Exploring Quantitative Yeast Phenomics with Single-Cell Analysis of DNA Damage Foci.
A significant challenge of functional genomics is to develop methods for genome-scale acquisition and analysis of cell biological data. Here, we present an integrated method that combines genome-wide genetic perturbation of Saccharomyces cerevisiae with high-content screening to facilitate the genetic description of sub-cellular structures and compartment morphology. As proof of principle, we used a Rad52-GFP marker to examine DNA damage foci in ∼20 million single cells from ∼5,000 different mutant backgrounds in the context of selected genetic or chemical perturbations. Phenotypes were classified using a machine learning-based automated image analysis pipeline. 345 mutants were identified that had elevated numbers of DNA damage foci, almost half of which were identified only in sensitized backgrounds. Subsequent analysis of Vid22, a protein implicated in the DNA damage response, revealed that it acts together with the Sgs1 helicase at sites of DNA damage and preferentially binds G-quadruplex regions of the genome. This approach is extensible to numerous other cell biological markers and experimental systems.
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada.
Erin B
EB
Styles
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada.
Karen J
KJ
Founk
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada; Department of Computer Sciences, University of Toronto, Toronto, ON M5S 3E1, Canada.
Lee A
LA
Zamparo
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada; Department of Biochemistry, University of Toronto, Toronto, ON M5S 3E1, Canada.
Tina L
TL
Sing
Department of Molecular Biology, NCCR Program "Frontiers in Genetics", Institute of Genetics, Genomics, Geneva (iGE3), University of Geneva, 30, quai Ernest-Ansermet, 1211 Geneva 4, Switzerland.
Dogus
D
Altintas
Department of Molecular Biology, NCCR Program "Frontiers in Genetics", Institute of Genetics, Genomics, Geneva (iGE3), University of Geneva, 30, quai Ernest-Ansermet, 1211 Geneva 4, Switzerland.
Cyril
C
Ribeyre
Department of Molecular Biology, NCCR Program "Frontiers in Genetics", Institute of Genetics, Genomics, Geneva (iGE3), University of Geneva, 30, quai Ernest-Ansermet, 1211 Geneva 4, Switzerland.
Virginie
V
Ribaud
Laboratory of Computational Systems Biology, Ecole Polytéchnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland.
Jacques
J
Rougemont
Department of Genetics and Center for Genome Sciences and Systems Biology, Washington University School of Medicine in St. Louis, St. Louis, MO 63108, USA.
David
D
Mayhew
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada.
Michael
M
Costanzo
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada.
Matej
M
Usaj
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada.
Adrian J
AJ
Verster
Department of Computer Science and Engineering, University of Minnesota, Minneapolis, MN 55455, USA.
Elizabeth N
EN
Koch
Dipartimento di Bioscienze, Universita' degli Studi di Milano, 20122 Milano, Italy.
Daniele
D
Novarina
Institute of Molecular Biology (IMB), Ackermannweg 4, Mainz 55128, Germany.
Marco
M
Graf
Institute of Molecular Biology (IMB), Ackermannweg 4, Mainz 55128, Germany.
Brian
B
Luke
Dipartimento di Bioscienze, Universita' degli Studi di Milano, 20122 Milano, Italy.
Marco
M
Muzi-Falconi
Department of Computer Science and Engineering, University of Minnesota, Minneapolis, MN 55455, USA.
Chad L
CL
Myers
Department of Genetics and Center for Genome Sciences and Systems Biology, Washington University School of Medicine in St. Louis, St. Louis, MO 63108, USA.
Robi David
RD
Mitra
Department of Molecular Biology, NCCR Program "Frontiers in Genetics", Institute of Genetics, Genomics, Geneva (iGE3), University of Geneva, 30, quai Ernest-Ansermet, 1211 Geneva 4, Switzerland.
David
D
Shore
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada; Department of Biochemistry, University of Toronto, Toronto, ON M5S 3E1, Canada.
Grant W
GW
Brown
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada.
Zhaolei
Z
Zhang
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada. Electronic address: charlie.boone@utoronto.ca.
Charles
C
Boone
The Donnelly Centre, University of Toronto, Toronto, ON M5S 3E1, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 3E1, Canada. Electronic address: brenda.andrews@utoronto.ca.
Brenda J
BJ
Andrews
D004249:DNA Damage; D004260:DNA Repair; D008565:Membrane Proteins; D051721:Rad52 DNA Repair and Recombination Protein; D053484:RecQ Helicases; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins
D008565:Membrane Proteins; D051721:Rad52 DNA Repair and Recombination Protein; D029701:Saccharomyces cerevisiae Proteins; C469825:VID22 protein, S cerevisiae; C090472:SGS1 protein, S cerevisiae; D053484:RecQ Helicases