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Uncovering transcriptional regulation of glycerol metabolism in Aspergilli through genome-wide gene expression data analysis

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Abstract

Glycerol is catabolized by a wide range of microorganisms including Aspergillus species. To identify the transcriptional regulation of glycerol metabolism in Aspergillus, we analyzed data from triplicate batch fermentations of three different Aspergilli (Aspergillus nidulans, Aspergillus oryzae and Aspergillus niger) with glucose and glycerol as carbon sources. Protein comparisons and cross-analysis with gene expression data of all three species resulted in the identification of 88 genes having a conserved response across the three Aspergilli. A promoter analysis of the up-regulated genes led to the identification of a conserved binding site for a putative regulator to be 5′-TGCGGGGA-3′, a binding site that is similar to the binding site for Adr1 in yeast and humans. We show that this Adr1 consensus binding sequence was over-represented on promoter regions of several genes in A. nidulans, A. oryzae and A. niger. Our transcriptome analysis indicated that genes involved in ethanol, glycerol, fatty acid, amino acids and formate utilization are putatively regulated by Adr1 in Aspergilli as in Saccharomyces cerevisiae and this transcription factor therefore is likely to be cross-species conserved among Saccharomyces and distant Ascomycetes. Transcriptome data were further used to evaluate the high osmolarity glycerol pathway. All the components of this pathway present in yeast have orthologues in the three Aspergilli studied and its gene expression response suggested that this pathway functions as in S. cerevisiae. Our study clearly demonstrates that cross-species evolutionary comparisons among filamentous fungi, using comparative genomics and transcriptomics, are a powerful tool for uncovering regulatory systems.

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Acknowledgments

The authors would like to thank Tina Johansen, Pia Friis and Lene Christiansen at Technical University of Denmark for assistance with the experimental work and Dr. Kim Hansen for supervising the fermentations with A. oryzae, Lone Vuholm and Anne Kejser Jensen at Novozymes for technical assistance. We would like to thank National Council of Research Conacyt-Mexico and Chalmers University of Technology for financial support to MS; Novozymes Bioprocess Academy and Chalmers University of Technology for financial support to WV; Danish Research Council for Technology and Production Sciences and Lundbeck Foundation for financial support to GP and Danish Research Agency for Technology and Production for financial support to MRA. We thank Dr. Gerald Hofmann for revision of the manuscript and good scientific discussion.

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Authors and Affiliations

  1. Department of Chemical and Biological Engineering, Chalmers University of Technology, 412 96, Göteborg, Sweden

    Margarita Salazar, Wanwipa Vongsangnak & Jens Nielsen

  2. Department of Systems Biology, Center for Microbial Biotechnology, Technical University of Denmark, 2800, Kongens Lyngby, Denmark

    Gianni Panagiotou & Mikael R. Andersen

Authors
  1. Margarita Salazar
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  2. Wanwipa Vongsangnak
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  3. Gianni Panagiotou
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  4. Mikael R. Andersen
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  5. Jens Nielsen
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Corresponding author

Correspondence to Jens Nielsen.

Additional information

Communicated by S. Hohmann.

M. Salazar and W. Vongsangnak contributed equally.

Electronic supplementary material

Below is the link to the electronic supplementary material.

438_2009_486_MOESM1_ESM.pdf

A. nidulans differentially expressed genes mapped to metabolic map of A. oryzae resulting from glucose versus glycerol t-test analysis (PDF 3.54 MB)

438_2009_486_MOESM2_ESM.pdf

A. oryzae differentially expressed genes mapped to metabolic map of A. oryzae resulting from glucose versus glycerol t-test analysis (PDF 3.54 MB)

438_2009_486_MOESM3_ESM.pdf

A. niger differentially expressed genes mapped to metabolic map of A. oryzae resulting from glucose versus glycerol t-test analysis (PDF 3.54 MB)

438_2009_486_MOESM4_ESM.pdf

Significant genes differentially expressed and mapped to the metabolic maps of A. niger and A. oryzae. Selected pathways included: central carbon metabolism, TCA cycle, C2 and C3 carbon metabolism and fatty acid metabolism. Complete metabolic maps of A. nidulans, A. oryzae and A. niger, using A. oryzae as a template are included in Supplementary Figs 1, 2 and 3, respectively. The abbreviation of metabolites is described as follows. C2 metabolism: ETH, ethanol; AC, acetate, ACAL, acetaldehyde; ACCOA, acetyl-CoA. C3 metabolism: GL, glycerol; GLYAL, D-glyceraldehyde; GLYN, glycerone; GL3P, sn-glycerol 3-phosphate; T3P2, glycerone phosphate. Pyruvate metabolism: F6P, Beta-D-fructose 6-phosphate; FDP, Beta-D-fructose 1,6-bisphosphate; T3P1, D-glyceraldehyde 3-phosphate; 13PDG, 1,3-Bisphospho-D-glycerate; 3PG, 3-Phospho-D-glycerate 2PG, 2-Phospho-D-glycerate; PEP, phosphoenolpyruvate; PYR, pyruvate; MTHGXL, methylglyoxal; RGT, glutathione; LACAL, D-lactaldehyde; LAC, D-lactate; LGT, (R)-S-lactoylglutathione; LLAC, L-lactate. TCA cycle: OA, oxaloacetate; CIT, citrate; ACO, Cis-aconitate; ICIT, isocitrate AKG, 2-oxoglutarate; SUCCOA, succinyl coenzyme A; SUCC, succinate; FUM, fumarate; MAL, (S)-malate; GABAL, 4-aminobutyraldehyde; GABA, 4-aminobutanoate; GLU, L-glutamate; SUCCSAL, succinate semialdehyde. Fatty acid catabolism: C120COA, dodecanoyl-Coenzyme A; C120CAR, dodecanoyl-carnitine; C12DCOA, dodecanoyl-dehydro-Coenzyme A; C12HCOA, dodecanoyl-Hydroxy-Coenzyme A; C12OCOA, dodecanoyl-oxo-Coenzyme A; C140COA, myristoyl-Coenzyme A; C140CAR, myristoyl-carnitine; C14DCOA, myristoyl-dehydro-Coenzyme A; C14HCOA, myristoyl-Hydroxy-Coenzyme A; C14OCOA, myristoyl-oxo-Coenzyme A; C160COA, hexadecanoyl-Coenzyme A; C160CAR, hexadecanoyl-carnitine; C16DCOA, hexadecanoyl-dehydro-Coenzyme A; C16HCOA, hexadecanoyl-Hydroxy-Coenzyme A; C160COA, hexadecanoyl-Coenzyme A; C180COA, stearoyl-Coenzyme A; C180CAR, octadecanoyl-carnitine; C18DCOA, stearoyl-dehydro-Coenzyme A; C18HCOA, stearoyl-Hydroxy-Coenzyme A; C180COA, Stearoyl-oxo-Coenzyme A. Extracellular metabolites are designated by subscript ‘e’; mitochondrial metabolites by subscript ‘m’ (PDF 90.2 KB)

Supplementary Table 1. (PDF 204 KB)

Supplementary Table 2. (XLS 50 KB)

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Supplementary Table 5. (PDF 24.6 KB)

Supplementary Table 6. (XLS 34.0 KB)

Supplementary Table 7. (XLS 274 KB)

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Salazar, M., Vongsangnak, W., Panagiotou, G. et al. Uncovering transcriptional regulation of glycerol metabolism in Aspergilli through genome-wide gene expression data analysis. Mol Genet Genomics 282, 571–586 (2009). https://doi.org/10.1007/s00438-009-0486-y

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