Complementation of Yeast Genes with Human Genes as an Experimental Platform for Functional Testing of Human Genetic Variants

Hamza, Akil; Tammpere, Erik; Kofoed, Megan; Keong, Christelle; Chiang, Jennifer; Giaever, Guri; Nislow, Corey; Hieter, Philip

doi:10.1534/genetics.115.181099

Cited by 72 publications

(75 citation statements)

References 53 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Among ~4,000 disease genes, examination of four systematic screens and curated literature suggests that ~5% of human disease genes currently have a yeast complementation assay (Hamza et al , 2015; Kachroo et al , 2015; Sun et al , 2016). This number could grow dramatically via systematic complementation testing under different environments and genetic backgrounds.…”

Section: Resultsmentioning

confidence: 99%

A framework for exhaustively mapping functional missense variants

Weile

Sun

Coté

et al. 2017

Molecular Systems Biology

172

286

View full text Add to dashboard Cite

Although we now routinely sequence human genomes, we can confidently identify only a fraction of the sequence variants that have a functional impact. Here, we developed a deep mutational scanning framework that produces exhaustive maps for human missense variants by combining random codon mutagenesis and multiplexed functional variation assays with computational imputation and refinement. We applied this framework to four proteins corresponding to six human genes: UBE2I (encoding SUMO E2 conjugase), SUMO1 (small ubiquitin‐like modifier), TPK1 (thiamin pyrophosphokinase), and CALM1/2/3 (three genes encoding the protein calmodulin). The resulting maps recapitulate known protein features and confidently identify pathogenic variation. Assays potentially amenable to deep mutational scanning are already available for 57% of human disease genes, suggesting that DMS could ultimately map functional variation for all human disease genes.

show abstract

Section: Resultsmentioning

confidence: 99%

A framework for exhaustively mapping functional missense variants

Weile

Sun

Coté

et al. 2017

Molecular Systems Biology

172

286

View full text Add to dashboard Cite

show abstract

“…For quantitative analysis, area under the curves were calculated for each condition and normalized to the fitness of the wild-type strain containing a vector-alone control as described in ref. 105.…”

Section: Methodsmentioning

confidence: 99%

Overexpression screens identify conserved dosage chromosome instability genes in yeast and human cancer

Duffy

Fam

Wang

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

View full text Add to dashboard Cite

Somatic copy number amplification and gene overexpression are common features of many cancers. To determine the role of gene overexpression on chromosome instability (CIN), we performed genome-wide screens in the budding yeast for yeast genes that cause CIN when overexpressed, a phenotype we refer to as dosage CIN (dCIN), and identified 245 dCIN genes. This catalog of genes reveals human orthologs known to be recurrently overexpressed and/or amplified in tumors. We show that two genes, TDP1, a tyrosyl-DNA-phosphdiesterase, and TAF12, an RNA polymerase II TATA-box binding factor, cause CIN when overexpressed in human cells. Rhabdomyosarcoma lines with elevated human Tdp1 levels also exhibit CIN that can be partially rescued by siRNA-mediated knockdown of TDP1. Overexpression of dCIN genes represents a genetic vulnerability that could be leveraged for selective killing of cancer cells through targeting of an unlinked synthetic dosage lethal (SDL) partner. Using SDL screens in yeast, we identified a set of genes that when deleted specifically kill cells with high levels of Tdp1. One gene was the histone deacetylase RPD3, for which there are known inhibitors. Both HT1080 cells overexpressing hTDP1 and rhabdomyosarcoma cells with elevated levels of hTdp1 were more sensitive to histone deacetylase inhibitors valproic acid (VPA) and trichostatin A (TSA), recapitulating the SDL interaction in human cells and suggesting VPA and TSA as potential therapeutic agents for tumors with elevated levels of hTdp1. The catalog of dCIN genes presented here provides a candidate list to identify genes that cause CIN when overexpressed in cancer, which can then be leveraged through SDL to selectively target tumors.dosage chromosome instability | overexpression | synthetic dosage lethality | TDP1 | rhabdomyosarcoma

show abstract

“…OD 600 measurements were measured by a Tecan M200 plate reader at 30 min intervals for 72 hr at 30°. Area-under-the-curve analysis was performed as previously described (Hamza et al 2015). …”

Section: Methodsmentioning

confidence: 99%

Dosage Mutator Genes in Saccharomyces cerevisiae: A Novel Mutator Mode-of-Action of the Mph1 DNA Helicase

et al. 2016

Self Cite

View full text Add to dashboard Cite

Mutations that cause genome instability are considered important predisposing events that contribute to initiation and progression of cancer. Genome instability arises either due to defects in genes that cause an increased mutation rate (mutator phenotype), or defects in genes that cause chromosome instability (CIN). To extend the catalog of genome instability genes, we systematically explored the effects of gene overexpression on mutation rate, using a forward-mutation screen in budding yeast. We screened ∼5100 plasmids, each overexpressing a unique single gene, and characterized the five strongest mutators, MPH1 (mutator phenotype 1), RRM3, UBP12, PIF1, and DNA2. We show that, for MPH1, the yeast homolog of Fanconi Anemia complementation group M (FANCM), the overexpression mutator phenotype is distinct from that of mph1Δ. Moreover, while four of our top hits encode DNA helicases, the overexpression of 48 other DNA helicases did not cause a mutator phenotype, suggesting this is not a general property of helicases. For Mph1 overexpression, helicase activity was not required for the mutator phenotype; in contrast Mph1 DEAH-box function was required for hypermutation. Mutagenesis by MPH1 overexpression was independent of translesion synthesis (TLS), but was suppressed by overexpression of RAD27, a conserved flap endonuclease. We propose that binding of DNA flap structures by excess Mph1 may block Rad27 action, creating a mutator phenotype that phenocopies rad27Δ. We believe this represents a novel mutator mode-of-action and opens up new prospects to understand how upregulation of DNA repair proteins may contribute to mutagenesis.

show abstract

Complementation of Yeast Genes with Human Genes as an Experimental Platform for Functional Testing of Human Genetic Variants

Cited by 72 publications

References 53 publications

A framework for exhaustively mapping functional missense variants

A framework for exhaustively mapping functional missense variants

Overexpression screens identify conserved dosage chromosome instability genes in yeast and human cancer

Dosage Mutator Genes in Saccharomyces cerevisiae: A Novel Mutator Mode-of-Action of the Mph1 DNA Helicase

Contact Info

Product

Resources

About