Deletion of the small GTPase rac1 in Trichoderma reesei provokes hyperbranching and impacts growth and cellulase production

Springer Science and Business Media LLC - 2019
Elisabeth Fitz1, Christian Gamauf2, Bernhard Schink1, Franziska Wanka3
1Research Division Biochemical Technology, Institute of Chemical, Environmental & Bioscience Engineering, TU Wien, Vienna, Austria
2Group Biotechnology, Clariant Produkte (Deutschland) GmbH, Planegg, Germany
3Austrian Centre of Industrial Biotechnology (ACIB) GmbH c/o Research Division Biochemical Technology, Institute of Chemical, Environmental & Bioscience Engineering, TU Wien, Vienna, Austria

Tóm tắt

AbstractBackgroundTrichoderma reeseiis widely known for its enormous protein secretion capacity and as an industrially relevant producer of cellulases and hemicellulases. Over the last decades, rational strain engineering was applied to further enhance homologous and heterologous enzyme yields. The introduction of hyperbranching is believed to increase protein secretion, since most exocytosis is located at the hyphal apical tip. There are several genetic modifications which can cause hyperbranching, for example the deletion of the small Rho GTPaserac. Rac plays a crucial role in actin dynamics and is involved in polarisation of the cell during germination and apical extension of the hyphae.ResultsWe deletedrac1in aT. reeseistrain with an ectopically overexpressed endoglucanase, CEL12A, under Pcdna1control. This deletion provoked a hyperbranching phenotype and strong apolar growth during germination and in mature hyphae. The strains displayed dichotomous branching and shorter total mycelium length with a larger hyphal diameter. Δrac1strains exhibited a decreased radial growth on solid media. Biomass formation in liquid cultures was carbon source dependent; similar to the reference strain during growth on lactose, increased ond-glucose and slightly enhanced on cellulose. While extracellular cellulase activities remained at parental strain levels ond-glucose and cellulose, the specific activity on lactose cultures was increased up to three times at 72 h accompanied by an upregulation of transcription of the main cellulases. Although the morphology of the Δrac1strains was considerably altered, the viscosity of the culture broth in fed-batch cultivations were not significantly different in comparison to the parental strain.ConclusionsDeletion of the small Rho GTPaserac1changes the morphology of the hyphae and provokes hyperbranching without affecting viscosity, independent of the carbon source. In contrast, biomass formation and cellulase production are altered in a carbon source dependent manner in the Δrac1strains.

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