Yeast strains used in this study.

<p>Yeast strains used in this study.</p

Structural parameters of mitochondria from different mutant strains.

<p>Electron micrographs of the tested strains were obtained as described in the Materials and Methods section. For the analysis of mitochondrial size, circumference and area 70 to 100 mitochondria were tested.</p

Growth analysis of yeast strains deleted of genes up-regulated in the absence of <i>PSD1</i>.

<p>Strains as indicated were grown on YPD, YPLac, MMLac, YPGlycerol, YPSorbitol, and YPD with 0.05% SDS. Cell suspensions of strains listed were spotted at dilutions 1, 1/10, 1/100, 1/1000, and 1/10000. Incubation was carried out at 30°C.</p

Analysis of organelles from wild type, <i>cho2Δopi3Δ</i> and <i>cki1Δdpl1Δeki1Δ</i> yeast strains.

<p>Yeast strains used were grown on YPO media. Data obtained with isolated peroxisomes (lane A), mitochondria (lane B) and endoplasmic reticulum (lane C) are shown. Line 1: Phospholipid pattern expressed as μg of individual phospholipids per mg protein. LP, lysophospholipids; PI, phosphatidylinositol; PS, phosphatidylserine; PC, phosphatidylcholine; PE, phosphatidylethanolamine; CL, cardiolipin; PA, phosphatidic acid. Line 2: PC to PE ratio in different organelles and strains. Line 3: Fatty acid composition of different organelles and strains. Line 4: Ratio of saturated (SFA) to unsaturated (UFA) fatty acids in different organelles and strains. Line 5: Ergosterol (ERG) to phospholipid (PL) ratio in different organelles and strains. Line 6: Anisotropy values obtained with different organelles and strains. For experimental details see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135084#sec002" target="_blank">Materials and Methods</a> section. For all experiments two independent biological samples were used which were analyzed 2 to 3 time, each.</p

Quantification of Western blot analysis of subcellular fractions from <i>S</i>. <i>cerevisiae</i>.

<p>ER (endoplasmic reticulum), M (mitochondria) and PX (peroxisomes) were compared to the H (homogenate) of the wild type strain (A), the <i>cki1</i>Δ<i>dpl1</i>Δ<i>eki1</i>Δ (B) and the <i>cho2</i>Δ<i>opi3</i>Δ (C) mutant strains.</p

Gene expression analysis.

<p>Relative gene expression of <i>PSD2</i>, <i>ALE1</i>, <i>EKI1</i>, <i>EPT1</i>, <i>DPL1</i>, <i>GPH1</i>, <i>GPM2</i> and <i>RSB1</i> was measured by qRT-PCR from isolated RNA of wild type and <i>Δpsd1</i>. Expression of the respective genes in wild type was set at 1 and values obtained with RNA isolated from <i>Δpsd1</i> were set in relation. Data are mean values from three independent experiments with the respective deviation.</p

Fatty acids in the homogenate from wild type, <i>cho2Δopi3Δ</i> and <i>cki1Δdpl1Δeki1Δ</i> yeast strains.

<p>Yeast strains used were grown on YPO media, and homogenate samples were prepared after cell disruption. The oleic acid content was quantified as described in the Materials and Methods section.</p

Pathways of phosphatidylcholine biosynthesis in the yeast <i>S</i>. <i>cerevisiae</i>.

<p>Cct1, cholinephosphate cytidylyltransferase; CDP-Cho, cytidine-diphosphocholine, CDP-Etn, cytidine-diphosphoethanolamine; Cho, Choline; Cho2, phosphatidylethanolamine methyltransferase; Cho-P, phosphocholine; Cki1, choline kinase; Cpt1, cholinephosphotransferase; DAG, diacylglycerol; Dpl1, sphingosine phosphate lyase; Ect1, phosphoethanolamine cytidylyltransferase; Eki1, ethanolamine kinase; Ept1, sn-1,2-diacylglycerol ethanolamine- and cholinephosphotranferase; Etn, ethanolamine; Etn-P, phosphoethanolamine; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PDME, phosphatidyldimethylethanolamine; PMME phosphatidylmonomethylethanolamine; PS, phosphatidylserine; Psd1, phosphatidylserine decarboxylase 1; Psd2, phosphatidylserine decarboxylase 2; Opi3, methylene-fatty-acyl-phospholipid synthase; SAM, S-adenosyl-L-methionine; SL, sphingolipids.</p

Growth phenotype of wild type, <i>cho2Δ</i>, <i>opi3Δ</i>, <i>cho2Δopi3Δ</i> and <i>cki1Δdpl1Δeki1Δ</i> yeast strains.

<p>(A): Drop test on YPD plates (30°C and 37°C); on minimal oleate media containing choline and ethanolamine; and on YPO plates are shown. (B) Growth of liquid cultures on YPO.</p

Gene expression of <i>GPH1</i>, <i>GPM2</i> and <i>RSB1</i> in <i>Δpsd1Δpsd2</i> with variable supplementation of ethanolamine.

<p>Relative gene expression of <i>GPH1</i>, <i>GPM2</i> and <i>RSB1</i> was measured by qRT-PCR from isolated RNA of wild type and <i>Δpsd1Δpsd2</i> cultivated in the presence of different amounts of ethanolamine (values in brackets). Expression of the respective genes in wild type supplemented to a final concentration of 5 mM was set at 1, and values obtained with RNA isolated from <i>Δpsd1Δpsd2</i> supplemented with 5 mM or 0.25 mM ethanolamine, respectively, were set in relation. Data are mean values from three independent experiments with the respective deviation.</p