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## Results: 6

1.

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**A**) Comparison of the growth curves of*S. cerevisiae*BY4741 in the absence (▪) or presence of 2% v/v (□), 4% v/v (Δ) and 6% v/v (○) of ethanol. (**B**) Comparison of the growth curves of*S. cerevisiae*BY4741 (▪, □) and the corresponding deletion mutant*fps1Δ*(♦, ◊) in the absence (closed symbols) or presence of 2% v/v of ethanol (open symbols).2.

Changes found to occur (as indicated by the black or white arrows) or not (□,▪) in yeast cells growing in the presence of inhibitory concentrations of ethanol (closed symbols), or upon

*FPS1*deletion (open symbols) are shown. The metabolic scheme was based on information gathered in the KEGG PATHWAY Database (http://www.genome.jp/kegg/pathway.html) and in Yeast Biochemical Pathway Database (http://pathway.yeastgenome.org/).3.

Samples were collected during the exponential phase of growth (OD

_{600 nm}= 1.0) in the same basal medium supplemented with 0 or 2% v/v of ethanol. (**A**) 2D score plot displaying the space formed by the two first principal components (R^{2}X for PC1 equal to 0.375 and R^{2}X for PC2 equal to 0.284), (**B**) dendrogram based on the PCA scores (ward clustering distance measure) considering all principal components of the model and (**C**) relative abundances of different metabolites in the exo-metabolome of the parental strain and the*fps1Δ*deletion mutant during the exponential growth phase (OD_{600 nm}= 1.0) in the presence of different ethanol concentrations (0 and 2% v/v). Experimental values are means of four independent experiments with standard deviation error bars.4.

Samples were collected during the exponential phase of growth (OD

_{600 nm}= 1.0) in the same basal medium supplemented with different ethanol concentrations (% v/v). (**A**) 2D score plot displaying the space formed by the two first principal components (R^{2}X for PC1 equal to 0.522 and R^{2}X for PC2 equal to 0.166), (**B**) 2D loading plot displaying the space formed by the two first principal components and presenting the characteristic bins for a set of metabolites and (**C**) relative abundance of different metabolites in the exo-metabolome of*S. cerevisiae*BY4741 cells during the exponential phase of growth (OD_{600 nm}= 1.0) in the presence of different ethanol concentrations (0, 2, 4 and 6% v/v). Experimental values are means of four independent experiments with standard deviation error bars. Key:*1*Orotic acid,*2*Succininc acid,*3*2-Oxoglutaric acid,*4*L-leucine,*5*L-methionine,*6*Glucose,*7*L-histidine,*8*Glycerol,*9*Formic acid,*10*Uracil,*11*Acetic acid,*12*Pyruvic acid,*13*Acetoin,*14*Acetaldehyde,*15*Lactic acid,*16*L-alanine and*17*Fumaric acid.5.

Yeast cells were harvested during the exponential phase of growth (OD

_{600 nm}= 1.0) in the same basal medium supplemented with 0 or 2% v/v of ethanol. (**A**) 2D score plot displaying the space formed by the two first principal components (R^{2}X for PC1 equal to 0.525 and R^{2}X for PC2 equal to 0.321), (**B**) 2D loading plot displaying the space formed by the two first principal components (with characteristic bins for a set of metabolites) and (**C**) relative abundances of different metabolites in the endo-metabolome of the parental strain and the*fps1Δ*deletion strain during the exponential growth phase (OD_{600 nm}= 1.0) in the presence of different ethanol concentrations (0 and 2% v/v). Experimental values are means of six independent experiments with standard deviation error bars. Key:*1*NAD^{+},*2*NADP^{+},*3*L-tyrosine,*4*Fumaric acid,*5*L-phenylalanine,*6*Citric acid,*7*Succininic acid,*8*Glycerol,*9*L-glutamic acid,*10*L-leucine,*11*L-methionine, 12 L-alanine,*13*L-ornithine,*14*L-threonine,*15*L-valine,*16*L-glutamine,*17*L-aspartic acid,*18*L-malic acid,*19*L-histidine,*20*L-serine,*21*L-asparagine,*22*L-lysine,*23*Thiamine,*24*NADH and*25*L-proline.6.

Yeast cells were harvested during the exponential phase of growth (OD

_{600 nm}= 1.0) in the same basal medium and supplemented with different ethanol concentrations (% v/v). (**A**) 2D score plot displaying the space formed by the two first principal components (R^{2}X for PC1 equal to 0.792 and R^{2}X for PC2 equal to 0.129), (**B**) 2D loading plot displaying the space formed by the two first principal components and presenting the characteristic bins for a set of metabolites and (**C**) Relative abundance of different metabolites in the endo-metabolome of*S. cerevisiae*BY4741 cells harvested during the exponential phase of growth (OD_{600 nm}= 1.0) in the presence of different ethanol concentrations (0, 2, 4 and 6% v/v). Each plot displays the variation in the relative abundance of a representative bin in the spectrum for each amino acid. Experimental values are means of six independent experiments with standard deviation error bars. Key:*1*Glycerol,*2*L-arginine,*3*NAD^{+},*4*NADP^{+},*5*L-tyrosine,*6*L-histidine,*7*Trehalose,*8*NADH,*9*Fumaric acid,*10*L-proline,*11*L-phenylalanine,*12*L-asparagine,*13*Citric acid,*14*L-serine,*15*Succininic acid,*16*L-aspartic acid,*17*Malic acid,*18*L-glutamine,*19*L-alanine,*20*L-lysine,*21*L-glycine,*22*L-threonine,*23*L-valine,*24*L-methionine,*25*L-leucine,*26*L-glutamic acid and*27*L-ornithine.