The effects of statins on the mevalonic acid pathway in recombinant yeast strains expressing human HMG-CoA reductase

BMC Biotechnology

Agata Maciejak¹  Agata Leszczynska¹  Ilona Warchol¹  Monika Gora¹  Joanna Kaminska¹  Danuta Plochocka¹  Monika Wysocka-Kapcinska¹  Dorota Tulacz¹  Joanna Siedlecka¹  Ewa Swiezewska¹  Maciej Sojka²  Witold Danikiewicz²  Norbert Odolczyk¹  Anna Szkopinska¹  Grazyna Sygitowicz³  Beata Burzynska¹ 

[1] Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawinskiego 5A, 02-106 Warsaw, Poland

[2] Institute of Organic Chemistry, Polish Academy of Sciences, Warsaw, Poland

[3] Department of Biochemistry and Clinical Chemistry, Medical University of Warsaw, Warsaw, Poland

关键词: Mevalonate pathway;    Heterologous proteins;    Yeast expression system;    Statins;    HMG-CoA reductase;   

Others  :  1089658
DOI  :  10.1186/1472-6750-13-68

 received in 2013-04-03, accepted in 2013-08-29,  发布年份 2013

【 摘 要 】

Background

The yeast Saccharomyces cerevisiae can be a useful model for studying cellular mechanisms related to sterol synthesis in humans due to the high similarity of the mevalonate pathway between these organisms. This metabolic pathway plays a key role in multiple cellular processes by synthesizing sterol and nonsterol isoprenoids. Statins are well-known inhibitors of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR), the key enzyme of the cholesterol synthesis pathway. However, the effects of statins extend beyond their cholesterol-lowering action, since inhibition of HMGR decreases the synthesis of all products downstream in the mevalonate pathway. Using transgenic yeast expressing human HMGR or either yeast HMGR isoenzyme we studied the effects of simvastatin, atorvastatin, fluvastatin and rosuvastatin on the cell metabolism.

Results

Statins decreased sterol pools, prominently reducing sterol precursors content while only moderately lowering ergosterol level. Expression of genes encoding enzymes involved in sterol biosynthesis was induced, while genes from nonsterol isoprenoid pathways, such as coenzyme Q and dolichol biosynthesis or protein prenylation, were diversely affected by statin treatment. Statins increased the level of human HMGR protein substantially and only slightly affected the levels of Rer2 and Coq3 proteins involved in non-sterol isoprenoid biosynthesis.

Conclusion

Statins influence the sterol pool, gene expression and protein levels of enzymes from the sterol and nonsterol isoprenoid biosynthesis branches and this effect depends on the type of statin administered. Our model system is a cheap and convenient tool for characterizing individual statins or screening for novel ones, and could also be helpful in individualized selection of the most efficient HMGR inhibitors leading to the best response and minimizing serious side effects.

【 授权许可】

   
2013 Maciejak et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 1.

【 参考文献 】

• [1]Shepherd J, Cobbe SM, Ford I, Isles CG, Lorimer AR, Macfarlane PW, McKillop JH, Packard CJ: Prevention of coronary heart disease with pravastatin in men with hypercholesterolemia. N Engl J Med 1995, 333:1301-1308.
• [2]Prosser LA, Stinnett AA, Goldman PA, Williams LW, Hunink MG, Goldman L, Weinstein MC: Cost-effectiveness of cholesterol-lowering therapies according to selected patient characteristics. Ann Intern Med 2000, 132:769-779.
• [3]Sever PS, Dahlöf B, Poulter NR, Wedel H, Beevers G, Caulfield M, Collins R, Kjeldsen SE, Kristinsson A, McInnes GT, Mehlsen J, Nieminen M, O’Brien E, Ostergren J, ASCOT investigators: Prevention of coronary and stroke events with atorvastatin in hypertensive patients who have average or lower-than-average cholesterol concentrations, in the Anglo-Scandinavian Cardiac Outcomes Trial-Lipid Lowering Arm (ASCOT-LLA): a multicentre randomised controlled trial. Lancet 2003, 361:1149-1158.
• [4]Buhaescu I, Izzedine H: Mevalonate pathway: a review of clinic and therapeutical implications. Clin Biochem 2007, 40:575-584.
• [5]Evans M, Rees A: Effects of HMG-CoA reductase inhibitors on skeletal muscle. Drug Saf 2002, 25:649-663.
• [6]Onofrei MD, Butler KL, Fuke DC, Miller HB: Safety of statin therapy in patients with preexisting liver disease. Pharmacotherapy 2008, 28:522-529.
• [7]Zhou Q, Liao JK: Statins and cardiovascular disease: from cholesterol lowering to pleiotropy. Curr Pharm Des 2009, 15:467-478.
• [8]Botstein D, Chervitz SA, Cherry JM: Yeast as a model organism. Science 1997, 277:1259-1260.
• [9]Sturgeon CM, Kemmer D, Anderson HJ, Roberge M: Yeast as a tool to uncover the cellular targets of drugs. Biotechnol J 2006, 1:289-298.
• [10]Marjanovic J, Chalupska D, Patenode C, Coster A, Arnold E, Ye A, Anesi G, Lu Y, Okun I, Tkachenko S, Haselkorn R, Gornicki P: Recombinant yeast screen for new inhibitors of human acetyl-CoA carboxylase 2 identifies potential drugs to treat obesity. Proc Natl Acad Sci USA 2010, 107:9093-9098.
• [11]Wysocka-Kapcinska M, Lutyk-Nadolska J, Kiliszek M, Plochocka D, Maciag M, Leszczynska A, Rytka J, Burzynska B: Functional expression of human HMG-CoA reductase in Saccharomyces cerevisiae: a system to analyse normal and mutated versions of the enzyme in the context of statin treatment. J Appl Microbiol 2009, 106:895-902.
• [12]Nielsen J: Systems biology of lipid metabolism: from yeast to human. FEBS Lett 2009, 583:3905-3913.
• [13]Smith AM, Ammar R, Nislow C, Giaever G: A survey of yeast genomic assays for drug and target discovery. Pharmacol Ther 2010, 127:156-164.
• [14]Morikawa S, Murakami T, Yamazaki H, Izumi A, Saito Y, Hamakubo T, Kodama T: Analysis of the global RNA expression profiles of skeletal muscle cells treated with statins. J Atheroscler Thromb 2005, 12:121-131.
• [15]Leszczynska A, Gora M, Plochocka D, Hoser G, Szkopinska A, Koblowska M, Iwanicka-Nowicka R, Kotlinski M, Rawa K, Kiliszek M, Burzynska B: Different statins produce highly divergent changes in gene expression profiles of human hepatoma cells: a pilot study. Acta Biochim Pol 2011, 58:635-639.
• [16]Gerber R, Ryan JD, Clark DS: Cell-based screen of HMG-CoA reductase inhibitors and expression regulators using LC-MS. Anal Biochem 2004, 329:28-34.
• [17]Hagemenas FC, Illingworth DR: Cholesterol homeostasis in mononuclear leukocytes from patients with familial hypercholesterolemia treated with lovastatin. Arteriosclerosis 1989, 9:355-361.
• [18]Leszczynska A, Burzynska B, Plochocka D, Kaminska J, Zimnicka M, Kania M, Kiliszek M, Wysocka-Kapcinska M, Danikiewicz W, Szkopinska A: Investigating the effects of statins on cellular lipid metabolism using a yeast expression system. PLoS One 2009, 4:e8499.
• [19]Mabuchi H, Higashikata T, Kawashiri M, Katsuda S, Mizuno M, Nohara A, Inazu A, Koizumi J, Kobayashi J: Reduction of serum ubiquinol-10 and ubiquinone-10 levels by atorvastatin in hypercholesterolemic patients. J Atheroscler Thromb 2005, 2:111-119.
• [20]Rundek T, Naini A, Sacco R: Atorvastatin decreases the coenzyme Q10 level in the blood of patients at risk for cardiovascular disease and stroke. Arch Neurol 2004, 61:889-892.
• [21]Swiezewska E, Danikiewicz W: Polyisoprenoids: structure, biosynthesis and function. Prog Lipid Res 2005, 44:235-258.
• [22]Bergamini E: Dolichol: an essential part in the antioxidant machinery of cell membranes? Biogerontology 2003, 4:337-339.
• [23]Larsson O, Wejde J: Dolichol delays G1-arrest for one cell cycle in human fibroblasts subjected to depletion of serum or mevalonate. J Cell Sci 1992, 103:1065-1072.
• [24]Baksi K, Tavarez-Pagan JJ, Martinez JA, Banerjee DK: Unique structural motif supports mannosylphospho dolichol synthase: an important angiogenesis regulator. Cirr Drug Targets 2008, 9:262-271.
• [25]Endres M, Laufs U: Effects of statins on endothelium and signaling mechanisms. Stroke 2004, 35:2708-2711.
• [26]Takemoto M, Node K, Nakagami H, Liao Y, Grimm M, Takemoto Y, Kitakaze M, Liao JK: Statins as antioxidant therapy for preventing cardiac myocyte hypertrophy. J Clin Invest 2001, 108:1429-1437.
• [27]Fowler DM, Cooper SJ, Stephany JJ, Hendon N, Nelson S, Fields S: Suppression of statin effectiveness by copper and zinc in yeast and human cells. Mol Biosyst 2011, 7:533-544.
• [28]Takami T, Fang Y, Zhou X, Jaiseng W, Ma Y, Kuno T: A genetic and pharmacological analysis of isoprenoid pathway by LC-MS/MS in fission yeast. PLoS One 2012, 7:e49004.
• [29]Janke C, Magiera MM, Rathfelder N, Taxis C, Reber S, Maekawa H, Moreno-Borchart A, Doenges G, Schwob E, Schiebel E, Knop M: A versatile toolbox for PCR-based tagging of yeast genes: new fluorescent proteins, more markers and promoter substitution cassettes. Yeast 2004, 11:947-962.
• [30]Sato M, Fujisaki S, Sato K, Nishimura Y, Nakano A: Yeast Saccharomyces cerevisiae has two cis-prenyltransferases with different properties and localizations. Implication for their distinct physiological roles in dolichol synthesis. Genes Cells 2001, 6:495-506.
• [31]Rose M, Winston F, Hieter P: Methods in Yeast Genetics. A Cold Spring Harbor Laboratory Course. New York: Cold Spring Harbor Press; 1990.
• [32]Pfaffl MW: A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 2001, 29:e45.
• [33]Pfaffl MW, Horgan GW, Dempfle L: Relative expression software tool (REST©) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Res 2002, 9:e36.
• [34]Volland C, Urban-Grimal D, Géraud G, Haguenauer-Tsapis R: Endocytosis and degradation of the yeast uracil permease under adverse conditions. J Biol Chem 1994, 269:9833-9841.