The examples of chemical genetics studies using
Abstract
The budding yeast Saccharomyces cerevisiae is a useful eukaryote model organism for application to chemical biology studies, for example, drug screening, drug evaluation, and target identification. To use yeast for chemical biology research, however, it has been necessary to construct yeast strains suitable for various compounds because of their high drug resistance. Hence, the deletion of all multidrug resistance genes except for those that are important for viability and for genetic experiments/manipulation could increase the drug sensitivity without influencing the transformation, mating, or sporulation efficiency. There are two major factors conferring multidrug resistance in S. cerevisiae: one is the drug efflux system and the other is the permeability barrier. We therefore constructed a strain which shows high sensitivity to multiple drugs by disrupting the drug efflux system using ATP-binding cassette transporters and suppressing the membrane barrier system by introducing an ERG6-inducible system. In this review, we discuss the construction of our multidrug-sensitive yeast strains and their application in chemical biology.
Keywords
- multidrug-sensitive yeast
- drug efflux system
- permeability barrier system
- drug target identification
- drug screening
1. Introduction
1.1. Screening and target identification of bioactive small molecules: important processes in chemical genetics
The screening of bioactive small molecule compounds is the most important process in drug development. Natural products which have structural diversity isolated from microorganisms, plants, and animals are useful sources in the field of drug development [1]. Structurally, new natural products might show novel activities such as antimicrobial, antiviral, and antitumor activities. These natural products also provide useful information for medicinal chemistry, and allow the development of new synthetic compounds as novel medicines. For example, eribulin, a semi-synthetic derivative of halichondrin B, has been approved as an anti-cancer drug [2, 3, 4]. Therefore, the screening and identification of new small molecules open new avenues for drug development. There are two major ways to identify bioactive small molecules: phenotypic screening and target-based screening. Phenotypic screening is based on cytotoxicity [5, 6, 7], cell cycle arrest [8], immune-suppression [9], and morphological changes [10] of drug-treated cells, fungi, and bacteria. Target-based screening is performed based on measurable readouts such as enzymatic activity inhibition [11] or drug-protein interaction [12]. These approaches have identified useful small molecules and medicines.
Target identification (Target ID) of small molecules is also quite important in order to develop safe and useful drugs [13]. Thalidomide, a cautionary example, was used as a sedative a half-century ago before it was found to be teratogenic and to cause multiple birth defects [14]. However, thalidomide is also used in the treatment of Hansen’s disease, myeloma [14], and so on. In addition, immunomodulatory drugs derived from thalidomide have been developed as a new class of anti-cancer drugs and novel medicines for treating ribosomopathies such as 5q-syndrome [15]. Recently, cereblon, a substrate receptor of the CRL4 E3 ubiquitin ligase, has been identified as a primary target of thalidomide teratogenic [16] and anti-cancer [15] activity. These lines of research provide useful information that cereblon may pose a risk of teratogenic activity and simultaneously serve as an attractive molecular target for immunomodulatory drug development. To identify the relevant target molecules and target pathways, indirect and direct approaches have been used [13]. The indirect approaches include phenotypic analysis and large-scale analysis such as proteomic and genome-wide analyses. Some specific changes in cell morphology, cell cycle arrest, and other phenotypes provide us useful information for predicting targets of the drugs. Based on this property, Morphobase, an encyclopedic database of the morphological changes that occur in drug-treated cells, has been constructed and applied to drug target discovery [17]. Large-scale analyses such as proteomics, metabolomics, and transcriptome analysis of drug-treated cells have been performed to predict the target pathways of bioactive small molecules [18]. Genome-wide genetic studies are also frequently used for drug target ID. For example, synthetic lethal/sick genetic interaction analyses [19, 20], genome-wide overexpression screening [21], and haploinsufficiency-chemical sensitive assays [22] have been used to analyze the mode of action of various drugs. On the other hand, direct approaches, such as affinity probe approaches and genetic analyses, are quite useful to identify the direct target molecules of drugs. By using affinity probe approaches, the targets of thalidomide [16] and FK506 [23] have been identified. Genetic analysis is another powerful method of identifying not only drug targets [24, 25, 26, 27, 28, 29] but also the signaling pathway affected by a drug. Genetic studies using model organisms such as yeast have contributed to identification of the target molecules of bioactive compounds.
The identification of new bioactive small molecules and elucidation of their target molecules/signaling pathways are important not only for developing medicines but also for basic science. Such compounds are a useful tool for understanding the fundamental protein functions in cells. Well-known examples are famous immunosuppressants such as FK506, cyclosporine, and rapamycin. These compounds inhibit immunophilin and T-cell activation through different mechanisms [30]. Studies of these compounds have revealed their detailed immunoreaction mechanisms [30]. Mitotic inhibitors are another example. Mitotic spindle formation and chromosome segregation are fast processes that are completed within approximately 1 hour. Therefore, by taking advantage of rapid pharmacological intervention, studies using microtubule inhibitors (αβ-tubulin inhibitors [31, 32, 33] or γ-tubulin inhibitor [12]), mitotic kinesins (Eg5 [34, 35]), and mitotic kinase inhibitors (aurora kinases [36, 37], Cdk1 [38], Plk1 [39, 40], Mps1 [41, 42]) highlighted useful information regarding the temporal regulation of mitotic spindle architecture and faithful chromosome segregation. These findings could in turn contribute to further drug development. Therefore, target ID of newly found useful bioactive compounds is quite an important process in both basic science and medicine development.
1.2. Saccharomyces cerevisiae , a useful model organism for chemical genetics
Compound | Approach | Finding | Ref. |
---|---|---|---|
Benomyl | Pathway analysis | Identification of Mad1, Mad2, Mad3 as mitotic spindle checkpoint proteins by using benomyl sensitive mutants | [31] |
Benomyl | Pathway analysis | Identification of Bub1, Bub2, Bub3 as mitotic spindle checkpoint proteins by using benomyl sensitive mutants | [32] |
Reveromycin A | Target ID | Identification of |
[27] |
Curvularol | Target ID | Identification of |
[28] |
Rapamycin | Target ID | Identification of |
[29] |
Eudistomin C | Target ID | Identification of |
[50] |
Splitomicin | Screening | Identification of splitomicin as a NAD+-dependent histone deacetylase inhibitor | [51] |
Mammalian cell line (HeLa) | Budding yeast (BY4741) | |
---|---|---|
Cycloheximide (μM) | 0.2 | 270 |
Digitonin (μM) | 0.4 | 1.9 |
Fluphenazine (μM) | 13 | 51 |
Latrunculin A (nM) | 0.2 | >240 |
4-Nitroquinoline 1-oxide (μM) | 0.1 | 7.1 |
Rapamycin (nM) | >300 | 7.1 |
Staurosporine (μM) | 0.1 | 15.1 |
Tunicamycin (μM) | 1.8 | >120 |
In this review, we discuss the construction of two multidrug-sensitive yeast strains, 12geneΔHSR [48] and 12geneΔHSR-iERG [49], which are available for genetic analysis. We also discuss the application of these strains in drug screening and target ID [50].
2. Construction and application of multidrug-sensitive yeast strains
2.1. Construction of multidrug-sensitive yeast strains
We constructed a multidrug-sensitive yeast strain by disrupting 12 ABC transporter-related genes and suppressing the
Transformation efficiency (Cfu/μg) | Mating efficiency (%) | Sporulation efficiency (%) | |
---|---|---|---|
BY4741 | 9.6 × 105 ± 2.2 × 105 | 17.7 ± 7.5 | 21.9 ± 6.8 |
55.0 ± 51.3 | 4.8 ± 1.7 | 9.4 ± 4.7 | |
12geneΔ0 | 1.2 × 105 ± 2.0 × 104 | 15.7 ± 5.3 | 5.0 ± 2.9 |
12geneΔ0HSR | N.D. | N.D. | 28.8 ± 4.6 |
12geneΔ0HSR-iERG6 (under glucose condition) | 7.0 ± 8.2 | 6.4 ± 2.2 | 0.0 ± 0.0 |
12geneΔ0HSR-iERG6 (under galactose condition) | 3.0 × 104 ± 2.4 × 104 | N.D. | 10.7 ± 3.0 |
2.2. Application 1: drug screening
2.2.1. Availability of 12geneΔ0HSR-iERG6 in drug screening
In general,
Number of broth | Number of hit broth | Hit ratio (%) | |
---|---|---|---|
Fungus | 2664 | 149 | 5.6 |
Actinomycetes | 5617 | 289 | 5.1 |
Total | 8281 | 438 | 5.3 |
Number of broth | Number of hit broth | Hit ratio (%) | |
---|---|---|---|
Fungus | 3144 | 270 | 8.6 |
Actinomycetes | 3067 | 253 | 8.2 |
Total | 6211 | 523 | 8.4 |
To identify the mitochondrial inhibitors, we used the difference in cell growth between the glucose medium and the glycerol medium. Yeast can use glycerol as a respiratory substance after the conversion to dihydroxyacetone phosphate via glycerol-3-phosphate by cytosolic and mitochondrial enzymes, GUT1p and GUT2p, respectively. Therefore, yeast could grow even in the presence of a mitochondrial inhibitor in glucose medium because of anaerobic respiration, but not in glycerol medium in which one of the metabolites in glycolysis, dihydroxyacetone phosphate, could not be produced. Therefore, we compared the growth inhibition induced by microbial broth samples on glucose medium (1% yeast extract, 2% polypeptone, 2% glucose, 1.5% agar) with that on glycerol medium (1% yeast extract, 2% polypeptone, 3% glycerol, 1.5% agar), and chose the broth which inhibited yeast growth on glycerol medium but not on glucose medium [55]. Growth inhibition activities of microbial broth samples were evaluated using the paper disc method on agar plates inoculated with recombinant
To determine whether it is possible to isolate the novel compounds or not, we selected the microbial broths which were detected using 12geneΔ0HSR-iERG6 but not using the quadruplex mutant. We found a total of 46 broths (fungus origin: 16 broths; actinomycetes origin: 30 broths) which inhibited the growth of 12geneΔ0HSR-iERG6 specifically. Among these broths, we selected two fungus broths for further purification of active metabolites, and isolated 4,6′-anhydrooxysporidinone (
2.2.2. Screening of readthrough compounds
Because the usefulness of our strains was confirmed, we next performed the preliminary screening of compounds that show readthrough activities. Readthrough compounds allow the translational machinery to skip nonsense mutations encoding premature termination codons (PTCs) and could become medicines for hereditary diseases caused by PTCs (Figure 4). To date, many small molecules have been developed as readthrough drug candidates. Several forms of aminoglycoside antibiotics, such as gentamicin (
To discover novel readthrough compounds, we constructed yeast strains for the screening of readthrough compounds using 12geneΔ0HSR.
Next, we initiated a high-throughput screening of the readthrough compounds based on the halo assay using chemical library. This screening is underway, but already several hit compounds have been found, including rapamycin (
2.3. Application 2: target ID
Since our strains show multidrug sensitivity without a decrease in genetic availability, they should also be useful for performing target ID for drugs and the mechanism evaluation of compounds, especially those which are only available in limited amounts, such as natural products. Here we show an example of target ID [50]. Eudistomin C (EudiC, Figure 8), a natural product isolated from the Caribbean tunicate
Collectively, our target ID studies of EudiC suggested the mode of action of EudiC cytotoxicity and indicated that our sensitive strains would be quite useful for performing drug target IDs in a relatively short period.
3. Conclusions and perspective
In the field of chemical biology, several model organisms, including yeast, worms, flies, and mice, have been used. Yeast is one of the most-used model organisms due to its ease of handling and its genetic availability, but its drug resistance is sometimes an obstacle to investigation. To overcome this problem, we constructed two multidrug-sensitive yeast strains, 12geneΔ0HSR and 12geneΔ0HSR-iERG6. These strains not only show a broad spectrum of drug sensitivities against compounds for which resistance is shown by both ABC transporters and ergosterol without influencing transformation, mating, or sporulation efficiency, but they are also useful for drug screening. Indeed, we performed a screening of antifungal compounds and protein translation regulators which skip stop codons and found some promising candidates. Using 12geneΔ0HSR-iERG6, we succeeded in improving the hit rate of drug screening from microbial broth. The screening of microbial broth which inhibits the growth of 12geneΔ0HSR-iERG6 but not of the quadruplex mutant identified novel compounds suggested that our multidrug-sensitive strain-based screening using previously tested chemical sources in yeast screening could identify new bioactive compounds. Furthermore, as our screening system for readthrough compounds, genetically modified multidrug-sensitive strains can be applied for several types of screening such as a yeast 2-hybrid system-based protein-protein interaction modulators screening. Recently, a yeast 3-hybrid system has been applied for drug-protein interaction analysis [78]. In this study, the
Recently, it has been reported that RNAseq combined with Crisper/Cas9-based genome-editing technologies is useful for target ID in mammalian cells [25]. Identification of the drug target using our multidrug-sensitive strains and confirmation of the identified mutation in mammalian cells by Crisper/Cas9-based genome editing will reveal the mechanisms of drugs in more detail. Our multidrug-sensitive strains have the potential to facilitate chemical genetic studies and contribute to the development of medicines in the future.
References
- 1.
Katz L, Baltz RH. Natural product discovery: Past, present, and future. Journal of Industrial Microbiology & Biotechnology. 2016; 43 (2):155-176. DOI: 10.1007/s10295-015-1723-5 - 2.
Towle MJ, Salvato KA, Budrow J, Wels BF, Kuznetsov G, Aalfs KK, et al. In vitro and in vivo anticancer activities of synthetic macrocyclic ketone analogues of halichondrin B. Cancer Research. 2001; 61 (3):1013-1021 - 3.
Nastrucci C, Cesario A, Russo P. Anticancer drug discovery from the marine environment. Recent Patents of Anticancer Drug Discovery. 2012; 7 :218-232 - 4.
Chiba H, Tagami K. Research and development of HALAVEN (Eribulin Mesylate). Journal of Synthetic Organic Chemistry, Japan. 2011; 69 (5):600-610. DOI: 10.5059/yukigoseikyokaishi.69.600 - 5.
Low WK, Dang Y, Schneider-Poetsch T, Shi Z, Choi NS, Merrick WC, et al. Inhibition of eukaryotic translation initiation by the marine natural product pateamine a. Molecular Cell. 2005; 20 (5):709-722. DOI: 10.1016/j.molcel.2005.10.008 - 6.
Feling RH, Buchanan GO, Mincer TJ, Kauffman CA, Jensen PR, Fenical W. Salinosporamide A: A highly cytotoxic proteasome inhibitor from a novel microbial source, a marine bacterium of the new genus Salinospora. Angewandte Chemie International Edition. 2003; 42 (3):355-357. DOI: 10.1002/anie.200390115 - 7.
Ueda H, Nakajima H, Hori Y, Fujita T, Nishimura M, Goto T, et al. FR901228, a novel antitumor bicyclic depsipeptide produced by Chromobacterium violaceum no. 968. I. Taxonomy, fermentation, isolation, physico-chemical and biological properties, and antitumor activity. Journal of Anitibiotics (Tokyo). 1994; 47 (3):301-310 - 8.
Yoshida M, Beppu T. Reversible arrest of proliferation of rat 3Y1 fibroblasts in both the G1 and G2. Experimental Cell Research. 1988; 177 (1):122-131 - 9.
Kino T, Hatanaka H, Hashimoto M, Nishiyama M, Goto T, Okuhara M, et al. FK-506, a novel immunosuppressant isolated from a Streptomyces. I. Fermentation, isolation, and physico-chemical and biological characteristics. Journal of Antibiotics (Tokyo). 1987; 40 (9):1249-1255 - 10.
Ingber D, Fujita T, Kishimoto S, Sudo K, Kanamaru T, Brem H, et al. Synthetic analogues of fumagillin that inhibit angiogenesis and suppress tumour growth. Nature. 1990; 348 (6301):555-557. DOI: 10.1038/348555a0 - 11.
Steegmaier M, Hoffmann M, Baum A, Lénárt P, Petronczki M, Krssák M, et al. BI 2536, a potent and selective inhibitor of polo-like kinase 1, inhibits tumor growth in vivo. Current Biology. 2007; 17 (4):316-322. DOI: 10.1016/j.cub.2006.12.037 - 12.
Chinen T, Liu P, Shioda S, Pagel J, Cerikan B, Lin TC, et al. The γ-tubulin-specific inhibitor gatastatin reveals temporal requirements of microtubule nucleation during the cell cycle. Nature Communications. 2015; 6 :8722. DOI: 10.1038/ncomms9722 - 13.
Schenone M, Dančík V, Wagner BK, Clemons PA. Target identification and mechanism of action in chemical biology and drug discovery. Nature Chemical Biology. 2013; 9 (4):232-240. DOI: 10.1038/nchembio.1199 - 14.
Laffitte E, Revuz J. Thalidomide: An old drug with new clinical applications. Expert Opinion on Drug Safety. 2004; 3 (1):47-56. DOI: 10.1517/14740338.3.1.47 - 15.
Ito T, Handa H. Cereblon and its downstream substrates as molecular targets of immunomodulatory drugs. International Journal of Hematology. 2016; 104 (3):293-299. DOI: 10.1007/s12185-016-2073-4 - 16.
Ito T, Ando H, Suzuki T, Ogura T, Hotta K, Imamura Y, et al. Identification of a primary target of thalidomide teratogenicity. Science. 2010; 327 (5971):1345-1350. DOI: 10.1126/science.1177319 - 17.
Futamura Y, Kawatani M, Kazami S, Tanaka K, Muroi M, Shimizu T, et al. Morphobase, an encyclopedic cell morphology database, and its use for drug target identification. Chemistry & Biology. 2012; 19 (12):1620-1630. DOI: 10.1016/j.chembiol.2012.10.014 - 18.
Muroi M, Kazami S, Noda K, Kondo H, Takayama H, Kawatani M, et al. Application of proteomic profiling based on 2D-DIGE for classification of compounds according to the mechanism of action. Chemistry & Biology. 2010; 17 (5):460-470. DOI: 10.1016/j.chembiol.2010.03.016 - 19.
Tong AH, Evangelista M, Parsons AB, Xu H, Bader GD, Pagé N, et al. Systematic genetic analysis with ordered arrays of yeast deletion mutants. Science. 2001; 294 (5550):2364-2368. DOI: 10.1126/science.1065810 - 20.
Parsons AB, Brost RL, Ding H, Li Z, Zhang C, Sheikh B, et al. Integration of chemical-genetic and genetic interaction data links bioactive compounds to cellular target pathways. Nature Biotechnology. 2004; 22 (1):62-69. DOI: 10.1038/nbt919 - 21.
Nishimura S, Arita Y, Honda M, Iwamoto K, Matsuyama A, Shirai A, et al. Marine antifungal theonellamides target 3β-hydroxysterol to activate Rho1 signaling. Nature Chemical Biology. 2010; 6 (7):519-526. DOI: 10.1038/nchembio.387 - 22.
Giaever G, Shoemaker DD, Jones TW, Liang H, Winzeler EA, Astromoff A, et al. Genomic profiling of drug sensitivities via induced haploinsufficiency. Nature Genetics. 1999; 21 (3):278-283. DOI: 10.1038/6791 - 23.
Harding MW, Galat A, Uehling DE, Schreiber SL. A receptor for the immunosuppressant FK506 is a cis-trans peptidyl-prolyl isomerase. Nature. 1989; 341 (6244):758-760. DOI: 10.1038/341758a0 - 24.
Luo L, Parrish CA, Nevins N, McNulty DE, Chaudhari AM, Carson JD, et al. ATP-competitive inhibitors of the mitotic kinesin KSP that function via an allosteric mechanism. Nature Chemical Biology. 2007; 3 (11):722-726. DOI: 10.1038/nchembio.2007.34 - 25.
Kasap C, Elemento O, Kapoor TM. DrugTargetSeqR: A genomics- and CRISPR-Cas9-based method to analyze drug targets. Nature Chemical Biology. 2014; 10 (8):626-628. DOI: 10.1038/nchembio.1551 - 26.
Wu CY, Feng Y, Cardenas ER, Williams N, Floreancig PE, De Brabander JK, et al. Studies toward the unique pederin family member psymberin: Structure-activity relationships, biochemical studies, and genetics identify the mode-of-action of psymberin. Journal of the American Chemical Society. 2012; 134 (46):18998-19003. DOI: 10.1021/ja 3057002 - 27.
Miyamoto Y, Machida K, Mizunuma M, Emoto Y, Sato N, Miyahara K, et al. Identification of Saccharomyces cerevisiae isoleucyl-tRNA synthetase as a target of the G1-specific inhibitor Reveromycin A. Journal of Biological Chemistry. 2002;277 (32):28810-28814 - 28.
Kobayashi Y, Mizunuma M, Osada H, Obayashi YK, Izunuma MM, Sada HO, et al. Identification of Saccharomyces cerevisiae ribosomal protein L3 as a target of curvularol, a G1-specific inhibitor of mammalian cells. Bioscience Biotechnology and Biochemistry. 2006;70 (10):2451-2459 - 29.
Heitman J, Movva NR, Hall MN. Targets for cell cycle arrest by the immunosuppressant rapamycin in yeast. Science. 1991; 253 (5022):905-909 - 30.
Schreiber SL. The immunophilins their immunosuppressive ligands molecular recognition by the immunophilins. Science. 1991; 251 (4991):283-287 - 31.
Li R, Murray AW. Feedback control of mitosis in budding yeast. Cell. 1991; 66 (3):519-531 - 32.
Hoyt MA, Totis L, Roberts BT. S. cerevisiae genes required for cell cycle arrest in response to loss of microtubule function. Cell. 1991;66 (3):507-517 - 33.
Kelling J, Sullivan K, Wilson L, Jordan MA. Suppression of centromere dynamics by Taxol in living osteosarcoma cells. Cancer Research. 2003; 63 (11):2794-2801. DOI: 10.1242/jcs.024018 - 34.
Groen AC, Needleman D, Brangwynne C, Gradinaru C, Fowler B, Mazitschek R, et al. A novel small-molecule inhibitor reveals a possible role of kinesin-5 in an astral spindle-pole assembly. Journal of Cell Science. 2008; 121 (14):2293-2300. DOI: 10.1242/jcs.024018 - 35.
Kapoor TM, Mayer TU, Coughlin ML, Mitchison TJ. Probing spindle assembly mechanisms with monastrol, a small molecule inhibitor of the mitotic kinesin, Eg5. Journal of Cell Biology. 2000; 150 (5):975-988 - 36.
Kesisova IA, Nakos KC, Tsolou A, Angelis D, Lewis J, Chatzaki A, et al. Tripolin A, a novel small-molecule inhibitor of aurora A kinase, reveals new regulation of HURP’s distribution on microtubules. PLoS One. 2013; 8 (3):e58485. DOI: 10.1371/journal.pone.0058485 - 37.
J-M W, Chen C-T, Coumar MS, Lin W-H, Chen Z-J, Hsu JT, et al. Aurora kinase inhibitors reveal mechanisms of HURP in nucleation of centrosomal and kinetochore microtubules. Proceedings of the National Academy of Sciences of the United States of America. 2013; 110 (19):E1779-E1787. DOI: 10.1073/pnas.1220523110 - 38.
Royou A, McCusker D, Kellogg DR, Sullivan W. Grapes (Chk1) prevents nuclear CDK1 activation by delaying cyclin B nuclear accumulation. Journal of Cell Biology. 2008; 183 (1):63-75. DOI: 10.1083/jcb.200801153 - 39.
Burkard ME, Randall CL, Larochelle S, Zhang C, Shokat KM, Fisher RP, et al. Chemical genetics reveals the requirement for Polo-like kinase 1 activity in positioning RhoA and triggering cytokinesis in human cells. Proceedings of the National Academy of Sciences of the United States of America. 2007; 104 (11):4383-4388. DOI: 10.1073/pnas.0701140104 - 40.
Lénárt P, Petronczki M, Steegmaier M, Di Fiore B, Lipp JJ, Hoffmann M, et al. The small-molecule inhibitor BI 2536 reveals novel insights into mitotic roles of Polo-like kinase 1. Current Biology. 2007; 17 (4):304-315. DOI: 10.1016/j.cub.2006.12.046 - 41.
Hewitt L, Tighe A, Santaguida S, White AM, Jones CD, Musacchio A, et al. Sustained Mps1 activity is required in mitosis to recruit O-Mad2 to the Mad1-C-Mad2 core complex. Journal of Cell Biology. 2010; 190 (1):25-34. DOI: 10.1083/jcb.201002133 - 42.
Santaguida S, Tighe A, D’Alise AM, Taylor SS, Musacchio A. Dissecting the role of MPS1 in chromosome biorientation and the spindle checkpoint through the small molecule inhibitor reversine. Journal of Cell Biology. 2010; 190 (1):73-87. DOI: 10.1083/jcb.201001036 - 43.
Simon J, Bedalov A. Yeast as a model system for anticancer drug discovery. Nature Reviews Cancer. 2004; 4 (6):481-492. DOI: 10.1038/nrc1372 - 44.
Hirao M, Posakony J, Nelson M, Hruby H, Jung M, Simon JA, et al. Identification of selective inhibitors of NAD+-dependent deacetylases using phenotypic screens in yeast. Journal of Biological Chemistry. 2003; 278 (52):52773-52782. DOI: 10.1074/jbc.M308966200 - 45.
Decottignies A, Goffeau A. Complete inventory of the yeast ABC proteins. Nature Genetics. 1997; 15 (2):137-145. DOI: 10.1038/ng0297-137 - 46.
Bauer BE, Wolfger H, Kuchler K. Inventory and function of yeast ABC proteins: About sex, stress, pleiotropic drug and heavy metal resistance. Biochimica et Biophysica Acta. 1999; 1461 (2):217-236 - 47.
Jungwirth H, Kuchler K, Yeast ABC. Transporters—A tale of sex, stress, drugs and aging. FEBS Letters. 2006; 580 :1131-1138. DOI: 10.1016/j.febslet.2005.12.050 - 48.
Chinen T, Ota Y, Nagumo Y, Masumoto H, Usui T. Construction of multidrug-sensitive yeast with high sporulation efficiency. Bioscience Biotechnology and Biochemistry. 2011; 75 (8):1588-1593. DOI: 10.1271/bbb.110311 - 49.
Chinen T, Nagumo Y, Usui T. Construction of a genetic analysis-available multidrug sensitive yeast strain by disruption of the drug efflux system and conditional repression of the membrane barrier system. The Journal of General and Applied Microbiology. 2014; 60 (4):160-162 - 50.
Ota Y, Chinen T, Yoshida K, Kudo S, Nagumo Y, Shiwa Y, et al. Eudistomin C, an antitumor and antiviral natural product, targets 40S ribosome and inhibits protein translation. ChemBioChem. 2016; 17 :1616-1620. DOI: 10.1002/cbic.201600075 - 51.
Bedalov A, Gatbonton T, Irvine WP, Gottschling DE, Simon JA. Identification of a small molecule inhibitor of Sir2p. Proceedings of the National Academy of Sciences of the United States of America. 2001; 98 (26):15113-15118. DOI: 10.1073/pnas.261574398 - 52.
Storici F, Lewis LK, Resnick MA. In vivo site-directed mutagenesis using oligonucleotides. Nature Biotechnology. 2001; 19 (8):773-776. DOI: 10.1038/90837 - 53.
Deutschbauer AM, Davis RW. Quantitative trait loci mapped to single-nucleotide resolution in yeast. Nature Genetics. 2005; 37 (12):1333-1340. DOI: 10.1038/ng1674 - 54.
Dimitrov LN, Brem RB, Kruglyak L, Gottschling DE. Polymorphisms in multiple genes contribute to the spontaneous mitochondrial genome instability of Saccharomyces cerevisiae S288C strains. Genetics. 2009;183 (1):365-383. DOI: 10.1534/genetics.109.104497 - 55.
Watanabe Y, Suga T, Narusawa S, Iwatsuki M, Nonaka K, Nakashima T, et al. Decatamariic acid, a new mitochondrial respiration inhibitor discovered by pesticidal screening using drug-sensitive Saccharomyces cerevisiae . Journal of Antibiotics (Tokyo). 2017;70 (4):395-399. DOI: 10.1038/ja.2016.164 - 56.
Wijeratne EMK, Gunatilaka AAL. Biomimetic conversion of (−)-fusoxypyridone and (−)-oxysporidinone to (−)-sambutoxin: Further evidence for the structure of the tricyclic pyridone alkaloid, (−)-fusoxypyridone. Bioorganic & Medicinal Chemistry Letters. 2011; 21 (8):2327-2329. DOI: 10.1016/j.bmcl.2011.02.091 - 57.
Zhan J, Burns AM, Liu MX, Faeth SH, Gunatilaka AAL. Search for cell motility and angiogenesis inhibitors with potential anticancer activity: Beauvericin and other constituents of two endophytic strains of Fusarium oxysporum . Journal of Natural Products. 2007;70 (2):227-232. DOI: 10.1021/np060394t - 58.
Wang Q-X, Li S-F, Zhao F, Dai H-Q, Bao L, Ding R, et al. Chemical constituents from endophytic fungus Fusarium oxysporum . Fitoterapia. 2011;82 (5):777-781. DOI: 10.1016/j.fitote.2011.04.002 - 59.
Zhang F, Ding G, Li L, Cai X, Si Y, Guo L, et al. Isolation, antimicrobial activity, and absolute configuration of the furylidene tetronic acid core of pestalotic acids A–G. Organic & Biomolecular Chemistry. 2012; 10 (27):5307-5314. DOI: 10.1039/c2ob25469g - 60.
Burke JF, Mogg AE. Suppression of a nonsense mutation in mammalian cells in vivo by the aminoglycoside antibiotics G-418 and paromomycin. Nucleic Acids Research. 1985; 13 (17):6265-6272 - 61.
Barton-Davis ER, Cordier L, Shoturma DI, Leland SE, Sweeney HL. Aminoglycoside antibiotics restore dystrophin function to skeletal muscles of mdx mice. Journal of Clinical Investigation. 1999; 104 (4):375-381. DOI: 10.1172/JCI7866 - 62.
Sabbavarapu NM, Shavit M, Degani Y, Smolkin B, Belakhov V, Baasov T. Design of novel aminoglycoside derivatives with enhanced suppression of diseases-causing nonsense mutations. ACS Medicinal Chemistry Letters. 2016; 7 (4):418-423. DOI: 10.1021/acsmedchemlett.6b00006 - 63.
Mingeot-Leclercq MP, Tulkens PM. Aminoglycosides: Nephrotoxicity. Antimicrobial Agents and Chemotherapy. 1999; 43 (5):1003-1012 - 64.
Hutchin T, Cortopassi G. Proposed molecular and cellular mechanism for aminoglycoside ototoxicity. Antimicrobial Agents and Chemotherapy. 1994; 38 (11):2517-2520 - 65.
Welch EM, Barton ER, Zhuo J, Tomizawa Y, Friesen WJ, Trifillis P, et al. PTC124 targets genetic disorders caused by nonsense mutations. Nature. 2007; 447 (7140):87-91. DOI: 10.1038/nature05756 - 66.
Hamada M, Takeuchi T, Kondo S, Ikeda Y, Naganawa H. A new antibiotic, negamycin. Journal of Antibiotics (Tokyo). 1970; 23 (3):170-171 - 67.
Arakawa M, Shiozuka M, Nakayama Y, Hara T, Hamada M, Kondo S, et al. Negamycin restores dystrophin expression in skeletal and cardiac muscles of mdx mice. Journal of Biochemistry. 2003; 134 (5):751-758 - 68.
Taguchi A, Hamada K, Kotake M, Shiozuka M, Nakaminami H, Pillaiyar T, et al. Discovery of natural products possessing selective eukaryotic readthrough activity: 3-epi-deoxynegamycin and its leucine adduct. ChemMedChem. 2014; 9 (10):2233-2237. DOI: 10.1002/cmdc.201402208 - 69.
Hamada K, Taguchi A, Kotake M, Aita S, Murakami S, Takayama K, et al. Structure-activity relationship studies of 3-epi-deoxynegamycin derivatives as potent readthrough drug candidates. ACS Medicinal Chemistry Letters. 2015; 6 (6):689-694. DOI: 10.1021/acsmedchemlett.5b00121 - 70.
Roman H. Studies of gene mutation in Saccharomyces. Cold Spring Harbor Symposia on Quantitative Biology. 1956; 21 :175-185 - 71.
Hieter P, Mann C, Snyder M, Davis RW. Mitotic stability of yeast chromosomes: A colony color assay that measures nondisjunction and chromosome loss. Cell. 1985; 40 (2):381-392 - 72.
Pal M, Ishigaki Y, Nagy E, Maquat LE. Evidence that phosphorylation of human Upf1 protein varies with intracellular location and is mediated by a wortmannin-sensitive and rapamycin-sensitive PI 3-kinase-related kinase signaling pathway. RNA. 2001; 7 :5-15 - 73.
Nickless A, Jackson E, Marasa J, Nugent P, Mercer RW, Piwnica-Worms D, et al. Intracellular calcium regulates nonsense-mediated mRNA decay. Nature Medicine. 2014; 20 (8):961-968. DOI: 10.1038/nm.3620 - 74.
Rinehart KL, Kobayashi J, Harbour GC, Hughes RG Jr, Mizsak SA, Scahill TA. Eudistomins C, E, K, and L, potent antiviral compounds containing a novel oxathiazepine ring from the Caribbean tunicate Eudistoma olivaceum. Journal of the American Chemical Society. 1984; 106 (5):1524-1526. DOI: 10.1021/ja00317a079 - 75.
Rinehart KL, Kobayashi J, Harbour GC, Gilmore J, Mascal M, Holt TG, et al. Eudistomins A-Q, β-carbolines from the antiviral Caribbean tunicate Eudistoma olivaceum . Journal of the American Chemical Society. 1987;109 (11):3378-3387. DOI: 10.1021/ja00245a031 - 76.
Lake RJ, Blunt JW, Munro MHG. Eudistomins from the New Zealand ascidian Ritterella sigillinoides . Australian Journal of Chemistry. 1989;42 (7):1201-1206. DOI: 10.1071/CH9891201 - 77.
Granneman S, Nandineni MR, Baserga SJ. The putative NTPase Fap7 mediates cytoplasmic 20S pre-rRNA processing through a direct interaction with Rps14. Molecular Cellular Biology. 2005; 25 (23):10352-10364. DOI: 10.1128/MCB.25.23.10352-10364.2005 - 78.
Chidley C, Haruki H, Pedersen MG, Muller E, Johnsson K. A yeast-based screen reveals that sulfasalazine inhibits tetrahydrobiopterin biosynthesis. Nature Chemical Biology. 2011; 7 (6):375-383. DOI: 10.1038/nchembio.557 - 79.
Puri M, Kaur I, Perugini MA, Gupta RC. Ribosome-inactivating proteins: Current status and biomedical applications. Drug Discovery Today. 2012; 17 (13-14):774-783. DOI: 10.1016/j.drudis.2012.03.007 - 80.
Ivashkiv LB, Donlin LT. Regulation of type I interferon responses. Nature Reviews Immunology. 2014; 14 (1):36-49. DOI: 10.1038/nri3581 - 81.
Diamond MS, Farzan M. The broad-spectrum antiviral functions of IFIT and IFITM proteins. Nature Reviews Immunology. 2012; 13 (1):46-57. DOI: 10.1038/nri3344 - 82.
Ben-Shem A, Garreau de Loubresse N, Melnikov S, Jenner L, Yusupova GYM. The structure of the eukaryotic ribosome at 3.0 Å resolution. Science. 2011; 334 (6062):1524-1529. DOI: 10.1126/science.1212642 - 83.
Babaylova E, Graifer D, Malygin A, Stahl J, Shatsky I, Karpova G. Positioning of subdomain IIId and apical loop of domain II of the hepatitis C IRES on the human 40S ribosome. Nucleic Acids Research. 2009; 37 (4):1141-1151. DOI: 10.1093/nar/gkn1026 - 84.
Boehringer D, Thermann R, Ostareck-Lederer A, Lewis JD, Stark H. Structure of the hepatitis C virus IRES bound to the human 80S ribosome: Remodeling of the HCV IRES. Structure. 2005; 13 (11):1695-1706. DOI: 10.1016/j.str.2005.08.008