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Gene expression of Grx4 regulated strains in the Ustilago maydis FB2 background

ABSTRACT: Gene expression of FB2 wt in minimal medium with 2 percent arabinose or glucose after 24h of growth was compared with the conditional promoter regulated strain Pcrg1:grx4 (#55). Glucose is a repressor of this promoter while arabinose acts as inducer. Grx4 was initially pre-depleted during growth on MM with glucose over night. Afterwards cells were transferred to MM with either glucose or arabinose and cells were collected after 24h. 3 biological repeats were performed for each strain and each condition. Cells were flash frozen and used to isolate total RNA. RNA sequencing was done by Genewiz. DEG’s between wt and mutant for the different conditions were obtained. Iron regulated genes such as the high affinity uptake system and the siderophore system of Ustilago maydis were repressed in the Pcrg1:grx4 (#55) strain when grown in glucose. Expression of these genes were similar for wt and the conditional strain when grown in arabinose. Grx4 was identified as an essential gene in Ustilago maydis which regulates the iron regulon and related genes.

ORGANISM(S): Ustilago maydis 521

PROVIDER: GSE239366 | GEO | 2023/12/08

REPOSITORIES: GEO

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Project description:L-Arabinose occurs at economically relevant levels in lignocellulosic hydrolysates. Especially at low concentrations of L-arabinose, its uptake via the Gal2 galactose transporter is an important rate-controlling step in the complete conversion of these feedstocks by engineered, pentose-metabolizing Saccharomyces cerevisiae strains. Chemostat-based transcriptome analysis yielded 16 putative sugar transporter genes in the filamentous fungus Penicillium chrysogenum whose transcript levels were at least three-fold higher in L-arabinose-limited cultures than in glucose-limited and ethanol-limited cultures. Of five genes that showed an over 30-fold higher transcript level in arabinose-grown cultures, only one (Pc20g01790) restored growth on L-arabinose upon expression in an engineered L-arabinose-fermenting S. cerevisiae strain in which GAL2 had been deleted. Sugar-transport assays indicated that Pc20g01790this transporter, designated as PcAraT, encodes functions as a high-affinity (Km = 0.13 mM) L-arabinose-proton symporter that does not transport xylose or glucose. An L-arabinose-metabolizing S. cerevisiae strain co-expressing Pc20g01790PcAraT and GAL2 showed lower residual substrate concentrations in L-arabinose-limited chemostat cultures (4 mg L-1) than a congenic strain in which L-arabinose import exclusively depended on Gal2 (1.8 g L-1). Inhibition of L-arabinose transport by these sugars was less pronounced than observed with Gal2. A hexose-phosphorylation-deficient, L-arabinose-metabolizing S. cerevisiae strain expressing PcAraT Pc20g0190 grew on 20 g L-1 L-arabinose in the presence of 20 g L-1 glucose, which completely inhibited growth on L-arabinose of a congenic strain dependent on L-arabinose transport via Gal2. Its high affinity and specificity for L-L-arabinose, combined with limited sensitivity to inhibition by glucose and D-D-xylose make PcAraT/ Pc20g01790 a valuable transporter gene for application in metabolic engineering strategies aimed at engineering S. cerevisiae strains for efficient conversion of lignocellulosic hydrolysates.

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Gene expression of Grx4 regulated strains in the Ustilago maydis FB2 background

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