Integrating high-throughput genetic interaction mapping and high-content screening to explore yeast spindle morphogenesis.
We describe the application of a novel screening approach that combines automated yeast genetics, synthetic genetic array (SGA) analysis, and a high-content screening (HCS) system to examine mitotic spindle morphogenesis. We measured numerous spindle and cellular morphological parameters in thousands of single mutants and corresponding sensitized double mutants lacking genes known to be involved in spindle function. We focused on a subset of genes that appear to define a highly conserved mitotic spindle disassembly pathway, which is known to involve Ipl1p, the yeast aurora B kinase, as well as the cell cycle regulatory networks mitotic exit network (MEN) and fourteen early anaphase release (FEAR). We also dissected the function of the kinetochore protein Mcm21p, showing that sumoylation of Mcm21p regulates the enrichment of Ipl1p and other chromosomal passenger proteins to the spindle midzone to mediate spindle disassembly. Although we focused on spindle disassembly in a proof-of-principle study, our integrated HCS-SGA method can be applied to virtually any pathway, making it a powerful means for identifying specific cellular functions.
Vizeacoumar
Franco J
FJ
Banting and Best Department of Medical Research, University of Toronto, Toronto, Ontario M5S 3E1, Canada.
van Dyk
Nydia
N
S Vizeacoumar
Frederick
F
Cheung
Vincent
V
Li
Jingjing
J
Sydorskyy
Yaroslav
Y
Case
Nicolle
N
Li
Zhijian
Z
Datti
Alessandro
A
Nislow
Corey
C
Raught
Brian
B
Zhang
Zhaolei
Z
Frey
Brendan
B
Bloom
Kerry
K
Boone
Charles
C
Andrews
Brenda J
BJ
D057205:Automation, Laboratory; D005821:Genetic Techniques; D009154:Mutation; D011485:Protein Binding; D025941:Protein Interaction Mapping; D025842:SUMO-1 Protein / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry* / Q000166:cytology* / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D008941:Spindle Apparatus / Q000737:chemistry* / Q000235:genetics
D025842:SUMO-1 Protein; D029701:Saccharomyces cerevisiae Proteins