The development of a heterologous transformation system for the cellulolytic fungus Trichoderma reesei based on a pyrG-negative mutant strain

Background:There is an oxidoreductive D-galactose pathway in filamentous fungi. Results:We identified an L-xylo-3-hexulose reductase that produces D-sorbitol and that is part of this pathway. Conclusion: This L-xylo-3-hexulose reductase is the missing link in the oxidoreductive D-galactose pathway. Significance: The alternative pathway for D-galactose catabolism in filamentous fungi is elucidated.

Section: Methodsmentioning

confidence: 99%

l-xylo-3-Hexulose Reductase Is the Missing Link in the Oxidoreductive Pathway for d-Galactose Catabolism in Filamentous Fungi

Mojžita

Herold

Metz

et al. 2012

“…Southern Blot Analysis-Fungal genomic DNA was isolated as described elsewhere (32) and Southern hybridization was carried out as described by Sambrook et al (29).…”

Section: Methodsmentioning

confidence: 99%

Phosphorylation Positively Regulates DNA Binding of the Carbon Catabolite Repressor Cre1 of Hypocrea jecorina(Trichoderma reesei)

Cziferszky¹,

Mach²,

Kubicek

2002

115

Cre1 of the ascomyceteCarbon catabolite repression is a means by which cells manage priority use of easy and fast metabolizable carbon sources over more complex ones. In Saccharomyces cerevisiae, mutations in MIG1 or in respective binding sites in Mig1 regulated promoters partially relieve the yeast from carbon catabolite repression (1-4). To form an active repressor complex, Mig1 requires the co-repressors Ssn6 (Cyc8)-Tup1 (5-8). Mig1 function is regulated by glucose-dependent nuclear import/export, Mig1 being nuclear when glucose is present and being transported to the cytoplasm when glucose becomes limiting (9). Concomitantly, Mig1 has been shown to be phosphorylated under derepressing conditions (10, 11). Present genetic evidence suggests that both of these events are mediated by the Snf1 kinase complex: a mutation in MIG1 suppresses the snf1 mutant defects in SUC2 and GAL1 expression (3, 4), indicating that SNF1 negatively regulates repression by Mig1. Furthermore, Mig1 is permanently localized in the nucleus in snf1

“…The primers Env1T101I5F and Env1T101I3R contain overlapping sequences for combination with the amdS marker cassette (63). The mutation was confirmed by sequencing the vector, and the amplified cassette was used for protoplast transformation of QM6a⌬env1 (51).…”

Section: Construction Of Env1-t101i Complementation Strains In T Reementioning

confidence: 99%

A Native Threonine Coordinates Ordered Water to Tune Light-Oxygen-Voltage (LOV) Domain Photocycle Kinetics and Osmotic Stress Signaling in Trichoderma reesei ENVOY

Lokhandwala

Vega

Hopkins

et al. 2016

Light-oxygen-voltage (LOV)3 domain-containing photoreceptors are widely distributed in nature, where they couple blue light absorption to regulation of a diverse array of signal transduction pathways (1). In general, LOV proteins can be divided into two subclasses: 1) short LOV proteins (sLOV) that exist as the isolated LOV domain with short ancillary N-or C-terminal caps (2-4) and 2) modular LOV proteins that couple blue light activation to allosteric regulation of effector domains (1). Although the modular LOV proteins are present in plants, bacteria, and all fungi, the sLOV variety are predominantly found in bacteria and only some fungi (4). Currently, LOV proteins have received widespread attention because of their important roles in regulation of circadian function (3, 5, 6), growth and development (7,8), stress responses (9 -11), and adaptation to and regulation of pathogenicity (12). In addition, their wide ranging utility has led to the development of optogenetic tools that harness their modular design (13,14). Despite substantial research, key questions involving LOV photocycles remain.LOV domain chemistry is characterized by blue light-induced formation of a covalent adduct between a bound flavin cofactor (FMN, FAD, or riboflavin) and a conserved Cys residue in a GXNCRFLQ motif. Concomitant with adduct formation is protonation of the N5 position of the isoalloxazine ring. Current models indicate that signal transduction is coupled to N5 protonation via allosteric regulation of N-or C-terminal effector elements remote from the flavin active site (3,15,16). The covalent adduct is defined by a broad UV-visible absorption band centered at ϳ390 nm (LOV 390 ). Upon return to the dark, the adduct state spontaneously decays to an oxidized flavin (LOV 450 ) on a timescale of seconds to days (17,18). Currently the biological role of the wide range in photocycle lifetimes is unknown; however, several studies have suggested that the range facilitates adaptation to changing levels of light intensity (17,19,20). For these reasons, chemical tuning of the LOV photocycle lifetime through understanding of the adduct decay mechanism has been attempted in several systems (2,17,18,(21)(22)(23)(24).Several lines of reasoning have led to a general mechanism of adduct decay. First, solvent isotope effect experiments indicate that a single proton abstraction event is rate-limiting (17, 18). Second, adduct decay can be catalyzed by the presence of small molecule bases such as imidazole (25). Third, residue substitutions at regions that regulate solvent access to the flavin active site have a substantial effect on LOV photocycle lifetimes (18,26). Combined, these experiments implicate N5 deprotonation as the rate-determining step in adduct decay. Consistent with such a model, mutation of residues that regulate accessibility of small molecules to the N5 position or that tune hydrogen bonding characteristics affect kinetics of LOV proteins (17,18,21,22,24,26,27). Importantly, the natural base responsible for N5 deprotonation remai...