not activated by the proteases in the lungs of mice (24, 25). The following report by Tashiro et al. (26) also supports the above-described view: a variant (F1-R) of Sendai virus which exhibited susceptibility to the activation by ubiquitous proteases as a result of the amino acid substitutions around the cleavage site and/or at the glycosylation site of the F2 subunit caused a systemic infection in mice.Recently, we isolated a variant of Sendai virus named TCs from the parent TR-5. As TCs was susceptible to both trypsin and chymotrypsin, we studied the mechanism of the activation of TCs by trypsin and the pneumopathogenicity of TCs in mice. The results obtained are presented in this report. The overall results will support our previous view that the in vitro trypsin sensitivity of Sendai virus correlates with pneumopathogenicity in mice.TCs was isolated by plaquing the trypsin-resistant mutant TR-5 in the presence of 3 ,ug of trypsin per ml instead of chymotrypsin for TR-5 in the agar overlay. The sensitivity of TCs to activation by trypsin and chymotrypsin was compared with those of TR-5, the wild-type virus of the Fushimi strain of Sendai virus, and trypsin-sensitive revertant TSrev (isolated from TR-5, also the parent of TCs, by passaging the virus in the presence of 1 pLg of trypsin per ml of the medium * Corresponding author. t Present address:[8]). TCs ...

Tryptase Clara, an activating protease for Sendai virus in rat lungs, is involved in pneumopathogenicity

“…The protease is localized exclusively in the nonciliated secretory cells, the so-called Clara cells, of the bronchial and bronchiolar epithelia of rats and is also secreted into the airway lumen. Tryptase Clara cleaves specifically single arginine residues of several peptides at neutral pH, and its spectrum of protease inhibitors is compatible with that suggested for the Sendai virus-activating protease(s) in mouse lungs (6,14,27,28,31). In addition, we have demonstrated that tryptase Clara cleaves the hemagglutinin of an influenza A virus and activates viral infectivity in vitro (17).…”

“…This may be explained by the apical budding phenotype of Sendai virus in the bronchial epithelial cells, the primary target of natural infections; spread of the progeny virus remains localized at the surface epithelia of the respiratory tract (32,33). Such a protease(s) for activation of Sendai virus has repeatedly been suggested to be present in the lungs as a key cellular determinant of pulmonary pathogenicity (8,14,26,30,31). This putative protease should also have characteristics similar, although not identical, to those of trypsin rather than those of chymotrypsin or elastase (7,13,27,28,31).…”

All Four Sendai Virus C Proteins Bind Stat1, but only the Larger Forms also Induce Its Mono-ubiquitination and Degradation

“…In many cases, the evasive measures specified by the viral genomes remain to be linked directly to the virulence of the pathogen in question, for lack of a relevant animal model. SeV, however, is a well-studied respiratory pathogen of lab mice (Itoh et al, 1990). The SeV C F170S mutation, moreover, was uncovered as one of two mutations (along with L E2050A ) in one of the most virulent virus strains reported to date (SeV M ; LD 50 of 40) that became avirulent (SeV MVC ; LD 50 of Ͼ800,000) on adaptation for growth in monkey kidney cells (Itoh et al, 1997).…”

Increased Induction of Apoptosis by a Sendai Virus Mutant Is Associated with Attenuation of Mouse Pathogenicity

“…In our previous work, we demonstrated that the pathogenicity of SeV closely correlates with virus replication in the mouse lung (35). We then showed that susceptibility of the fusion (F) envelope glycoprotein to trypsin and to the activating proteases in the mouse respiratory tract determines the efficiency of virus replication, and therefore the virulence, by supporting multiple-cycle replication through cleavage and activation of the F protein (16). Kato et al (20) reported that an SeV mutant lacking the V protein replicated less efficiently in the lungs of mice and was strongly attenuated.…”