Dataset Information

Characterizing the PRRSV nsp2 Deubiquitinase Reveals Dispensability of Cis-Activity for Replication and a Link of nsp2 to Inflammation Induction.

ABSTRACT: The papain-like cysteine protease 2 (PLP2) within the N-terminus of the porcine reproductive and respiratory syndrome virus (PRRSV) nsp2 replicase protein specifies a deubiquitinating enzyme (DUB), but its biochemical properties and the role in infection have remained poorly defined. By using in vitro assays, we found that the purified PLP2 could efficiently cleave K63 and K48 linked polyubiquitin chains Ub3-7 in vitro although displaying a differential activity in converting the respective ubiquitin dimers to monomer. The subsequent mutagenesis analyses revealed that the requirement for PLP2 DUB activity surprisingly resembled that for cis-cleavage activity, as several mutations (e.g., D91R, D85R, etc.) that largely ablated the DUB function also blocked the cis- but not trans-proteolytic cleavage of nsp2/3 polyprotein. Moreover, the analyses identified key mutations that could differentiate DUB from PLP2 cis- and trans-cleavage activities. Further reverse genetics analyses revealed the following findings: (i) mutations that largely blocked the DUB activity were all lethal to the virus, (ii) a point mutation T88G that selectively blocked the cis-cleavage activity of PLP2 did not affect viral viability in cell culture, and (iii) an E90Q mutation that did not affect either of the PLP2 activities led to rescue of WT-like virus but displayed significantly reduced ability to induce TNF-? production. Our findings support the possibility that the PLP2 DUB activity, but not cis-cleavage activity, is essential for PRRSV replication. The data also establish a strong link of nsp2 to pro-inflammatory cytokine induction during infection that operates in a manner independent of PLP2 DUB activity.

SUBMITTER: Zhou S

PROVIDER: S-EPMC6832237 | biostudies-literature | 2019 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Characterizing the PRRSV nsp2 Deubiquitinase Reveals Dispensability of <i>Cis</i>-Activity for Replication and a Link of nsp2 to Inflammation Induction.

Zhou Shaochuan S Ge Xinna X Kong Can C Liu Teng T Liu Aijing A Gao Peng P Song Jiangwei J Zhou Lei L Guo Xin X Han Jun J Yang Hanchun H

Viruses 20190926 10

The papain-like cysteine protease 2 (PLP2) within the N-terminus of the porcine reproductive and respiratory syndrome virus (PRRSV) nsp2 replicase protein specifies a deubiquitinating enzyme (DUB), but its biochemical properties and the role in infection have remained poorly defined. By using in vitro assays, we found that the purified PLP2 could efficiently cleave K63 and K48 linked polyubiquitin chains Ub3-7 in vitro although displaying a differential activity in converting the respective ubiq ...[more]

PMID: 31561412

Similar Datasets

Project description:Porcine reproductive and respiratory syndrome virus (PRRSV) causes a highly contagious disease and brings huge economic losses to commercial pork production worldwide. PRRSV causes severe reproductive failure in sows and respiratory distress in piglets. To trace the evolution of PRRSV in pigs with respiratory diseases in some regions of China, 112 samples were collected from nine provinces in China during 2016-2018. All samples were detected by RT-PCR and analyzed by the Nsp2/ORF5 (ORF5a)-genes-phylogeny. Sequence analysis and recombination analysis were conducted on the Nsp2/ORF5 (ORF5a) genes of the identified strain in the study. The RT-PCR result shown that the positive rate of PRRSV was 50.89% (57/112). Phylogenetic analysis showed that the identified PRRSV strains were all NA genotype and belonged to lineage 1, 3, and 8. The Nsp2 gene of identified PRRSV strains exhibited nucleotide homologies of 53.0 ~ 99.8%, and amino acid homologies of 46.8 ~ 99.7%. The ORF5 gene of identified PRRSV strains exhibited nucleotide homologies of 82.4 ~ 100%, and amino acid homologies of 79.6 ~ 100%. Sequence analysis revealed that a discontinuous 30-amino-acid deletion (positions 481 and 533-561) and a 131-amino-acid discontinuity deletion (positions 323-433, 481, and 533-551) in Nsp2 of PPRSV isolates; all identified strains in this study may be wild strains, and most identified strains may be highly virulent strains. Sequence analysis of ORF5 and ORF5a revealed that the mutation sites of GP5 were mainly concentrated in the signal peptide and epitopes region, while the mutation sites of ORF5a were mainly concentrated in the transmembrane and the intramembrane region. The recombination analysis indicated that there may be multiple recombination regions in identified strains, and the recombination pattern was more complex. This study showed that the prevalent PRRSV strain in some regions of China was still HP-PRRSV, while NADC30 strain also occupied a certain proportion; different types of PRRSV strains showed different patterns and variation in China. This study suggested that the monitoring of PRRSV prevalence and genetic variation should be further strengthened.

Dataset Information

Characterizing the PRRSV nsp2 Deubiquitinase Reveals Dispensability of Cis-Activity for Replication and a Link of nsp2 to Inflammation Induction.

Publications

Characterizing the PRRSV nsp2 Deubiquitinase Reveals Dispensability of <i>Cis</i>-Activity for Replication and a Link of nsp2 to Inflammation Induction.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets