JunShima FumikoTanaka AkiraAndo ToshihideNakamura HiroshiTakagi Gene Expression Omnibus (GEO) GEO NCBI NLM NIH http://www.ncbi.nlm.nih.gov/geo geo@ncbi.nlm.nih.gov GPL90 1 mRNA T128 Saccharomyces cerevisiae Typical commercial baker's yeast used in Japan Cell pellets (11,700 OD units) were suspended in 390 ml of LF medium in a 500-ml flask and then fermented at 30C for 300 min.To investigate gene expression profiles during initial stages of dough-fermentation, cell samples for DNA microarray analysis were obtained from each culture medium at 15 min, 30 min, and 60 min.(shima_15, shima_30, shima_60) Mini-scale fed-batch pre-cultivation was done using a jar fermentor (1-liter, B. E. Marubishi, Japan) and peristaltic pump (EYELA, Japan) with a sequential controller (B. E. Marubishi). After 18 h cultivation, cells in stationary phase were collected by centrifugation (2,700g for 5 min). Some of the cell pellets were suspended in 900 ml of sterilized water (4C). Cells for no-fermentation control were harvested after the fed-butch cultivation and stored at -80C until RNA extraction. (shima_0) polyA RNA Each time samples of total RNA were extracted using the hot phenol method. Poly (A)+ RNA was isolated from the total RNA by using an Oligotex dt30 (super) mRNA purification kit (Takara, Japan) according to the manufacturer's instructions. Yeast genome arrays (YGS98 GeneChip, Affymetrix, Santa Clara, CA) were used as DNA microarrays in this study. Biotin Labeling was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. DNA microarray hybridization was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. In brief, hybridization was at 45C in a hybridization oven (model 640, Affymetrix) at 60 rpm for 16 h. The EukGE-WS2v4 fluidics protocol of the Affymetrix Microarray Suite Version 5.0 (MASv5.0) software was used to perform staining and washing procedures. The arrays were subsequently read using a GeneArray Scanner (Agilent technologies, Palo Alto, CA). before fermentation Affymetrix Microarray Suite Version 5.0 Sample1.cel Sample1.exp ID_REF VALUE 0min ABS_CALL flag 11378_at 2.7 A 11379_at 1271.1 P 11380_at 172.2 P 11381_at 45.1 A 11382_at 17.2 A 11383_at 3.7 A 11384_at 18.5 A 11385_s_at 819.8 P 11386_at 4733.9 P 11387_at 695.5 P 11388_at 4420.3 P 11389_at 383.8 P 11390_at 24.3 A 11391_at 22.6 A 11392_at 210.9 P 11393_at 186.7 P 11356_at 5778.8 P 11357_at 1450.1 P 11358_at 227.3 P 11359_at 1273.6 P 1 mRNA T128 Saccharomyces cerevisiae Typical commercial baker's yeast used in Japan Cell pellets (11,700 OD units) were suspended in 390 ml of LF medium in a 500-ml flask and then fermented at 30C for 300 min.To investigate gene expression profiles during initial stages of dough-fermentation, cell samples for DNA microarray analysis were obtained from each culture medium at 15 min, 30 min, and 60 min.(after_15min, after_30min, after_60min) Mini-scale fed-batch pre-cultivation was done using a jar fermentor (1-liter, B. E. Marubishi, Japan) and peristaltic pump (EYELA, Japan) with a sequential controller (B. E. Marubishi). After 18 h cultivation, cells in stationary phase were collected by centrifugation (2,700g for 5 min). Some of the cell pellets were suspended in 900 ml of sterilized water (4C). Cells for no-fermentation control were harvested after the fed-butch cultivation and stored at -80C until RNA extraction. (before_fermentation) polyA RNA Each time samples of total RNA were extracted using the hot phenol method. Poly (A)+ RNA was isolated from the total RNA by using an Oligotex dt30 (super) mRNA purification kit (Takara, Japan) according to the manufacturer's instructions. Yeast genome arrays (YGS98 GeneChip, Affymetrix, Santa Clara, CA) were used as DNA microarrays in this study. Biotin Labeling was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. DNA microarray hybridization was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. In brief, hybridization was at 45C in a hybridization oven (model 640, Affymetrix) at 60 rpm for 16 h. The EukGE-WS2v4 fluidics protocol of the Affymetrix Microarray Suite Version 5.0 (MASv5.0) software was used to perform staining and washing procedures. The arrays were subsequently read using a GeneArray Scanner (Agilent technologies, Palo Alto, CA). before fermentation (replicate) Affymetrix Microarray Suite Version 5.0 Sample2.cel Sample2.exp ID_REF VALUE before_fermentation_2 ABS_CALL flag 11378_at 2.7 A 11379_at 15.5 A 11380_at 55.9 P 11381_at 1.5 A 11382_at 2.7 A 11383_at 2.2 A 11384_at 6.6 A 11385_s_at 175.5 P 11386_at 73 P 11387_at 2.7 A 11388_at 24.8 P 11389_at 10.8 A 11390_at 3.3 A 11391_at 6.5 A 11392_at 8 A 11393_at 22.8 A 11356_at 40.9 P 11357_at 20.1 A 11358_at 10.7 A 11359_at 20.7 A 1 mRNA T128 Saccharomyces cerevisiae Typical commercial baker's yeast used in Japan Cell pellets (11,700 OD units) were suspended in 390 ml of LF medium in a 500-ml flask and then fermented at 30C for 300 min.To investigate gene expression profiles during initial stages of dough-fermentation, cell samples for DNA microarray analysis were obtained from each culture medium at 15 min, 30 min, and 60 min.(after_15min, after_30min, after_60min) Mini-scale fed-batch pre-cultivation was done using a jar fermentor (1-liter, B. E. Marubishi, Japan) and peristaltic pump (EYELA, Japan) with a sequential controller (B. E. Marubishi). After 18 h cultivation, cells in stationary phase were collected by centrifugation (2,700g for 5 min). Some of the cell pellets were suspended in 900 ml of sterilized water (4C). Cells for no-fermentation control were harvested after the fed-butch cultivation and stored at -80C until RNA extraction. (before_fermentation) polyA RNA Each time samples of total RNA were extracted using the hot phenol method. Poly (A)+ RNA was isolated from the total RNA by using an Oligotex dt30 (super) mRNA purification kit (Takara, Japan) according to the manufacturer's instructions. Yeast genome arrays (YGS98 GeneChip, Affymetrix, Santa Clara, CA) were used as DNA microarrays in this study. Biotin Labeling was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. DNA microarray hybridization was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. In brief, hybridization was at 45C in a hybridization oven (model 640, Affymetrix) at 60 rpm for 16 h. The EukGE-WS2v4 fluidics protocol of the Affymetrix Microarray Suite Version 5.0 (MASv5.0) software was used to perform staining and washing procedures. The arrays were subsequently read using a GeneArray Scanner (Agilent technologies, Palo Alto, CA). fermentation 15min Affymetrix Microarray Suite Version 5.0 Sample3.cel Sample3.exp ID_REF VALUE 15min ABS_CALL flag 11378_at 2 A 11379_at 307.7 P 11380_at 24.1 P 11381_at 22.3 A 11382_at 1.9 A 11383_at 1.6 A 11384_at 8.5 A 11385_s_at 476.2 P 11386_at 554.1 P 11387_at 37.2 A 11388_at 439.1 P 11389_at 556.5 P 11390_at 23.3 A 11391_at 20.2 A 11392_at 1.3 A 11393_at 11.8 A 11356_at 238.3 P 11357_at 407.6 P 11358_at 42 P 11359_at 66.9 P 1 mRNA T128 Saccharomyces cerevisiae Typical commercial baker's yeast used in Japan Cell pellets (11,700 OD units) were suspended in 390 ml of LF medium in a 500-ml flask and then fermented at 30C for 300 min.To investigate gene expression profiles during initial stages of dough-fermentation, cell samples for DNA microarray analysis were obtained from each culture medium at 15 min, 30 min, and 60 min.(after_15min, after_30min, after_60min) Mini-scale fed-batch pre-cultivation was done using a jar fermentor (1-liter, B. E. Marubishi, Japan) and peristaltic pump (EYELA, Japan) with a sequential controller (B. E. Marubishi). After 18 h cultivation, cells in stationary phase were collected by centrifugation (2,700g for 5 min). Some of the cell pellets were suspended in 900 ml of sterilized water (4C). Cells for no-fermentation control were harvested after the fed-butch cultivation and stored at -80C until RNA extraction. (before_fermentation) polyA RNA Each time samples of total RNA were extracted using the hot phenol method. Poly (A)+ RNA was isolated from the total RNA by using an Oligotex dt30 (super) mRNA purification kit (Takara, Japan) according to the manufacturer's instructions. Yeast genome arrays (YGS98 GeneChip, Affymetrix, Santa Clara, CA) were used as DNA microarrays in this study. Biotin Labeling was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. DNA microarray hybridization was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. In brief, hybridization was at 45C in a hybridization oven (model 640, Affymetrix) at 60 rpm for 16 h. The EukGE-WS2v4 fluidics protocol of the Affymetrix Microarray Suite Version 5.0 (MASv5.0) software was used to perform staining and washing procedures. The arrays were subsequently read using a GeneArray Scanner (Agilent technologies, Palo Alto, CA). fermentation 15min (replicate) Affymetrix Microarray Suite Version 5.0 Sample4.cel Sample4.exp ID_REF VALUE 15min ABS_CALL flag 11378_at 6.1 A 11379_at 4.6 A 11380_at 9.2 A 11381_at 2.8 A 11382_at 12.5 A 11383_at 5.7 A 11384_at 14.2 A 11385_s_at 59.9 P 11386_at 34.6 A 11387_at 12.1 A 11388_at 55.4 P 11389_at 167.9 A 11390_at 4.2 A 11391_at 49.2 A 11392_at 3 A 11393_at 17.7 A 11356_at 24.3 A 11357_at 86.6 P 11358_at 32.1 A 11359_at 5.8 A 1 mRNA T128 Saccharomyces cerevisiae Typical commercial baker's yeast used in Japan Cell pellets (11,700 OD units) were suspended in 390 ml of LF medium in a 500-ml flask and then fermented at 30C for 300 min.To investigate gene expression profiles during initial stages of dough-fermentation, cell samples for DNA microarray analysis were obtained from each culture medium at 15 min, 30 min, and 60 min.(after_15min, after_30min, after_60min) Mini-scale fed-batch pre-cultivation was done using a jar fermentor (1-liter, B. E. Marubishi, Japan) and peristaltic pump (EYELA, Japan) with a sequential controller (B. E. Marubishi). After 18 h cultivation, cells in stationary phase were collected by centrifugation (2,700g for 5 min). Some of the cell pellets were suspended in 900 ml of sterilized water (4C). Cells for no-fermentation control were harvested after the fed-butch cultivation and stored at -80C until RNA extraction. (before_fermentation) polyA RNA Each time samples of total RNA were extracted using the hot phenol method. Poly (A)+ RNA was isolated from the total RNA by using an Oligotex dt30 (super) mRNA purification kit (Takara, Japan) according to the manufacturer's instructions. Yeast genome arrays (YGS98 GeneChip, Affymetrix, Santa Clara, CA) were used as DNA microarrays in this study. Biotin Labeling was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. DNA microarray hybridization was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. In brief, hybridization was at 45C in a hybridization oven (model 640, Affymetrix) at 60 rpm for 16 h. The EukGE-WS2v4 fluidics protocol of the Affymetrix Microarray Suite Version 5.0 (MASv5.0) software was used to perform staining and washing procedures. The arrays were subsequently read using a GeneArray Scanner (Agilent technologies, Palo Alto, CA). fermentation 30min Affymetrix Microarray Suite Version 5.0 Sample5.cel Sample5.exp ID_REF VALUE 30min ABS_CALL flag 11378_at 3.8 A 11379_at 185.7 P 11380_at 15.3 P 11381_at 119.7 P 11382_at 4 A 11383_at 1.5 A 11384_at 8.6 A 11385_s_at 1667.8 P 11386_at 220.4 P 11387_at 15.5 A 11388_at 307.1 P 11389_at 510.1 P 11390_at 39.9 A 11391_at 19.1 P 11392_at 4.6 A 11393_at 16.6 A 11356_at 22.4 A 11357_at 140.2 P 11358_at 34.6 P 11359_at 62.8 P 1 mRNA T128 Saccharomyces cerevisiae Typical commercial baker's yeast used in Japan Cell pellets (11,700 OD units) were suspended in 390 ml of LF medium in a 500-ml flask and then fermented at 30C for 300 min.To investigate gene expression profiles during initial stages of dough-fermentation, cell samples for DNA microarray analysis were obtained from each culture medium at 15 min, 30 min, and 60 min.(after_15min, after_30min, after_60min) Mini-scale fed-batch pre-cultivation was done using a jar fermentor (1-liter, B. E. Marubishi, Japan) and peristaltic pump (EYELA, Japan) with a sequential controller (B. E. Marubishi). After 18 h cultivation, cells in stationary phase were collected by centrifugation (2,700g for 5 min). Some of the cell pellets were suspended in 900 ml of sterilized water (4C). Cells for no-fermentation control were harvested after the fed-butch cultivation and stored at -80C until RNA extraction. (before_fermentation) polyA RNA Each time samples of total RNA were extracted using the hot phenol method. Poly (A)+ RNA was isolated from the total RNA by using an Oligotex dt30 (super) mRNA purification kit (Takara, Japan) according to the manufacturer's instructions. Yeast genome arrays (YGS98 GeneChip, Affymetrix, Santa Clara, CA) were used as DNA microarrays in this study. Biotin Labeling was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. DNA microarray hybridization was done according to the Affymetrix GeneChip Expression Analysis Technical Manual. In brief, hybridization was at 45C in a hybridization oven (model 640, Affymetrix) at 60 rpm for 16 h. The EukGE-WS2v4 fluidics protocol of the Affymetrix Microarray Suite Version 5.0 (MASv5.0) software was used to perform staining and washing procedures. The arrays were subsequently read using a GeneArray Scanner (Agilent technologies, Palo Alto, CA). fermentation 60min Affymetrix Microarray Suite Version 5.0 Sample6.cel Sample6.exp ID_REF VALUE 60min ABS_CALL flag 11378_at 8 A 11379_at 421.2 P 11380_at 77 P 11381_at 65.5 P 11382_at 2.3 A 11383_at 3.3 A 11384_at 41.4 M 11385_s_at 900.7 P 11386_at 338.6 P 11387_at 32.7 A 11388_at 916.8 P 11389_at 536.3 P 11390_at 56.3 A 11391_at 40.1 A 11392_at 18.9 A 11393_at 27.5 A 11356_at 271.4 P 11357_at 200.3 P 11358_at 60.4 P 11359_at 455.1 P

Gene expression profiles of baker's yeast during initial dough-fermentation were investigated using liquid fermentation media to obtain insights at the molecular level into rapid adaptation mechanisms of baker's yeast. Results showed that onset of fermentation caused drastic changes in gene expression profiles within 15 min. Genes involved in the tricarboxylic acid (TCA) cycle were down-regulated and genes involved in glycolysis were up-regulated, indicating a metabolic shift from respiration to fermentation. Genes involved in ethanol production (PDC genes and ADH1), in glycerol synthesis (GPD1 and HOR2), and in low-affinity hexose transporters (HXT1 and HXT3) were up-regulated at the beginning of model dough-fermentation. Among genes up-regulated at 15 min, several genes classified as transcription were down-regulated within 30 min. These down-regulated genes are involved in messenger RNA splicing and ribosomal protein biogenesis, in zinc finger transcription factor proteins, and in transcriptional regulator (SRB8, MIG1). In contrast, genes involved in amino acid metabolism and in vitamin metabolism, such as arginine biosynthesis, riboflavin biosynthesis, and thiamin biosynthesis, were subsequently up-regulated after 30 min. Interestingly, the genes involved in the unfolded protein response (UPR) pathway were also subsequently up-regulated. Our study presents the first overall description of the transcriptional response of baker's yeast during dough-fermentation, and will thus help clarify genomic responses to various stresses during commercial fermentation processes.

Saccharomyces cerevisiae T128 was used as a model of typical commercial baker's yeast used in Japan. After 18 h cultivation, cells in stationary phase were collected by centrifugation (2,700g for 5 min). Some of the cell pellets were suspended in 900 ml of sterilized water. Cells for no-fermentation control were harvested after the fed-butch cultivation and stored until RNA extraction. Cell pellets (11,700 OD units) were suspended in 390 ml of lequid fermentation (LF) medium in a 500-ml flask and then fermented for 300 min. To investigate gene expression profiles during initial stages of dough-fermentation, cell samples for DNA microarray analysis were obtained from each culture medium at 15 min, 30 min, and 60 min. Cells in stationary phase were then collected by centrifugation (2700 G for 5 min), and stored until RNA extraction. fermentation time before fermentation time fermentation 15 min time fermentation 30 min time fermentation 60 min