title: Silent mutations at codons 65 and 66 in reverse transcriptase alleviate indel formation and restore fitness in subtype B HIV-1 containing D67N and K70R drug resistance mutations.
abstract: Resistance to combined antiretroviral therapy (cART) in HIV-1-infected individuals is typically due to nonsynonymous mutations that change the protein sequence; however, the selection of synonymous or 'silent' mutations in the HIV-1 genome with cART has been reported. These silent K65K and K66K mutations in the HIV-1 reverse transcriptase (RT) occur in over 35% of drug-experienced individuals and are highly associated with the thymidine analog mutations D67N and K70R, which confer decreased susceptibility to most nucleoside and nucleotide RT inhibitors. However, the basis for selection of these silent mutations under selective drug pressure is unknown. Using Illumina next-generation sequencing, we demonstrate that the D67N/K70R substitutions in HIV-1 RT increase indel frequency by 100-fold at RT codons 65-67, consequently impairing viral fitness. Introduction of either K65K or K66K into HIV-1 containing D67N/K70R reversed the error-prone DNA synthesis at codons 65-67 in RT and improved viral replication fitness, but did not impact RT inhibitor drug susceptibility. These data provide new mechanistic insights into the role of silent mutations selected during antiretroviral therapy and have broader implications for the relevance of silent mutations in the evolution and fitness of RNA viruses.
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria 3004, Australia Department of Microbiology, Monash University, Clayton, Victoria 3800, Australia.
authors.forename: Sushama
authors.initials: S
authors.lastname: Telwatte
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria 3004, Australia Department of Infectious Diseases, Monash University, Melbourne, Victoria 3004, Australia.
authors.forename: Anna C
authors.initials: AC
authors.lastname: Hearps
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria 3004, Australia.
authors.forename: Adam
authors.initials: A
authors.lastname: Johnson
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria 3004, Australia.
authors.forename: Catherine F
authors.initials: CF
authors.lastname: Latham
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria 3004, Australia.
authors.forename: Katie
authors.initials: K
authors.lastname: Moore
authors.affiliation: Centre for Population Health, Burnet Institute, Melbourne, Victoria 3004, Australia.
authors.forename: Paul
authors.initials: P
authors.lastname: Agius
authors.affiliation: CSIRO Biosecurity Flagship, Australian Animal Health Laboratory, Geelong, Victoria 3220, Australia.
authors.forename: Mary
authors.initials: M
authors.lastname: Tachedjian
authors.affiliation: Department of Microbiology and Immunology, University of Melbourne, Parkville, Victoria 3010, Australia.
authors.forename: Secondo
authors.initials: S
authors.lastname: Sonza
authors.affiliation: Department of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Sluis-Cremer
authors.affiliation: British Columbia Centre for Excellence in HIV/AIDS, Vancouver, BC V6Z1Y6, Canada.
authors.forename: P Richard
authors.initials: PR
authors.lastname: Harrigan
authors.affiliation: Centre for Biomedical Research, Burnet Institute, Melbourne, Victoria 3004, Australia Department of Microbiology, Monash University, Clayton, Victoria 3800, Australia Department of Infectious Diseases, Monash University, Melbourne, Victoria 3004, Australia Department of Microbiology and Immunology, University of Melbourne, Parkville, Victoria 3010, Australia gildat@burnet.edu.au.
authors.forename: Gilda
authors.initials: G
authors.lastname: Tachedjian
mesh_terms: D019380:Anti-HIV Agents / Q000494:pharmacology; D001483:Base Sequence; D002460:Cell Line; D003062:Codon; D024882:Drug Resistance, Viral / Q000235:genetics; D057809:HEK293 Cells; D015658:HIV Infections / Q000188:drug therapy / Q000821:virology; D054303:HIV Reverse Transcriptase / Q000037:antagonists & inhibitors / Q000235:genetics*; D015497:HIV-1 / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics*; D006801:Humans; D054643:INDEL Mutation; D009154:Mutation; D012367:RNA, Viral / Q000235:genetics; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology
chemical_list: D019380:Anti-HIV Agents; D003062:Codon; D012367:RNA, Viral; D018894:Reverse Transcriptase Inhibitors; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase