Identification of protein complexes required for efficient sister chromatid cohesion.
Ctf8p is a component of Ctf18-RFC, an alternative replication factor C-like complex required for efficient sister chromatid cohesion in Saccharomyces cerevisiae. We performed synthetic genetic array (SGA) analysis with a ctf8 deletion strain as a primary screen to identify other nonessential genes required for efficient sister chromatid cohesion. We then assessed proficiency of cohesion at three chromosomal loci in strains containing deletions of the genes identified in the ctf8 SGA screen. Deletion of seven genes (CHL1, CSM3, BIM1, KAR3, TOF1, CTF4, and VIK1) resulted in defective sister chromatid cohesion. Mass spectrometric analysis of immunoprecipitated complexes identified a physical association between Kar3p and Vik1p and an interaction between Csm3p and Tof1p that we confirmed by coimmunoprecipitation from cell extracts. These data indicate that synthetic genetic array analysis coupled with specific secondary screens can effectively identify protein complexes functionally related to a reference gene. Furthermore, we find that genes involved in mitotic spindle integrity and positioning have a previously unrecognized role in sister chromatid cohesion.
Mayer
Melanie L
ML
Centre for Molecular Medicine and Therapeutics, University of British Columbia, Vancouver, British Columbia, Canada V5Z 4H4.
Pot
Isabelle
I
Chang
Michael
M
Xu
Hong
H
Aneliunas
Victoria
V
Kwok
Teresa
T
Newitt
Rick
R
Aebersold
Ruedi
R
Boone
Charles
C
Brown
Grant W
GW
Hieter
Philip
P
D002842:Chromatids / Q000648:ultrastructure*; D015825:Chromosomes, Fungal; D000939:Epitopes / Q000737:chemistry; D005656:Fungal Proteins / Q000502:physiology; D017353:Gene Deletion; D005821:Genetic Techniques; D016681:Genome, Fungal; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D013058:Mass Spectrometry; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D011233:Precipitin Tests; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins / Q000502:physiology*
D000939:Epitopes; D005656:Fungal Proteins; D008164:Luminescent Proteins; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins