Systematic yeast synthetic lethal and synthetic dosage lethal screens identify genes required for chromosome segregation

Abstract

Accurate chromosome segregation requires the execution and coordination of many processes during mitosis, including DNA replication, sister chromatid cohesion, and attachment of chromosomes to spindle microtubules via the kinetochore complex. Additional pathways are likely involved because faithful chromosome segregation also requires proteins that are not physically associated with the chromosome. Using kinetochore mutants as a starting point, we have identified genes with roles in chromosome stability by performing genome-wide screens employing synthetic genetic array methodology. Two genetic approaches (a series of synthetic lethal and synthetic dosage lethal screens) isolated 211 nonessential deletion mutants that were unable to tolerate defects in kinetochore function. Although synthetic lethality and synthetic dosage lethality are thought to be based upon similar genetic principles, we found that the majority of interactions associated with these two screens were nonoverlapping. To functionally characterize genes isolated in our screens, a secondary screen was performed to assess defects in chromosome segregation. Genes identified in the secondary screen were enriched for genes with known roles in chromosome segregation. We also uncovered genes with diverse functions, such as RCS1, which encodes an iron transcription factor. RCS1 was one of a small group of genes identified in all three screens, and we used genetic and cell biological assays to confirm that it is required for chromosome stability. Our study shows that systematic genetic screens are a powerful means to discover roles for uncharacterized genes and genes with alternative functions in chromosome maintenance that may not be discovered by using proteomics approaches.chromosome stability ͉ synthetic genetic array ͉ kinetochore

Synthetic genetic array screen identifies PP2A as a therapeutic target in Mad2-overexpressing tumors

“…As Mad2 is overexpressed in many cancer cells (Table S1) (8), this phenotype can be used to specifically kill Mad2-overexpressing tumor cells. We hypothesized that a specific gene whose inhibition causes synthetic dosage lethality (SDL) with Mad2 overexpression can be a target of cancer therapy (13,14).…”

A Genomewide Suppressor and Enhancer Analysis of cdc13-1 Reveals Varied Cellular Processes Influencing Telomere Capping in Saccharomyces cerevisiae

“…We combined the S. cerevisiae gene deletion collection with the well-defined, temperature-sensitive, and reversible cdc13-1 mutation using the SGA strain construction technique and we developed and optimized high-throughput yeast growth assays. Although others have looked systematically for synthetic lethality with temperature-sensitive mutations (e.g., Measday et al 2005;Baetz et al 2006), to the best of our knowledge, a genomewide screen for gene deletions that confer subtle suppressor/enhancer phenotypes on a temperature-sensitive mutation has not previously been reported. Our approach therefore complements other high-throughput genetic techniques such as SGA (Tong et al 2001;Roguev et al 2007) and synthetic dosage lethal analysis (Measday et al 2005).…”

“…Although others have looked systematically for synthetic lethality with temperature-sensitive mutations (e.g., Measday et al 2005;Baetz et al 2006), to the best of our knowledge, a genomewide screen for gene deletions that confer subtle suppressor/enhancer phenotypes on a temperature-sensitive mutation has not previously been reported. Our approach therefore complements other high-throughput genetic techniques such as SGA (Tong et al 2001;Roguev et al 2007) and synthetic dosage lethal analysis (Measday et al 2005). We screened for gene deletions that suppress the lethality of cdc13-1 at the nonpermissive temperature and genes that compromise or contribute to the ''reversibility'' of cdc13-1 mutants.…”

Specific synthetic lethal killing of RAD54B-deficient human colorectal cancer cells by FEN1 silencing

“…Conceptually, SL/SGD is of particular interest in the treatment of cancers, because it would only adversely affect tumor cells harboring the primary sensitizing somatic mutations, and leave the normal tissue unaffected. SL/SGD interactions have been studied extensively in yeast (26,27,36,37) and much data are currently available for yeast genes whose (putative) human orthologs are known to be somatically mutated in human CIN tumors. Although several groups have used rad54 as a gene query (21,(25)(26)(27), rdh54 has never been used as a query, but rather has only been identified as a hit (26).…”