Title
C-terminal flap endonuclease (rad27) mutations: lethal interactions with a DNA Ligase I mutation (cdc9-p) and suppression by proliferating cell nuclear antigen (POL30) in Saccharomyces cerevisiae.
Abstract
We recovered the flap endonuclease mutation rad27-K325* in a
synthetic lethal screen with cdc9-p, a DNA ligase mutation with two
substitutions (F43A/F44A) in its proliferating cell nuclear antigen (PCNA)
interaction domain. We created two additional rad27 alleles, rad27-A358* with a
stop codon at residue 358 and rad27-pX8 with substitutions of all eight residues
of the PCNA interaction domain. Tests of mutation avoidance and DNA repair
showed that rad27-K325* confers severe phenotypes similar to rad27Δ, rad27-
A358* confers mild phenotypes and rad27-pX8 confers phenotypes
intermediate between the other two alleles. rad27-K325* behaves similarly to
rad27Δ in being lethal with exo1Δ and rad51Δ and not with rad2Δ. Interestingly,
rad27-pX8 is lethal with rad51Δ, while rad27-A358* is lethal with rad51Δ at an
elevated temperature. High copy expression of POL30 (PCNA) suppresses the
canavanine mutation rate of all the rad27 alleles, including rad27Δ. rad27-K325*
has an absolute lethality with the PCNA mutation pol30-90 that is not
possessed by rad27Δ. These studies show the importance of the C-terminus of
the flap endonuclease in mutation avoidance, and, by virtue of the initial screen,
the role that PCNA plays in coordinating the entry of DNA ligase and the flap
endonuclease in replication and repair.
Description
University of Minnesota Ph.D. dissertation. May 2009 Major: Biochemistry, Molecular Biology, and Biophysics. Advisor: Dennis Livingston. 1 computer file (PDF); vi, 141 pages.
Suggested Citation
Karanja, Kenneth Kimani.
(2009).
C-terminal flap endonuclease (rad27) mutations: lethal interactions with a DNA Ligase I mutation (cdc9-p) and suppression by proliferating cell nuclear antigen (POL30) in Saccharomyces cerevisiae..
Retrieved from the University of Minnesota Digital Conservancy,
http://hdl.handle.net/11299/52143.