Mapping DNA damage‐dependent genetic interactions in yeast via party mating and barcode fusion genetics

• Published in: Molecular systems biology 2018-05, Vol.14 (5), p.e7985-n/a
• Main Authors: Díaz‐Mejía, J Javier, Celaj, Albi, Mellor, Joseph C, Coté, Atina, Balint, Attila, Ho, Brandon, Bansal, Pritpal, Shaeri, Fatemeh, Gebbia, Marinella, Weile, Jochen, Verby, Marta, Karkhanina, Anna, Zhang, YiFan, Wong, Cassandra, Rich, Justin, Prendergast, D'Arcy, Gupta, Gaurav, Öztürk, Sedide, Durocher, Daniel, Brown, Grant W, Roth, Frederick P
• Format: Article
• Language: English
• Subjects: Bar codes; Bleomycin; Chromosome Mapping; condition‐dependent; Damage; Deoxyribonucleic acid; DNA; DNA barcode; DNA Barcoding, Taxonomic; DNA Damage; DNA Repair; DNA sequencing; EMBO17; EMBO22; EMBO26; en masse; Epistasis, Genetic; Gene Deletion; Gene mapping; Genes; Genetic engineering; genetic interaction; Genetic Loci; Genetics; Growth conditions; High-Throughput Nucleotide Sequencing; Kinases; Mapping; Mating; Method; Methods; Methyl Methanesulfonate; Models, Theoretical; Mutants; Promoter Regions, Genetic; Protein kinase; Proteins; Recombination; Repair; Reproducibility of Results; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae Proteins - genetics; sequencing; Yeast
• ISSN: 1744-4292; 1744-4292
• DOI: 10.15252/msb.20177985

Condition‐dependent genetic interactions can reveal functional relationships between genes that are not evident under standard culture conditions. State‐of‐the‐art yeast genetic interaction mapping, which relies on robotic manipulation of arrays of double‐mutant strains, does not scale readily to multi‐condition studies. Here, we describe barcode fusion genetics to map genetic interactions (BFG‐GI), by which double‐mutant strains generated via en masse “party” mating can also be monitored en masse for growth to detect genetic interactions. By using site‐specific recombination to fuse two DNA barcodes, each representing a specific gene deletion, BFG‐GI enables multiplexed quantitative tracking of double mutants via next‐generation sequencing. We applied BFG‐GI to a matrix of DNA repair genes under nine different conditions, including methyl methanesulfonate (MMS), 4‐nitroquinoline 1‐oxide (4NQO), bleomycin, zeocin, and three other DNA‐damaging environments. BFG‐GI recapitulated known genetic interactions and yielded new condition‐dependent genetic interactions. We validated and further explored a subnetwork of condition‐dependent genetic interactions involving MAG1 , SLX4, and genes encoding the Shu complex, and inferred that loss of the Shu complex leads to an increase in the activation of the checkpoint protein kinase Rad53. Synopsis A new method, Barcode Fusion Genetics to Map Genetic Interactions (BFG‐GI) allows generating double mutants and measuring condition‐dependent genetic interactions en masse . Application of BFG‐GI to DNA repair genes reveals a new function for the Shu complex. BFG‐GI involves generating double‐mutant‐specific fused barcodes, enabling to measure the abundance of double mutants en masse by next generation sequencing. Once a double mutant BFG‐GI pool has been generated genetic interactions can be tested in new growth conditions. BFG‐GI is applied to 26 genes related to DNA damage repair in nine different conditions, including seven DNA‐damaging agents. A novel relationship is reported between the Shu complex and the checkpoint protein kinase Rad53. Graphical Abstract A new method, Barcode Fusion Genetics to Map Genetic Interactions (BFG‐GI) allows generating double mutants and measuring condition‐dependent genetic interactions en masse . Application of BFG‐GI to DNA repair genes reveals a new function for the Shu complex.