Quantitative site-specific phosphoproteomics of Trichoderma reesei signaling pathways upon induction of hydrolytic enzyme production

Elizabeth V. Nguyen, Susumu Y. Imanishi, Pekka Haapaniemi, Avinash Yadav, Markku Saloheimo, Garry L. Corthals (Corresponding Author), Tiina M. Pakula (Corresponding Author)

Research output: Contribution to journalArticleScientificpeer-review

8 Citations (Scopus)

Abstract

The filamentous fungus Trichoderma reesei is used for industrial production of secreted enzymes including carbohydrate active enzymes, such as cellulases and hemicellulases. The production of many of these enzymes by T. reesei is influenced by the carbon source it grows on, where the regulation system controlling hydrolase genes involves various signaling pathways. T. reesei was cultivated in the presence of sorbitol, a carbon source that does not induce the production of cellulases and hemicellulases, and then exposed to either sophorose or spent-grain extract, which are efficient inducers of the enzyme production. Specific changes at phosphorylation sites were investigated in relation to the production of cellulases and hemicellulases using an MS-based framework. Proteome-wide phosphorylation following carbon source exchange was investigated in the early stages of induction: 0, 2, 5, and 10 min. The workflow involved sequential trypsin digestion, TiO2 enrichment, and MS analysis using a Q Exactive mass spectrometer. We report on the identification and quantitation of 1721 phosphorylation sites. Investigation of the data revealed a complex signaling network activated upon induction involving components related to light-mediated cellulase induction, osmoregulation, and carbon sensing. Changes in protein phosphorylation were detected in the glycolytic pathway, suggesting an inhibition of glucose catabolism at 10 min after the addition of sophorose and as early as 2 min after the addition of spent-grain extract. Differential phosphorylation of factors related to carbon storage, intracellular trafficking, cytoskeleton, and cellulase gene regulation were also observed.
Original languageEnglish
Pages (from-to)457-467
JournalJournal of Proteome Research
Volume15
Issue number2
DOIs
Publication statusPublished - 2016
MoE publication typeA1 Journal article-refereed

Fingerprint

Phosphorylation
Enzyme Induction
Trichoderma
Carbon
Cellulases
Enzymes
Cellulase
Osmoregulation
Sorbitol
Workflow
Mass spectrometers
Hydrolases
Proteome
Fungi
Cytoskeleton
Gene expression
Trypsin
Genes
Digestion
Carbohydrates

Keywords

  • cellulase
  • hemicellulase
  • mass spectrometry
  • MS/MS
  • phosphoenrichment
  • phosphoproteomics
  • post-translational modification
  • TiO2
  • Trichoderma reesei

Cite this

Nguyen, Elizabeth V. ; Imanishi, Susumu Y. ; Haapaniemi, Pekka ; Yadav, Avinash ; Saloheimo, Markku ; Corthals, Garry L. ; Pakula, Tiina M. / Quantitative site-specific phosphoproteomics of Trichoderma reesei signaling pathways upon induction of hydrolytic enzyme production. In: Journal of Proteome Research. 2016 ; Vol. 15, No. 2. pp. 457-467.
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abstract = "The filamentous fungus Trichoderma reesei is used for industrial production of secreted enzymes including carbohydrate active enzymes, such as cellulases and hemicellulases. The production of many of these enzymes by T. reesei is influenced by the carbon source it grows on, where the regulation system controlling hydrolase genes involves various signaling pathways. T. reesei was cultivated in the presence of sorbitol, a carbon source that does not induce the production of cellulases and hemicellulases, and then exposed to either sophorose or spent-grain extract, which are efficient inducers of the enzyme production. Specific changes at phosphorylation sites were investigated in relation to the production of cellulases and hemicellulases using an MS-based framework. Proteome-wide phosphorylation following carbon source exchange was investigated in the early stages of induction: 0, 2, 5, and 10 min. The workflow involved sequential trypsin digestion, TiO2 enrichment, and MS analysis using a Q Exactive mass spectrometer. We report on the identification and quantitation of 1721 phosphorylation sites. Investigation of the data revealed a complex signaling network activated upon induction involving components related to light-mediated cellulase induction, osmoregulation, and carbon sensing. Changes in protein phosphorylation were detected in the glycolytic pathway, suggesting an inhibition of glucose catabolism at 10 min after the addition of sophorose and as early as 2 min after the addition of spent-grain extract. Differential phosphorylation of factors related to carbon storage, intracellular trafficking, cytoskeleton, and cellulase gene regulation were also observed.",
keywords = "cellulase, hemicellulase, mass spectrometry, MS/MS, phosphoenrichment, phosphoproteomics, post-translational modification, TiO2, Trichoderma reesei",
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year = "2016",
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Quantitative site-specific phosphoproteomics of Trichoderma reesei signaling pathways upon induction of hydrolytic enzyme production. / Nguyen, Elizabeth V.; Imanishi, Susumu Y.; Haapaniemi, Pekka; Yadav, Avinash; Saloheimo, Markku; Corthals, Garry L. (Corresponding Author); Pakula, Tiina M. (Corresponding Author).

In: Journal of Proteome Research, Vol. 15, No. 2, 2016, p. 457-467.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - Quantitative site-specific phosphoproteomics of Trichoderma reesei signaling pathways upon induction of hydrolytic enzyme production

AU - Nguyen, Elizabeth V.

AU - Imanishi, Susumu Y.

AU - Haapaniemi, Pekka

AU - Yadav, Avinash

AU - Saloheimo, Markku

AU - Corthals, Garry L.

AU - Pakula, Tiina M.

PY - 2016

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N2 - The filamentous fungus Trichoderma reesei is used for industrial production of secreted enzymes including carbohydrate active enzymes, such as cellulases and hemicellulases. The production of many of these enzymes by T. reesei is influenced by the carbon source it grows on, where the regulation system controlling hydrolase genes involves various signaling pathways. T. reesei was cultivated in the presence of sorbitol, a carbon source that does not induce the production of cellulases and hemicellulases, and then exposed to either sophorose or spent-grain extract, which are efficient inducers of the enzyme production. Specific changes at phosphorylation sites were investigated in relation to the production of cellulases and hemicellulases using an MS-based framework. Proteome-wide phosphorylation following carbon source exchange was investigated in the early stages of induction: 0, 2, 5, and 10 min. The workflow involved sequential trypsin digestion, TiO2 enrichment, and MS analysis using a Q Exactive mass spectrometer. We report on the identification and quantitation of 1721 phosphorylation sites. Investigation of the data revealed a complex signaling network activated upon induction involving components related to light-mediated cellulase induction, osmoregulation, and carbon sensing. Changes in protein phosphorylation were detected in the glycolytic pathway, suggesting an inhibition of glucose catabolism at 10 min after the addition of sophorose and as early as 2 min after the addition of spent-grain extract. Differential phosphorylation of factors related to carbon storage, intracellular trafficking, cytoskeleton, and cellulase gene regulation were also observed.

AB - The filamentous fungus Trichoderma reesei is used for industrial production of secreted enzymes including carbohydrate active enzymes, such as cellulases and hemicellulases. The production of many of these enzymes by T. reesei is influenced by the carbon source it grows on, where the regulation system controlling hydrolase genes involves various signaling pathways. T. reesei was cultivated in the presence of sorbitol, a carbon source that does not induce the production of cellulases and hemicellulases, and then exposed to either sophorose or spent-grain extract, which are efficient inducers of the enzyme production. Specific changes at phosphorylation sites were investigated in relation to the production of cellulases and hemicellulases using an MS-based framework. Proteome-wide phosphorylation following carbon source exchange was investigated in the early stages of induction: 0, 2, 5, and 10 min. The workflow involved sequential trypsin digestion, TiO2 enrichment, and MS analysis using a Q Exactive mass spectrometer. We report on the identification and quantitation of 1721 phosphorylation sites. Investigation of the data revealed a complex signaling network activated upon induction involving components related to light-mediated cellulase induction, osmoregulation, and carbon sensing. Changes in protein phosphorylation were detected in the glycolytic pathway, suggesting an inhibition of glucose catabolism at 10 min after the addition of sophorose and as early as 2 min after the addition of spent-grain extract. Differential phosphorylation of factors related to carbon storage, intracellular trafficking, cytoskeleton, and cellulase gene regulation were also observed.

KW - cellulase

KW - hemicellulase

KW - mass spectrometry

KW - MS/MS

KW - phosphoenrichment

KW - phosphoproteomics

KW - post-translational modification

KW - TiO2

KW - Trichoderma reesei

U2 - 10.1021/acs.jproteome.5b00796

DO - 10.1021/acs.jproteome.5b00796

M3 - Article

VL - 15

SP - 457

EP - 467

JO - Journal of Proteome Research

JF - Journal of Proteome Research

SN - 1535-3893

IS - 2

ER -