Functional analysis of the yeast genome

Winzeler, Elizabeth A.; Davis, Ronald W.

doi:10.1016/s0959-437x(97)80039-1

Cited by 55 publications

(19 citation statements)

References 47 publications

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“…tra1 - L3733A was introduced into a group of the knock-out collection of strains (Winzeler and Davis 1997) to examine genetic interactions with SAGA and NuA4 component genes (Table 5). Growth of the double mutants was compared with either single mutant on YPD and YPD containing 3% ethanol.…”

Section: Resultsmentioning

confidence: 99%

Mutational analysis of the C-terminal FATC domain of Saccharomyces cerevisiae Tra1

et al. 2010

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Tra1 is a component of the Saccharomyces cerevisiae SAGA and NuA4 complexes and a member of the PIKK family, which contain a C-terminal phosphatidylinositol 3-kinase-like (PI3K) domain followed by a 35-residue FATC domain. Single residue changes of L3733A and F3744A, within the FATC domain, resulted in transcriptional changes and phenotypes that were similar but not identical to those caused by mutations in the PI3K domain or deletions of other SAGA or NuA4 components. The distinct nature of the FATC mutations was also apparent from the additive effect of tra1-L3733A with SAGA, NuA4, and tra1 PI3K domain mutations. Tra1-L3733A associates with SAGA and NuA4 components and with the Gal4 activation domain, to the same extent as wild-type Tra1; however, steady-state levels of Tra1-L3733A were reduced. We suggest that decreased stability of Tra1-L3733A accounts for the phenotypes since intragenic suppressors of tra1-L3733A restored Tra1 levels, and reducing wild-type Tra1 led to comparable growth defects. Also supporting a key role for the FATC domain in the structure/function of Tra1, addition of a C-terminal glycine residue resulted in decreased association with Spt7 and Esa1, and loss of cellular viability. These findings demonstrate the regulatory potential of mechanisms targeting the FATC domains of PIKK proteins.Electronic supplementary materialThe online version of this article (doi:10.1007/s00294-010-0313-3) contains supplementary material, which is available to authorized users.

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Section: Resultsmentioning

confidence: 99%

Mutational analysis of the C-terminal FATC domain of Saccharomyces cerevisiae Tra1

et al. 2010

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“…All strains used (Supplementary Table S1) were derived from either BY4741, BY4742 or BY4743 (30). The tti2 disruption strains CY6070 and CY6857 have been described (26).…”

Section: Methodsmentioning

confidence: 99%

Genetic selection for mistranslation rescues a defective co-chaperone in yeast

Hoffman

Berg

Shilton

et al. 2016

Nucleic Acids Research

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Despite the general requirement for translation fidelity, mistranslation can be an adaptive response. We selected spontaneous second site mutations that suppress the stress sensitivity caused by a Saccharomyces cerevisiae tti2 allele with a Leu to Pro mutation at residue 187, identifying a single nucleotide mutation at the same position (C70U) in four tRNAProUGG genes. Linkage analysis and suppression by SUF9G3:U70 expressed from a centromeric plasmid confirmed the causative nature of the suppressor mutation. Since the mutation incorporates the G3:U70 identity element for alanyl-tRNA synthetase into tRNAPro, we hypothesized that suppression results from mistranslation of Pro187 in Tti2L187P as Ala. A strain expressing Tti2L187A was not stress sensitive. In vitro, tRNAProUGG (C70U) was mis-aminoacylated with alanine by alanyl–tRNA synthetase, but was not a substrate for prolyl–tRNA synthetase. Mass spectrometry from protein expressed in vivo and a novel GFP reporter for mistranslation confirmed substitution of alanine for proline at a rate of ∼6%. Mistranslating cells expressing SUF9G3:U70 induce a partial heat shock response but grow nearly identically to wild-type. Introducing the same G3:U70 mutation in SUF2 (tRNAProAGG) suppressed a second tti2 allele (tti2L50P). We have thus identified a strategy that allows mistranslation to suppress deleterious missense Pro mutations in Tti2.

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“…Yeast strains, listed in Table 1, are derivatives of BY4741 and BY4742 (Winzeler and Davis 1997). HIS3 -linked tra1 strains, wild-type TRA1 (CY4353), tra1-L3733A (CY4057 and CY4103) and tra1-F3744A (CY4350 and CY4351), are described in Hoke et al (2010).…”

Section: Methodsmentioning

confidence: 99%

Genetic Evidence Links the ASTRA Protein Chaperone Component Tti2 to the SAGA Transcription Factor Tra1

et al. 2012

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Tra1 is a 3744-residue component of the Saccharomyces cerevisiae SAGA, NuA4, and ASTRA complexes. Tra1 contains essential C-terminal PI3K and FATC domains, but unlike other PIKK (phosphoinositide three-kinase–related kinase) family members, lacks kinase activity. To analyze functions of the FATC domain, we selected for suppressors of tra1-F3744A, an allele that results in slow growth under numerous conditions of stress. Two alleles of TTI2, tti2-F328S and tti2-I336F, acted in a partially dominant fashion to suppress the growth-related phenotypes associated with tra1-F3744A as well as its resulting defects in transcription. tti2-F328S suppressed an additional FATC domain mutation (tra1-L3733A), but not a mutation in the PI3K domain or deletions of SAGA or NuA4 components. We find eGFP-tagged Tti2 distributed throughout the cell. Tti2 is a component of the ASTRA complex, and in mammalian cells associates with molecular chaperones in complex with Tti1 and Tel2. Consistent with this finding, Tra1 levels are reduced in a strain with a temperature-sensitive allele of tel2. Further agreeing with a possible role for Tti2 in the folding or stabilization of Tra1, tra1-F3744A was mislocalized to the cytoplasm, particularly under conditions of stress. Since an intragenic mutation of tra1-R3590I also suppressed F3744A, we propose that Tti2 is required for the folding/stability of the C-terminal FATC and PI3K domains of Tra1 into their functionally active form.

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Functional analysis of the yeast genome

Cited by 55 publications

References 47 publications

Mutational analysis of the C-terminal FATC domain of Saccharomyces cerevisiae Tra1

Mutational analysis of the C-terminal FATC domain of Saccharomyces cerevisiae Tra1

Genetic selection for mistranslation rescues a defective co-chaperone in yeast

Genetic Evidence Links the ASTRA Protein Chaperone Component Tti2 to the SAGA Transcription Factor Tra1

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