EVOLUTIONARY-OBTAINED saccharomyces cerevisiae, RESISTANT TO DMEM CULTURED WITH PROSTATE CANCER CELLS

Abstract



The complexity of the cell is a major problem to study on cancer treatment. In order to find an accurate solution to cancer, it is necessary to know the cell metabolism in detail, but the available knowledge and technology are inadequate. Herein, there is a need for alternative approaches to suggest a treatment way against cancer instead of trying to understand the cell metabolism in details. Therefore, this study aimed to obtain the desired phenotype of Saccharomyces cerevisiae which can easily grow in the DMEM pre-cultured with PC3 (PCM).

Evolutionary engineering was successfully applied to wild type (WT) yeast population to randomly generate a variety of genetic phenotypes. The resistant mutants against PCM were randomly selected from an agar plate. Furthermore, the best individual mutant being resistant in PCM was determined with genetic stability tests.

In the current study, the resistant yeasts were obtained by evolutionary engineering against PCM restricting the growth of WT. The growth fitness of selected mutants dramatically increased in the PCM, when compared to WT. The best mutant, MY2, is an example to be resistant in extreme environment with directing the instinct of survival of organisms. To suggest as a cancer treatment, the secondary metabolites of MY2 on cancer cells have to be tested.

Keywords

• Saccharomyces cerevisiae,cancer,evolutionary engineering,PC3,yeast

EVOLUTIONARY-OBTAINED saccharomyces cerevisiae, RESISTANT TO DMEM CULTURED WITH PROSTATE CANCER CELLS

Abstract

Hücrenin karmaşıklığı kanser tedavisi geliştirmek için büyük bir sorundur. Kansere karşı doğru bir çözüm bulmak için, hücre metabolizmasını ayrıntılı olarak bilmek gerekir, ancak mevcut bilgi ve teknoloji bunun için yetersizdir. Bu noktada, kanser tedavisi için alternatif yaklaşımlara ihtiyaç duyulmaktadır. Bu nedenle, bu çalışma, PC3 (PCM) ile önceden kültürlenmiş DMEM'de kolaylıkla büyüyebilen mutant Saccharomyces cerevisiae fenotipinin elde edilmesini amaçlamıştır.

 

Evrimsel mühendislik yöntemi başarılı bir şekilde uygulanarak, doğal tip maya (WT) popülasyonunda çeşitli genetik fenotipler rasgele üretildi. PCM'ye karşı dirençli mutantlar bir agar plakasından rastgele olacak şekilde seçildi. Ayrıca, Seçilen mutant feneotipler içinden PCM'de dirençli en iyi bireysel mutant genetik kararlılık testleri ile belirlendi.

 

Mevcut çalışmada, WT'nin büyümesini sınırlayan PCM'ye karşı evrimsel mühendislik yöntemiyle dirençli muatantlar elde edildi. Seçilen mutantların PCM içindeki büyüme kapasitesi, WT'ye kıyasla anlamlı biçimde arttı. En iyi mutant olan MY2, organizmaların hayatta kalma içgüdüsünü yönlendiren uç koşullarda dirençli olmanın bir örneğidir. Kanser hücresi büyütülen ortamda rahatlıkla büyüye bilen MY2, üretiği mikincil metabolitlerin kanser hücreleri üzerinde test edilmesi gerekir. Elde edilen sonuçlara göre kanser tedavisinde kullanılabilirliği değerlendirilmelidir.

Keywords

• saccharomyces cerevisiae,cancer,Evolutionary engineering,PC3,yeast

References

1. [1] Meng, X., Zhong, J., Liu, S., Murray, M., Gonzalez-Angulo, AM., A New Hypothesis for the Cancer Mechanism, Cancer and Metas. Rev. 31(1–2), 247–68, 2012.
2. [2] Bertram, JS., The Molecular Biology of Cancer, Mol. Asp. of Med. 21, 167-223. 2001.
3. [3] Hanahan, D., Weinberg, RA., Hallmarks of Cancer: The next Generation, Cell 144(5), 646–74, 2011.
4. [4] Chen, JLY., Lucas, JE., Schroeder, T., Mori, S., Wu, J., Nevins, J., Dewhirst, M., West, M., Chi1, JT., The Genomic Analysis of Lactic Acidosis and Acidosis Response in Human Cancers, PLoS Genet. 4(12), e1000293, 2008.
5. [5] DeBerardinis, RJ., Lum, JJ., Hatzivassiliou, G., Thompson, CB., The Biology of Cancer: Metabolic Reprogramming Fuels Cell Growth and Proliferation, Cell Metab. 7(1), 11–20, 2008.
6. [6] Vogelstein, B., Kinzler, KW., Cancer Genes and the Pathways They Control, Nat, Med. 10(8), 789–99, 2004.
7. [7] Semenza GL., HIF-1: Upstream and Downstream of Cancer Metabolism, Current Opinion in Genet, Dev. 20(1), 51–56, 2010.
8. [8] Dawson, MA., Kouzarides, T., Cancer Epigenetics: From Mechanism to Therapy, Cell 150(1), 12–27, 2012.

9. [9] Francescone, R., Hou, V., Grivennikov, SI., Microbiome, Inflammation and Cancer, Cancer J. 20(3), 181–89, 2015.
10. [10] Louis, P., Hold, GL., Flint, HJ., The Gut Microbiota, Bacterial Metabolites and Colorectal Cancer, Nat Rev Microbiol 12(10), 661–72, 2014.
11. [11] Bultman, SJ., Emerging Roles of the Microbiome in Cancer, Carcinogenesis, 35(2), 249–55, 2014.
12. [12] Ma, D., Forsythe, P., Bienenstock, J.,. Live Lactobacillus Reuteri Is Essential for the Inhibitory Effect on Tumor Necrosis Factor Alpha-Induced Interleukin-8 Expression Live Lactobacillus Reuteri Is Essential for the Inhibitory Effect on Tumor Necrosis Factor Alpha-Induced Interleukin-8 Expres, Infect. Immun. 72(9), 5308–5314, 2004.
13. [13] Vaishnava, S., Behrendta, CL., Ismaila, AS., Eckmannb, L., Hooper, L., Paneth Cells Directly Sense Gut Commensals and Maintain Homeostasis at the Intestinal Host-Microbial Interface, Proc. Natl. Acad. Sci. 105(52), 20858–20863, 2008.
14. [14] Garrett, WS., Gordon, JI., Glimcher, LH., Homeostasis and Inflammation in the Intestine, Cell 140(6), 859–70, 2010.
15. [15] Holmes, E., Li, JV., Marchesi, JR., Nicholson, JK., Gut Microbiota Composition and Activity in Relation to Host Metabolic Phenotype and Disease Risk, Cell Metab. 16(5), 559–564, 2012.
16. [16] Laxminarayan, R., Duse, A., Wattal, C., Zaidi, AK., Wertheim, HF., Sumpradit, N., Vlieghe, E., Hara, GL., Gould, IM., Goossens, H., Greko, C., So, AD., Bigdeli, M., Tomson, G., Woodhouse, W, Ombaka, E., Peralta, AQ., Qamar, FN., Mir, F., Kariuki, S., Bhutta, ZA., Coates, A., Bergstrom, R., Wright, GD., Brown, ED., Cars, O., Antibiotic resistance the need for global solutions, Lancet Infect. Dis 13, 1057–1098, 2013.
17. [17] Zinder, D., Bedford, T., Gupta, S., Pascual, M., The Roles of Competition and Mutation in Shaping Antigenic and Genetic Diversity in Influenza, PLoS Pathog. 9(1), e1003104, 2013.
18. [18] Fritz, JV., Desai, MS., Shah, P., Schneider, JG., Wilmes, P., From Meta-Omics to Causality: Experimental Models for Human Microbiome Research, Microbiome 1(1), 14, 2013.
19. [19] Bailey, JE., Toward a Science of Metabolic Engineering, Science, 252 (5013), 1668–75, 1991.
20. [20] Cakar, ZP., Metabolic and evolutionary engineering research in Turkey and beyond, Biotechnol. J. 4, 1–11, 2009.
21. [21] Cakar, ZP., Turanlı-Yıldız, B., Alkım, C., Yılmaz, Ü., Evolutionary Engineering of Saccharomyces Cerevisiae for Improved Industrially Important Properties, FEMS Yeast Res. 12(3), 171–182, 2012.
22. [22] Bailey, JE., Sburlati, A., Hatzimanikatis, V., Lee, K., Renner, WA., Tsai, PS., Inverse Metabolic Engineering: A Strategy for Directed Genetic Engineering of Useful Phenotypes, Biotechnol. Bioeng. 52(1), 109–121, 1996.
23. [23] Küçükgöze, G., Alkım, C., Yılmaz, Ü., Kısakesen, HI., Gündüz, S., Akman, S., Çakar, ZP., Evolutionary Engineering and Transcriptomic Analysis of Nickel-Resistant Saccharomyces Cerevisiae, FEMS Yeast Res. 13(8), 731–746, 2013.
24. [24] Almario, MP., Reyes, LH., Kao, KC., Evolutionary Engineering of Saccharomyces Cerevisiae for Enhanced Tolerance to Hydrolysates of Lignocellulosic Biomass, Biotech, Bioeng. 110(10), 2616–23, 2013.
25. [25] Şen, M., Yılmaz, Ü., Baysal, A., Akman, S., Çakar, ZP., In Vivo Evolutionary Engineering of a Boron-Resistant Bacterium : Bacillus Boroniphilus, Antonie van Leewenhoek 99, 825–35, 2011.
26. [26] Liu, L., Pana, A., Spofford, C., Zhoua, N., Alper, HS., An Evolutionary Metabolic Engineering Approach for Enhancing Lipogenesis in Yarrowia Lipolytica, Metab. Eng. 29, 36–45, 2015.
27. [27] Morales, P., Gentina, JC., Aroca, G., Mussatto, SI., Development of an Acetic Acid Tolerant Spathaspora Passalidarum Strain through Evolutionary Engineering with Resistance to Inhibitors Compounds of Autohydrolysate of Eucalyptus Globulus, Ind. Crops Prod. 106, 5-11, 2017.
28. [28] Lee, S., Jellison, T., Alper, HS., Systematic and Evolutionary Engineering of a Xylose Isomerase-Based Pathway in Saccharomyces Cerevisiae for Efficient Conversion Yields, Biotechnology for biofuels, 7(1), 122, 2014.
29. [29] Lee, SW., Oh MK., A Synthetic Suicide Riboswitch for the High-Throughput Screening of Metabolite Production in Saccharomyces Cerevisiae, Metab. Eng. 28, 143–150, 2015.
30. [30] Zheng, DQ., Wu, XC., Tao, XL., Wang, PM., Li, P., Chi, XQ., Li, YD., Yan, QF., Zhao, YH., Screening and Construction of Saccharomyces Cerevisiae Strains with Improved Multi-Tolerance and Bioethanol Fermentation Performance, Bioresour. Technol. 102(3), 3020–3027, 2011.
31. [31] Zumbansen, M., Altrogge, LM., Spottke, NUE., Spicker, S., Offizier, SM., Domzalski, SBS., Amand, ALS., Toell, A., Leake, D., Mueller- Hartmann, HA., First siRNA Library Screening in Hard-to-Transfect HUVEC Cells, J. RNAi Gene Silencing, 6(1), 354–360, 2010.
32. [32] Cakar, ZP., Alkım, C., Turanlı, B., Tokman, N., Akman, S., Sarıkaya, M., Tamerler, C., Benbadis, L., François, JM., Isolation of Cobalt Hyper-Resistant Mutants of Saccharomyces Cerevisiae by in Vivo Evolutionary Engineering Approach, J. Biotechno, 143,130–138, 2009.
33. [33] Lawrence, CW., Classical Mutagenesis Techniques, Methods Enzymol. 350(1988), 189–199, 2002.
34. [34] Russek, E., Colwell, RR., Computation of Most Probable Numbers, Appl. Environ. Microbiol. 45(5), 1646–1650, 1983.
35. [35] Memarian, N., Matthew, J., Javad, A., Nadereh, MR., Jianhua, X., Mehri, Z., Myron, S., Ashkan, G., Colony size measurement of the yeast gene deletion strains for functional genomics, BMC Bioinformatics, 8(1), 117, 2007.
36. [36] Liu, Y., Zuckier, LS., Ghesani, NV., Dominant Uptake of Fatty Acid over Glucose by Prostate Cells: A Potential New Diagnostic and Therapeutic Approach, Anticancer Res. 30 (2), 369–374, 2010.
37. [37] Berridge, MV., Herst, PM., Tan, AS., Metabolic Flexibility and Cell Hierarchy in Metastatic Cancer, Mitochondrion, 10(6), 584–88, 2010.
38. [38] Jang, M., Kim, SS., Lee, J., Cancer Cell Metabolism: Implications for Therapeutic Targets, Exp. Mol. Med. 45(10), e45, 2013.
39. [39] Meacham, CE., Morrison, SJ., Tumour Heterogeneity and Cancer Cell Plasticity, Nature, 501(7467), 328–37, 2013.
40. [40] Salari, R., Salari, R., Investigation of the Best Saccharomyces Cerevisiae Growth Condition, Electronic Physician, 9(1), 3592–97, 2017.
41. [41] Patnaik, R., Engineering complex phenotypes in industrial strains, Biotechnol. Prog. 24(1), 38–47, 2008.
42. [42] Tilloy, V., Cadière, A., Ehsani, M., Dequin, S., Reducing Alcohol Levels in Wines through Rational and Evolutionary Engineering of Saccharomyces Cerevisiae, Int. J. Food Microbiol. 213, 49–58, 2015.
43. [43] Dragosits, M., Mozhayskiy, V., Quinones-Soto, S., Park, J., Tagkopoulos, I., Evolutionary Potential, Cross-Stress Behavior and the Genetic Basis of Acquired Stress Resistance in Escherichia Coli, Mol. Syst. Biol. 9 (510), 1-13, 2013.