1 research outputs found
Quantitative Site-Specific Phosphoproteomics of <i>Trichoderma reesei</i> Signaling Pathways upon Induction of Hydrolytic Enzyme Production
The
filamentous fungus <i>Trichoderma reesei</i> 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 <i>T. reesei</i> is influenced
by the carbon source it grows on, where the regulation system controlling
hydrolase genes involves various signaling pathways. <i>T. reesei</i> 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, TiO<sub>2</sub> 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