title: Genome wide analysis of protein production load in Trichoderma reesei.
abstract: BACKGROUND
  The filamentous fungus Trichoderma reesei (teleomorph Hypocrea jecorina) is a widely used industrial host organism for protein production. In industrial cultivations, it can produce over 100 g/l of extracellular protein, mostly constituting of cellulases and hemicellulases. In order to improve protein production of T. reesei the transcriptional regulation of cellulases and secretory pathway factors have been extensively studied. However, the metabolism of T. reesei under protein production conditions has not received much attention.
  
  
  RESULTS
  To understand the physiology and metabolism of T. reesei under protein production conditions we carried out a well-controlled bioreactor experiment with extensive analysis. We used minimal media to make the data amenable for modelling and three strain pairs to cover different protein production levels. With RNA-sequencing transcriptomics we detected the concentration of the carbon source as the most important determinant of the transcriptome. As the major transcriptional response concomitant to protein production we detected the induction of selected genes that were putatively regulated by xyr1 and were related to protein transport, amino acid metabolism and transcriptional regulation. We found novel metabolic responses such as production of glycerol and a cellotriose-like compound. We then used this cultivation data for flux balance analysis of T. reesei metabolism and demonstrate for the first time the use of genome wide stoichiometric metabolic modelling for T. reesei. We show that our model can predict protein production rate and provides novel insight into the metabolism of protein production. We also provide this unprecedented cultivation and transcriptomics data set for future modelling efforts.
  
  
  CONCLUSIONS
  The use of stoichiometric modelling can open a novel path for the improvement of protein production in T. reesei. Based on this we propose sulphur assimilation as a major limiting factor of protein production. As an organism with exceptional protein production capabilities modelling of T. reesei can provide novel insight also to other less productive organisms.
authors.affiliation: VTT Technical Research Centre of Finland, Tietotie 2, P.O. Box FI-1000, 02044 Espoo, Finland.
authors.forename: Tiina M
authors.initials: TM
authors.lastname: Pakula
authors.affiliation: VTT Technical Research Centre of Finland, Tietotie 2, P.O. Box FI-1000, 02044 Espoo, Finland.
authors.forename: Heli
authors.initials: H
authors.lastname: Nygren
authors.affiliation: VTT Technical Research Centre of Finland, Tietotie 2, P.O. Box FI-1000, 02044 Espoo, Finland.
authors.forename: Dorothee
authors.initials: D
authors.lastname: Barth
authors.affiliation: Department of Information and Computer Science, Aalto University, PO Box 15400, 00076 Espoo, Finland ; Helsinki Institute for Information Technology HIIT, Espoo, Finland.
authors.forename: Markus
authors.initials: M
authors.lastname: Heinonen
authors.affiliation: VTT Technical Research Centre of Finland, Tietotie 2, P.O. Box FI-1000, 02044 Espoo, Finland.
authors.forename: Sandra
authors.initials: S
authors.lastname: Castillo
authors.affiliation: VTT Technical Research Centre of Finland, Tietotie 2, P.O. Box FI-1000, 02044 Espoo, Finland.
authors.forename: Merja
authors.initials: M
authors.lastname: Penttilä
authors.affiliation: VTT Technical Research Centre of Finland, Tietotie 2, P.O. Box FI-1000, 02044 Espoo, Finland.
authors.forename: Mikko
authors.identifier: 0000-0002-6902-8488
authors.initials: M
authors.lastname: Arvas
keywords: Flux balance analysis; Hypocrea jecorina; Metabolic modelling; Protein production; RNA sequencing; Stoichiometric model; Transcriptomics; Trichoderma reesei