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Transcriptome analysis of a respiratory Saccharomyces cerevisiae strain suggests the expression of its phenotype is glucose insensitive and predominantly controlled by Hap4, Cat8 and Mig1.

Bonander N, Ferndahl C, Mostad P, Wilks MD, Chang C, Showe L, Gustafsson L, Larsson C, Bill RM - BMC Genomics (2008)

Bottom Line: In addition, 13% were found to have a binding site for Cat8 and 21% had a binding site for Mig1.Our dataset gives a remarkably complete view of the involvement of genes in the TCA cycle, glyoxylate cycle and respiratory chain in the expression of the phenotype of V5.TM6*P.Furthermore, 88% of the transcriptional response of the induced genes in our dataset can be related to the potential activities of just three proteins: Hap4, Cat8 and Mig1.

View Article: PubMed Central - HTML - PubMed

Affiliation: School of Life and Health Sciences, Aston University, Aston Triangle, Birmingham, B4 7ET, UK. n.bonander@aston.ac.uk

ABSTRACT

Background: We previously described the first respiratory Saccharomyces cerevisiae strain, KOY.TM6*P, by integrating the gene encoding a chimeric hexose transporter, Tm6*, into the genome of an hxt yeast. Subsequently we transferred this respiratory phenotype in the presence of up to 50 g/L glucose to a yeast strain, V5 hxt1-7Delta, in which only HXT1-7 had been deleted. In this study, we compared the transcriptome of the resultant strain, V5.TM6*P, with that of its wild-type parent, V5, at different glucose concentrations.

Results: cDNA array analyses revealed that alterations in gene expression that occur when transitioning from a respiro-fermentative (V5) to a respiratory (V5.TM6*P) strain, are very similar to those in cells undergoing a diauxic shift. We also undertook an analysis of transcription factor binding sites in our dataset by examining previously-published biological data for Hap4 (in complex with Hap2, 3, 5), Cat8 and Mig1, and used this in combination with verified binding consensus sequences to identify genes likely to be regulated by one or more of these. Of the induced genes in our dataset, 77% had binding sites for the Hap complex, with 72% having at least two. In addition, 13% were found to have a binding site for Cat8 and 21% had a binding site for Mig1. Unexpectedly, both the up- and down-regulation of many of the genes in our dataset had a clear glucose dependence in the parent V5 strain that was not present in V5.TM6*P. This indicates that the relief of glucose repression is already operable at much higher glucose concentrations than is widely accepted and suggests that glucose sensing might occur inside the cell.

Conclusion: Our dataset gives a remarkably complete view of the involvement of genes in the TCA cycle, glyoxylate cycle and respiratory chain in the expression of the phenotype of V5.TM6*P. Furthermore, 88% of the transcriptional response of the induced genes in our dataset can be related to the potential activities of just three proteins: Hap4, Cat8 and Mig1. Overall, our data support genetic remodelling in V5.TM6*P consistent with a respiratory metabolism which is insensitive to external glucose concentrations.

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(A) The influence of multiple Hap complex sites on the change in gene expression of the genes in Additional file 1. A Fisher's exact test was performed for genes containing predicted binding sites for the Hap complex and their association with induction or repression. Genes were grouped into either those with one Hap complex binding site or those with two or more according to Additional file 1. (B) The influence of Hap4, Cat8 and Mig1 on the magnitude of induction of the genes in Additional file 1. The average fold-induction of genes in Additional file 1, grouped according to the transcription factor binding sites they contain, was calculated. It was observed that those containing binding sites for each of the Hap complex (where Hap4 is the activator), Cat8 and Mig1, had the highest average fold induction with Mig1 being a dominant factor in high induction. A bar for Cat8 is not included as only one gene (YOR019W, factor change 2.1) has a Cat8 site alone. (C) A Venn diagram of the distribution of binding sites for the Hap complex, Cat8 and Mig1 in the genes in Additional file 1. 88% of the genes in Additional file 1 contain a binding site for one or more of the Hap complex, Cat8 and Mig1. The range of induction for the 190 genes from Additional file 1 is 2.0 to 21.7.
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Figure 3: (A) The influence of multiple Hap complex sites on the change in gene expression of the genes in Additional file 1. A Fisher's exact test was performed for genes containing predicted binding sites for the Hap complex and their association with induction or repression. Genes were grouped into either those with one Hap complex binding site or those with two or more according to Additional file 1. (B) The influence of Hap4, Cat8 and Mig1 on the magnitude of induction of the genes in Additional file 1. The average fold-induction of genes in Additional file 1, grouped according to the transcription factor binding sites they contain, was calculated. It was observed that those containing binding sites for each of the Hap complex (where Hap4 is the activator), Cat8 and Mig1, had the highest average fold induction with Mig1 being a dominant factor in high induction. A bar for Cat8 is not included as only one gene (YOR019W, factor change 2.1) has a Cat8 site alone. (C) A Venn diagram of the distribution of binding sites for the Hap complex, Cat8 and Mig1 in the genes in Additional file 1. 88% of the genes in Additional file 1 contain a binding site for one or more of the Hap complex, Cat8 and Mig1. The range of induction for the 190 genes from Additional file 1 is 2.0 to 21.7.

Mentions: Since six of the genes from the HAP4 over-expression study did not contain Hap complex binding sites, we did a further in silico analysis of the induced genes of Additional file 1. We noted that the genes of Table 2, contained motifs that could be sub-divided into 2 distinct families: CCAATG (which we denoted as a Hap4_1 site) and (G/C)CAA(G/T)CAA (a Hap4_2 site). The bold sequences are the conserved sequences previously identified in the HAP4 over-expression strain study [19]. The genes of Additional file 1 were then examined 500 nucleotides upstream and 200 nucleotides downstream from the start codon using WebMOTIF [24] for these two biologically-relevant sites since it has been previously demonstrated that Hap complex binding sites are statistically over-represented up to 400 nucleotides upstream from the start codon [19]: we found that 112/146 (77%) induced genes and 127/190 (67%) of the complete dataset in Additional file 1 had one or more Hap complex binding site (Additional files 1 and 4). Our analysis included presumed binding sites such as CAAATC and CCAAAC which arose from the biologically-verified data on transcription factors (Table 2). For example, AQY2 is identified as Hap complex-dependent, and has two CCAAAC sites, but no CCAATNA sites. There are only two other genes that do not contain CCAATNA sites in Table 2, namely CRC1 and ALD6. In addition it was notable that Hap complex consensus sequences are predominantly found upstream of mitochondrial genes with 84/127 (66%) of such genes having at least a partial mitochondrial location. Furthermore, of the induced genes with a known cellular location and at least two Hap complex binding sites, 69/80 (86%) are localized to the mitochondria. It appears, then, that Hap4 activation is preferentially directed at mitochondrially-related functions especially within the TCA-cycle and respiratory chain, but cytosolic support via the glyoxylate cycle is also potentially regulated by Hap4 activation (Additional file 1). It was further noted that multiple Hap complex binding sites are associated with induction (Fig. 3A). A Fisher's exact test showed that there was a statistically-significant association between the change in gene expression and the number of Hap complex binding sites in the gene (p = 0.00). This supported the fact that genes with two or more Hap complex binding sites are likely to be induced, whereas those with one site may be either induced or repressed.


Transcriptome analysis of a respiratory Saccharomyces cerevisiae strain suggests the expression of its phenotype is glucose insensitive and predominantly controlled by Hap4, Cat8 and Mig1.

Bonander N, Ferndahl C, Mostad P, Wilks MD, Chang C, Showe L, Gustafsson L, Larsson C, Bill RM - BMC Genomics (2008)

(A) The influence of multiple Hap complex sites on the change in gene expression of the genes in Additional file 1. A Fisher's exact test was performed for genes containing predicted binding sites for the Hap complex and their association with induction or repression. Genes were grouped into either those with one Hap complex binding site or those with two or more according to Additional file 1. (B) The influence of Hap4, Cat8 and Mig1 on the magnitude of induction of the genes in Additional file 1. The average fold-induction of genes in Additional file 1, grouped according to the transcription factor binding sites they contain, was calculated. It was observed that those containing binding sites for each of the Hap complex (where Hap4 is the activator), Cat8 and Mig1, had the highest average fold induction with Mig1 being a dominant factor in high induction. A bar for Cat8 is not included as only one gene (YOR019W, factor change 2.1) has a Cat8 site alone. (C) A Venn diagram of the distribution of binding sites for the Hap complex, Cat8 and Mig1 in the genes in Additional file 1. 88% of the genes in Additional file 1 contain a binding site for one or more of the Hap complex, Cat8 and Mig1. The range of induction for the 190 genes from Additional file 1 is 2.0 to 21.7.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
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Figure 3: (A) The influence of multiple Hap complex sites on the change in gene expression of the genes in Additional file 1. A Fisher's exact test was performed for genes containing predicted binding sites for the Hap complex and their association with induction or repression. Genes were grouped into either those with one Hap complex binding site or those with two or more according to Additional file 1. (B) The influence of Hap4, Cat8 and Mig1 on the magnitude of induction of the genes in Additional file 1. The average fold-induction of genes in Additional file 1, grouped according to the transcription factor binding sites they contain, was calculated. It was observed that those containing binding sites for each of the Hap complex (where Hap4 is the activator), Cat8 and Mig1, had the highest average fold induction with Mig1 being a dominant factor in high induction. A bar for Cat8 is not included as only one gene (YOR019W, factor change 2.1) has a Cat8 site alone. (C) A Venn diagram of the distribution of binding sites for the Hap complex, Cat8 and Mig1 in the genes in Additional file 1. 88% of the genes in Additional file 1 contain a binding site for one or more of the Hap complex, Cat8 and Mig1. The range of induction for the 190 genes from Additional file 1 is 2.0 to 21.7.
Mentions: Since six of the genes from the HAP4 over-expression study did not contain Hap complex binding sites, we did a further in silico analysis of the induced genes of Additional file 1. We noted that the genes of Table 2, contained motifs that could be sub-divided into 2 distinct families: CCAATG (which we denoted as a Hap4_1 site) and (G/C)CAA(G/T)CAA (a Hap4_2 site). The bold sequences are the conserved sequences previously identified in the HAP4 over-expression strain study [19]. The genes of Additional file 1 were then examined 500 nucleotides upstream and 200 nucleotides downstream from the start codon using WebMOTIF [24] for these two biologically-relevant sites since it has been previously demonstrated that Hap complex binding sites are statistically over-represented up to 400 nucleotides upstream from the start codon [19]: we found that 112/146 (77%) induced genes and 127/190 (67%) of the complete dataset in Additional file 1 had one or more Hap complex binding site (Additional files 1 and 4). Our analysis included presumed binding sites such as CAAATC and CCAAAC which arose from the biologically-verified data on transcription factors (Table 2). For example, AQY2 is identified as Hap complex-dependent, and has two CCAAAC sites, but no CCAATNA sites. There are only two other genes that do not contain CCAATNA sites in Table 2, namely CRC1 and ALD6. In addition it was notable that Hap complex consensus sequences are predominantly found upstream of mitochondrial genes with 84/127 (66%) of such genes having at least a partial mitochondrial location. Furthermore, of the induced genes with a known cellular location and at least two Hap complex binding sites, 69/80 (86%) are localized to the mitochondria. It appears, then, that Hap4 activation is preferentially directed at mitochondrially-related functions especially within the TCA-cycle and respiratory chain, but cytosolic support via the glyoxylate cycle is also potentially regulated by Hap4 activation (Additional file 1). It was further noted that multiple Hap complex binding sites are associated with induction (Fig. 3A). A Fisher's exact test showed that there was a statistically-significant association between the change in gene expression and the number of Hap complex binding sites in the gene (p = 0.00). This supported the fact that genes with two or more Hap complex binding sites are likely to be induced, whereas those with one site may be either induced or repressed.

Bottom Line: In addition, 13% were found to have a binding site for Cat8 and 21% had a binding site for Mig1.Our dataset gives a remarkably complete view of the involvement of genes in the TCA cycle, glyoxylate cycle and respiratory chain in the expression of the phenotype of V5.TM6*P.Furthermore, 88% of the transcriptional response of the induced genes in our dataset can be related to the potential activities of just three proteins: Hap4, Cat8 and Mig1.

View Article: PubMed Central - HTML - PubMed

Affiliation: School of Life and Health Sciences, Aston University, Aston Triangle, Birmingham, B4 7ET, UK. n.bonander@aston.ac.uk

ABSTRACT

Background: We previously described the first respiratory Saccharomyces cerevisiae strain, KOY.TM6*P, by integrating the gene encoding a chimeric hexose transporter, Tm6*, into the genome of an hxt yeast. Subsequently we transferred this respiratory phenotype in the presence of up to 50 g/L glucose to a yeast strain, V5 hxt1-7Delta, in which only HXT1-7 had been deleted. In this study, we compared the transcriptome of the resultant strain, V5.TM6*P, with that of its wild-type parent, V5, at different glucose concentrations.

Results: cDNA array analyses revealed that alterations in gene expression that occur when transitioning from a respiro-fermentative (V5) to a respiratory (V5.TM6*P) strain, are very similar to those in cells undergoing a diauxic shift. We also undertook an analysis of transcription factor binding sites in our dataset by examining previously-published biological data for Hap4 (in complex with Hap2, 3, 5), Cat8 and Mig1, and used this in combination with verified binding consensus sequences to identify genes likely to be regulated by one or more of these. Of the induced genes in our dataset, 77% had binding sites for the Hap complex, with 72% having at least two. In addition, 13% were found to have a binding site for Cat8 and 21% had a binding site for Mig1. Unexpectedly, both the up- and down-regulation of many of the genes in our dataset had a clear glucose dependence in the parent V5 strain that was not present in V5.TM6*P. This indicates that the relief of glucose repression is already operable at much higher glucose concentrations than is widely accepted and suggests that glucose sensing might occur inside the cell.

Conclusion: Our dataset gives a remarkably complete view of the involvement of genes in the TCA cycle, glyoxylate cycle and respiratory chain in the expression of the phenotype of V5.TM6*P. Furthermore, 88% of the transcriptional response of the induced genes in our dataset can be related to the potential activities of just three proteins: Hap4, Cat8 and Mig1. Overall, our data support genetic remodelling in V5.TM6*P consistent with a respiratory metabolism which is insensitive to external glucose concentrations.

Show MeSH
Related in: MedlinePlus