Replication protein A (RPA) hampers the processive action of APOBEC3G cytosine Deaminase on single-stranded DNA

Artem G. Lada, Irina S R Waisertreiger, Corinn E. Grabow, Aishwarya Prakash, Gloria E Borgstahl, Igor B. Rogozin, Youri I Pavlov

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

Background: Editing deaminases have a pivotal role in cellular physiology. A notable member of this superfamily, APOBEC3G (A3G), restricts retroviruses, and Activation Induced Deaminase (AID) generates antibody diversity by localized deamination of cytosines in DNA. Unconstrained deaminase activity can cause genome-wide mutagenesis and cancer. The mechanisms that protect the genomic DNA from the undesired action of deaminases are unknown. Using the in vitro deamination assays and expression of A3G in yeast, we show that replication protein A (RPA), the eukaryotic single-stranded DNA (ssDNA) binding protein, severely inhibits the deamination activity and processivity of A3G. Principal Findings/Methodology: We found that mutations induced by A3G in the yeast genomic reporter are changes of a single nucleotide. This is unexpected because of the known property of A3G to catalyze multiple deaminations upon one substrate encounter event in vitro. The addition of recombinant RPA to the oligonucleotide deamination assay severely inhibited A3G activity. Additionally, we reveal the inverse correlation between RPA concentration and the number of deaminations induced by A3G in vitro on long ssDNA regions. This resembles the "hit and run" single base substitution events observed in yeast. Significance: Our data suggest that RPA is a plausible antimutator factor limiting the activity and processivity of editing deaminases in the model yeast system. Because of the similar antagonism of yeast RPA and human RPA with A3G in vitro, we propose that RPA plays a role in the protection of the human genome cell from A3G and other deaminases when they are inadvertently diverged from their natural targets. We propose a model where RPA serves as one of the guardians of the genome that protects ssDNA from the destructive processive activity of deaminases by non-specific steric hindrance.

Original languageEnglish (US)
Article numbere24848
JournalPloS one
Volume6
Issue number9
DOIs
StatePublished - Sep 15 2011

Fingerprint

cytosine deaminase
Cytosine Deaminase
Replication Protein A
single-stranded DNA
Single-Stranded DNA
Deamination
deamination
Yeast
Yeasts
yeasts
proteins
Genes
mutagenesis
Assays
Antibody Diversity
genome
Genome
Retroviridae
Recombinant proteins
Mutagenesis

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

Cite this

Replication protein A (RPA) hampers the processive action of APOBEC3G cytosine Deaminase on single-stranded DNA. / Lada, Artem G.; Waisertreiger, Irina S R; Grabow, Corinn E.; Prakash, Aishwarya; Borgstahl, Gloria E; Rogozin, Igor B.; Pavlov, Youri I.

In: PloS one, Vol. 6, No. 9, e24848, 15.09.2011.

Research output: Contribution to journalArticle

Lada, Artem G. ; Waisertreiger, Irina S R ; Grabow, Corinn E. ; Prakash, Aishwarya ; Borgstahl, Gloria E ; Rogozin, Igor B. ; Pavlov, Youri I. / Replication protein A (RPA) hampers the processive action of APOBEC3G cytosine Deaminase on single-stranded DNA. In: PloS one. 2011 ; Vol. 6, No. 9.
@article{0ccb8fe3d73b422cacdcbb674f4b58a5,
title = "Replication protein A (RPA) hampers the processive action of APOBEC3G cytosine Deaminase on single-stranded DNA",
abstract = "Background: Editing deaminases have a pivotal role in cellular physiology. A notable member of this superfamily, APOBEC3G (A3G), restricts retroviruses, and Activation Induced Deaminase (AID) generates antibody diversity by localized deamination of cytosines in DNA. Unconstrained deaminase activity can cause genome-wide mutagenesis and cancer. The mechanisms that protect the genomic DNA from the undesired action of deaminases are unknown. Using the in vitro deamination assays and expression of A3G in yeast, we show that replication protein A (RPA), the eukaryotic single-stranded DNA (ssDNA) binding protein, severely inhibits the deamination activity and processivity of A3G. Principal Findings/Methodology: We found that mutations induced by A3G in the yeast genomic reporter are changes of a single nucleotide. This is unexpected because of the known property of A3G to catalyze multiple deaminations upon one substrate encounter event in vitro. The addition of recombinant RPA to the oligonucleotide deamination assay severely inhibited A3G activity. Additionally, we reveal the inverse correlation between RPA concentration and the number of deaminations induced by A3G in vitro on long ssDNA regions. This resembles the {"}hit and run{"} single base substitution events observed in yeast. Significance: Our data suggest that RPA is a plausible antimutator factor limiting the activity and processivity of editing deaminases in the model yeast system. Because of the similar antagonism of yeast RPA and human RPA with A3G in vitro, we propose that RPA plays a role in the protection of the human genome cell from A3G and other deaminases when they are inadvertently diverged from their natural targets. We propose a model where RPA serves as one of the guardians of the genome that protects ssDNA from the destructive processive activity of deaminases by non-specific steric hindrance.",
author = "Lada, {Artem G.} and Waisertreiger, {Irina S R} and Grabow, {Corinn E.} and Aishwarya Prakash and Borgstahl, {Gloria E} and Rogozin, {Igor B.} and Pavlov, {Youri I}",
year = "2011",
month = "9",
day = "15",
doi = "10.1371/journal.pone.0024848",
language = "English (US)",
volume = "6",
journal = "PLoS One",
issn = "1932-6203",
publisher = "Public Library of Science",
number = "9",

}

TY - JOUR

T1 - Replication protein A (RPA) hampers the processive action of APOBEC3G cytosine Deaminase on single-stranded DNA

AU - Lada, Artem G.

AU - Waisertreiger, Irina S R

AU - Grabow, Corinn E.

AU - Prakash, Aishwarya

AU - Borgstahl, Gloria E

AU - Rogozin, Igor B.

AU - Pavlov, Youri I

PY - 2011/9/15

Y1 - 2011/9/15

N2 - Background: Editing deaminases have a pivotal role in cellular physiology. A notable member of this superfamily, APOBEC3G (A3G), restricts retroviruses, and Activation Induced Deaminase (AID) generates antibody diversity by localized deamination of cytosines in DNA. Unconstrained deaminase activity can cause genome-wide mutagenesis and cancer. The mechanisms that protect the genomic DNA from the undesired action of deaminases are unknown. Using the in vitro deamination assays and expression of A3G in yeast, we show that replication protein A (RPA), the eukaryotic single-stranded DNA (ssDNA) binding protein, severely inhibits the deamination activity and processivity of A3G. Principal Findings/Methodology: We found that mutations induced by A3G in the yeast genomic reporter are changes of a single nucleotide. This is unexpected because of the known property of A3G to catalyze multiple deaminations upon one substrate encounter event in vitro. The addition of recombinant RPA to the oligonucleotide deamination assay severely inhibited A3G activity. Additionally, we reveal the inverse correlation between RPA concentration and the number of deaminations induced by A3G in vitro on long ssDNA regions. This resembles the "hit and run" single base substitution events observed in yeast. Significance: Our data suggest that RPA is a plausible antimutator factor limiting the activity and processivity of editing deaminases in the model yeast system. Because of the similar antagonism of yeast RPA and human RPA with A3G in vitro, we propose that RPA plays a role in the protection of the human genome cell from A3G and other deaminases when they are inadvertently diverged from their natural targets. We propose a model where RPA serves as one of the guardians of the genome that protects ssDNA from the destructive processive activity of deaminases by non-specific steric hindrance.

AB - Background: Editing deaminases have a pivotal role in cellular physiology. A notable member of this superfamily, APOBEC3G (A3G), restricts retroviruses, and Activation Induced Deaminase (AID) generates antibody diversity by localized deamination of cytosines in DNA. Unconstrained deaminase activity can cause genome-wide mutagenesis and cancer. The mechanisms that protect the genomic DNA from the undesired action of deaminases are unknown. Using the in vitro deamination assays and expression of A3G in yeast, we show that replication protein A (RPA), the eukaryotic single-stranded DNA (ssDNA) binding protein, severely inhibits the deamination activity and processivity of A3G. Principal Findings/Methodology: We found that mutations induced by A3G in the yeast genomic reporter are changes of a single nucleotide. This is unexpected because of the known property of A3G to catalyze multiple deaminations upon one substrate encounter event in vitro. The addition of recombinant RPA to the oligonucleotide deamination assay severely inhibited A3G activity. Additionally, we reveal the inverse correlation between RPA concentration and the number of deaminations induced by A3G in vitro on long ssDNA regions. This resembles the "hit and run" single base substitution events observed in yeast. Significance: Our data suggest that RPA is a plausible antimutator factor limiting the activity and processivity of editing deaminases in the model yeast system. Because of the similar antagonism of yeast RPA and human RPA with A3G in vitro, we propose that RPA plays a role in the protection of the human genome cell from A3G and other deaminases when they are inadvertently diverged from their natural targets. We propose a model where RPA serves as one of the guardians of the genome that protects ssDNA from the destructive processive activity of deaminases by non-specific steric hindrance.

UR - http://www.scopus.com/inward/record.url?scp=80052842909&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=80052842909&partnerID=8YFLogxK

U2 - 10.1371/journal.pone.0024848

DO - 10.1371/journal.pone.0024848

M3 - Article

C2 - 21935481

AN - SCOPUS:80052842909

VL - 6

JO - PLoS One

JF - PLoS One

SN - 1932-6203

IS - 9

M1 - e24848

ER -