Vaccinia virus-encoded ribonucleotide reductase subunits are differentially required for replication and pathogenesis.

Ribonucleotide reductases (RRs) are evolutionarily-conserved enzymes that catalyze the rate-limiting step during dNTP synthesis in mammals. RR consists of both large (R1) and small (R2) subunits, which are both required for catalysis by the R1(2)R2(2) heterotetrameric complex. Poxviruses also encode...

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Autores principales:	Don B Gammon, Branawan Gowrishankar, Sophie Duraffour, Graciela Andrei, Chris Upton, David H Evans
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2010
Materias:	Immunologic diseases. Allergy RC581-607 Biology (General) QH301-705.5
Acceso en línea:	https://doaj.org/article/57798b4ea27142bca5e2f085d3137229
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spelling	oai:doaj.org-article:57798b4ea27142bca5e2f085d31372292021-12-02T20:00:30ZVaccinia virus-encoded ribonucleotide reductase subunits are differentially required for replication and pathogenesis.1553-73661553-737410.1371/journal.ppat.1000984https://doaj.org/article/57798b4ea27142bca5e2f085d31372292010-07-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/20628573/?tool=EBIhttps://doaj.org/toc/1553-7366https://doaj.org/toc/1553-7374Ribonucleotide reductases (RRs) are evolutionarily-conserved enzymes that catalyze the rate-limiting step during dNTP synthesis in mammals. RR consists of both large (R1) and small (R2) subunits, which are both required for catalysis by the R1(2)R2(2) heterotetrameric complex. Poxviruses also encode RR proteins, but while the Orthopoxviruses infecting humans [e.g. vaccinia (VACV), variola, cowpox, and monkeypox viruses] encode both R1 and R2 subunits, the vast majority of Chordopoxviruses encode only R2 subunits. Using plaque morphology, growth curve, and mouse model studies, we investigated the requirement of VACV R1 (I4) and R2 (F4) subunits for replication and pathogenesis using a panel of mutant viruses in which one or more viral RR genes had been inactivated. Surprisingly, VACV F4, but not I4, was required for efficient replication in culture and virulence in mice. The growth defects of VACV strains lacking F4 could be complemented by genes encoding other Chordopoxvirus R2 subunits, suggesting conservation of function between poxvirus R2 proteins. Expression of F4 proteins encoding a point mutation predicted to inactivate RR activity but still allow for interaction with R1 subunits, caused a dominant negative phenotype in growth experiments in the presence or absence of I4. Co-immunoprecipitation studies showed that F4 (as well as other Chordopoxvirus R2 subunits) form hybrid complexes with cellular R1 subunits. Mutant F4 proteins that are unable to interact with host R1 subunits failed to rescue the replication defect of strains lacking F4, suggesting that F4-host R1 complex formation is critical for VACV replication. Our results suggest that poxvirus R2 subunits form functional complexes with host R1 subunits to provide sufficient dNTPs for viral replication. Our results also suggest that R2-deficient poxviruses may be selective oncolytic agents and our bioinformatic analyses provide insights into how poxvirus nucleotide metabolism proteins may have influenced the base composition of these pathogens.Don B GammonBranawan GowrishankarSophie DuraffourGraciela AndreiChris UptonDavid H EvansPublic Library of Science (PLoS)articleImmunologic diseases. AllergyRC581-607Biology (General)QH301-705.5ENPLoS Pathogens, Vol 6, Iss 7, p e1000984 (2010)
institution	DOAJ
collection	DOAJ
language	EN
topic	Immunologic diseases. Allergy RC581-607 Biology (General) QH301-705.5
spellingShingle	Immunologic diseases. Allergy RC581-607 Biology (General) QH301-705.5 Don B Gammon Branawan Gowrishankar Sophie Duraffour Graciela Andrei Chris Upton David H Evans Vaccinia virus-encoded ribonucleotide reductase subunits are differentially required for replication and pathogenesis.
description	Ribonucleotide reductases (RRs) are evolutionarily-conserved enzymes that catalyze the rate-limiting step during dNTP synthesis in mammals. RR consists of both large (R1) and small (R2) subunits, which are both required for catalysis by the R1(2)R2(2) heterotetrameric complex. Poxviruses also encode RR proteins, but while the Orthopoxviruses infecting humans [e.g. vaccinia (VACV), variola, cowpox, and monkeypox viruses] encode both R1 and R2 subunits, the vast majority of Chordopoxviruses encode only R2 subunits. Using plaque morphology, growth curve, and mouse model studies, we investigated the requirement of VACV R1 (I4) and R2 (F4) subunits for replication and pathogenesis using a panel of mutant viruses in which one or more viral RR genes had been inactivated. Surprisingly, VACV F4, but not I4, was required for efficient replication in culture and virulence in mice. The growth defects of VACV strains lacking F4 could be complemented by genes encoding other Chordopoxvirus R2 subunits, suggesting conservation of function between poxvirus R2 proteins. Expression of F4 proteins encoding a point mutation predicted to inactivate RR activity but still allow for interaction with R1 subunits, caused a dominant negative phenotype in growth experiments in the presence or absence of I4. Co-immunoprecipitation studies showed that F4 (as well as other Chordopoxvirus R2 subunits) form hybrid complexes with cellular R1 subunits. Mutant F4 proteins that are unable to interact with host R1 subunits failed to rescue the replication defect of strains lacking F4, suggesting that F4-host R1 complex formation is critical for VACV replication. Our results suggest that poxvirus R2 subunits form functional complexes with host R1 subunits to provide sufficient dNTPs for viral replication. Our results also suggest that R2-deficient poxviruses may be selective oncolytic agents and our bioinformatic analyses provide insights into how poxvirus nucleotide metabolism proteins may have influenced the base composition of these pathogens.
format	article
author	Don B Gammon Branawan Gowrishankar Sophie Duraffour Graciela Andrei Chris Upton David H Evans
author_facet	Don B Gammon Branawan Gowrishankar Sophie Duraffour Graciela Andrei Chris Upton David H Evans
author_sort	Don B Gammon
title	Vaccinia virus-encoded ribonucleotide reductase subunits are differentially required for replication and pathogenesis.
title_short	Vaccinia virus-encoded ribonucleotide reductase subunits are differentially required for replication and pathogenesis.
title_full	Vaccinia virus-encoded ribonucleotide reductase subunits are differentially required for replication and pathogenesis.
title_fullStr	Vaccinia virus-encoded ribonucleotide reductase subunits are differentially required for replication and pathogenesis.
title_full_unstemmed	Vaccinia virus-encoded ribonucleotide reductase subunits are differentially required for replication and pathogenesis.
title_sort	vaccinia virus-encoded ribonucleotide reductase subunits are differentially required for replication and pathogenesis.
publisher	Public Library of Science (PLoS)
publishDate	2010
url	https://doaj.org/article/57798b4ea27142bca5e2f085d3137229
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Vaccinia virus-encoded ribonucleotide reductase subunits are differentially required for replication and pathogenesis.

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