Plasmids recovered from the high copy suppressor screen
| Chromosome (coordinates) . | Number of isolates . | Hygromycin B suppression . | Open-reading frames in insert* . | Proposed function of protein encoded by the suppressing gene . |
|---|---|---|---|---|
| (empty vector) | - | - | - | - |
| II (567382-574763) | 7 | ++++ | ARL1, UBS1, TYR1, POP7, YBR168w | G protein of the Arf-like family |
| X (103703-109978) | 1 | ++++ | ERG20, QCR8, HAL5 | Ser/Thr kinase regulating K+ transporters, Trk1 and Trk2 |
| VI (228886-239723) | 1 | +++ | YFR038w, YFR039c, SAP155, ERJ5 | Regulator of Sit4p phosphatase |
| I (84115-90958) | 1 | ++ | FUN21, SNC1 | Function unknown now |
| VII (430112-435243) | 9+1§ | ++ | MIG1, YGL034c** | Transcriptional repressor of the ENA1 gene, encoding the Na+/Li+ efflux pump |
| X (67860-73441) | 1¶ | ++ | ELO1, YJL195c**, CDC6, YJL193w | Shows 31% identity (50% similarity) to Sly41p, a triose phosphate transporter |
| IV (829075-835598) | 1† | ++ | ATC1, YDR185c, YDR186c | High copy suppressor of the Li+-sensitive phenotype of calcineurin mutants |
| XII overlapping region‡ | 2 | ++ | YLR132C, CKI1 | Choline kinase, catalyzes the committed step in phosphatidylcholine biosynthesis via the Kennedy pathway |
| Chromosome (coordinates) . | Number of isolates . | Hygromycin B suppression . | Open-reading frames in insert* . | Proposed function of protein encoded by the suppressing gene . |
|---|---|---|---|---|
| (empty vector) | - | - | - | - |
| II (567382-574763) | 7 | ++++ | ARL1, UBS1, TYR1, POP7, YBR168w | G protein of the Arf-like family |
| X (103703-109978) | 1 | ++++ | ERG20, QCR8, HAL5 | Ser/Thr kinase regulating K+ transporters, Trk1 and Trk2 |
| VI (228886-239723) | 1 | +++ | YFR038w, YFR039c, SAP155, ERJ5 | Regulator of Sit4p phosphatase |
| I (84115-90958) | 1 | ++ | FUN21, SNC1 | Function unknown now |
| VII (430112-435243) | 9+1§ | ++ | MIG1, YGL034c** | Transcriptional repressor of the ENA1 gene, encoding the Na+/Li+ efflux pump |
| X (67860-73441) | 1¶ | ++ | ELO1, YJL195c**, CDC6, YJL193w | Shows 31% identity (50% similarity) to Sly41p, a triose phosphate transporter |
| IV (829075-835598) | 1† | ++ | ATC1, YDR185c, YDR186c | High copy suppressor of the Li+-sensitive phenotype of calcineurin mutants |
| XII overlapping region‡ | 2 | ++ | YLR132C, CKI1 | Choline kinase, catalyzes the committed step in phosphatidylcholine biosynthesis via the Kennedy pathway |
MA03 was transformed with the high copy number plasmid library as described in Materials and Methods. After confirming that the suppressing function was contained on the plasmid, the plasmids were analyzed by restriction digestion with EcoRI, HindIII and BglII, and sequenced. The nucleotide coordinates of each distinct plasmid obtained from the screen is shown, as well as the level of ability to suppress the hygromycin-B-sensitive phenotype. Individual open-reading frames were subcloned and tested as described in Results and the `bold' gene or open-reading frame was able to suppress alone.
Open reading frames according to the Saccharomyces Genome Database (http://www.yeastgenome.org).
We also obtained a second clone nearly identical to this one, but one from which the region encoding the C-terminal 20% of ATC1 (35 amino acids) was replaced with sequences from the vector encoding 39 amino acids.
Overlapping fragments: pDH43 contained nucleotides 405602-411144 and pDH34 contained nucleotides 406804-411134. Both contained YLR132C and CKI1.
A tenth isolate, pDH39, contained a slightly larger fragment (chromosome VII, nucleotides 429435-435243), but nevertheless included both MIG1 and YGL034c.
We obtained a second clone, pRR193, that overlapped with this one and contained YJL193w and YJL192c (chromosome X, nucleotides 69942-74433).
These have been denoted as `dubious ORFs' according to the Saccharomyces Genome Database.