Characterization of nuclease-dependent functions of Exo1p in Saccharomyces cerevisiae

Phuoc T. Tran, Naz Erdeniz, Sandra Dudley, Robert (Mike) Liskay

Research output: Contribution to journalArticle

88 Citations (Scopus)

Abstract

Exo1p is a member of the Rad2p family of structure-specific nucleases that contain conserved N and I nuclease domains. Exo1p has been implicated in numerous DNA metabolic processes, such as recombination, double-strand break repair and DNA mismatch repair (MMR). In this report, we describe in vitro and in vivo characterization of full-length wild-type and mutant forms of Exo1p. Herein, we demonstrate that full-length yeast Exo1p possesses an intrinsic 5′-3′ exonuclease activity as reported previously, but also possesses a flap-endonuclease activity. Our study indicates that Exo1p shares similar, but not identical structure--function relationships to other characterized members of the Rad2p family in the N and I nuclease domains. The two exo1p mutants we examined, showed deficiencies for both double-stranded DNA (dsDNA) 5′-3′ exonuclease and flap-endonuclease activities. Examining the genetic interaction of these two exo1 mutations with rad27Δ suggest that the Exo1p flap-endonuclease activity and not the dsDNA 5′-3′ exonuclease is redundant to Rad27p for viability. In addition, our in vivo results also indicate that many exo1Δ phenotypes are dependent on the complete catalytic activities of Exo1p. Finally, our findings plus those of other investigators suggest that Exo1p functions both in a catalytic and a structural capacity during DNA MMR.

Original languageEnglish (US)
Pages (from-to)895-912
Number of pages18
JournalDNA Repair
Volume1
Issue number11
DOIs
StatePublished - Nov 1 2002

Fingerprint

Flap Endonucleases
Exonucleases
Yeast
Saccharomyces cerevisiae
DNA Mismatch Repair
Repair
Genetic Recombination
Catalyst activity
Yeasts
Research Personnel
Phenotype
Mutation
DNA

Keywords

  • DNA mismatch repair
  • DNA nuclease
  • EXO1
  • MutLα
  • Rad2p family

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

Cite this

Characterization of nuclease-dependent functions of Exo1p in Saccharomyces cerevisiae. / Tran, Phuoc T.; Erdeniz, Naz; Dudley, Sandra; Liskay, Robert (Mike).

In: DNA Repair, Vol. 1, No. 11, 01.11.2002, p. 895-912.

Research output: Contribution to journalArticle

Tran, Phuoc T. ; Erdeniz, Naz ; Dudley, Sandra ; Liskay, Robert (Mike). / Characterization of nuclease-dependent functions of Exo1p in Saccharomyces cerevisiae. In: DNA Repair. 2002 ; Vol. 1, No. 11. pp. 895-912.
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AB - Exo1p is a member of the Rad2p family of structure-specific nucleases that contain conserved N and I nuclease domains. Exo1p has been implicated in numerous DNA metabolic processes, such as recombination, double-strand break repair and DNA mismatch repair (MMR). In this report, we describe in vitro and in vivo characterization of full-length wild-type and mutant forms of Exo1p. Herein, we demonstrate that full-length yeast Exo1p possesses an intrinsic 5′-3′ exonuclease activity as reported previously, but also possesses a flap-endonuclease activity. Our study indicates that Exo1p shares similar, but not identical structure--function relationships to other characterized members of the Rad2p family in the N and I nuclease domains. The two exo1p mutants we examined, showed deficiencies for both double-stranded DNA (dsDNA) 5′-3′ exonuclease and flap-endonuclease activities. Examining the genetic interaction of these two exo1 mutations with rad27Δ suggest that the Exo1p flap-endonuclease activity and not the dsDNA 5′-3′ exonuclease is redundant to Rad27p for viability. In addition, our in vivo results also indicate that many exo1Δ phenotypes are dependent on the complete catalytic activities of Exo1p. Finally, our findings plus those of other investigators suggest that Exo1p functions both in a catalytic and a structural capacity during DNA MMR.

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