Genetics. Published Articles Ahead of Print: March 31, 2005, Copyright © 2005
doi:10.1534/genetics.104.040295


A more recent version of this article appeared on June 1, 2005.

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Sensitivity to Phosphonoacetic Acid: a New Phenotoype to Probe DNA Polymerase Delta in Saccharomyces cerevisiae

Lei Li 1, Kelly M Murphy 1, Uliana Kanevets 1 and Linda J. Reha-Krantz 1*

1 University of Alberta

* To whom correspondence should be addressed. E-mail: lreha{at}gpu.srv.ualberta.ca.

Submitted on December 22, 2004
Revised on February 9, 2005
Accepted on 15 February 2005


Abstract

A mutant allele (pol3-L612M) of the DNA polymerase delta gene in Saccharomyces cerevisiae was constructed that confers sensitivity to the antiviral drug, phosphonoacetic acid (PAA). We find that PAA-sensitivity tagging DNA polymerases is a useful method to selectively and reversibly inhibit one type of DNA polymerase. Our initial studies reveal that replication by the L612M-DNA pol delta requires Rad27 flap endonuclease activity since the pol3-L612M strain in not viable in the absence of RAD27 function. The L612M-DNA pol delta also strongly depends on mismatch repair (MMR). Reduced viability is observed in the absence of any of the core MMR proteins - Msh2, Mlh1 or Pms1 and severe sensitivity to PAA is observed in the absence of the core proteins, Msh6 or Exo1, but not Msh3. We propose that pol3-L612M cells need the Rad27 flap endonuclease and MMR complexes composed of Msh2/Msh6, Mlh1/Pms1 and Exo 1 for correct processing of Okazaki fragments.

Key Words: Rad27 flap endonuclease, chemically-sensitive DNA pol delta, lagging strand replication, mismatch repair, phosphonoacetic acid




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