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Second-generation (2G) bioethanol production, derived from lignocellulosic biomass, has emerged as a sustainable alternative to fossil fuels by addressing growing energy demands and environmental concerns. Fungal sugar transporters (STs) play a critical role in this process, enabling the uptake of monosaccharides such as glucose and xylose, which are released during the enzymatic hydrolysis of biomass. This mini-review explores recent advances in the structural and functional characterization of STs in filamentous fungi and yeasts, highlighting their roles in processes such as cellulase induction, carbon catabolite repression, and sugar signaling pathways. The review also emphasizes the potential of genetic engineering to enhance the specificity and efficiency of these transporters, overcoming challenges such as substrate competition and limited pentose metabolism in industrial strains. By integrating the latest research findings, this work underscores the pivotal role of fungal STs in optimizing lignocellulosic bioethanol production and advancing the bioeconomy. Future prospects for engineering transport systems and their implications for industrial biotechnology are also discussed. Key points STs present a conserved structure with different sugar affinities STs are involved in the signaling and transport of sugars derived from plant biomass Genetic engineering of STs can improve 2G bioethanol production

Background In hematologic cancers, including leukemia, cells depend on amino acids for rapid growth. Anti-metabolites that prevent their synthesis or promote their degradation are considered potential cancer treatment agents. Amino acid deprivation triggers proliferation inhibition, autophagy, and programmed cell death. l -lysine, an essential amino acid, is required for tumor growth and has been investigated for its potential as a target for cancer treatment. l -lysine α-oxidase, a flavoenzyme that degrades l -lysine, has been studied for its ability to induce apoptosis and prevent cancer cell proliferation. In this study, we describe the use of l -lysine α-oxidase (LO) from the filamentous fungus Trichoderma harzianum for cancer treatment. Results The study identified and characterized a novel LO from T. harzianum and demonstrated that the recombinant protein (rLO) has potent and selective cytotoxic effects on leukemic cells by triggering the apoptotic cascade through mitochondrial dysfunction. Conclusions The results support future translational studies using the recombinant LO as a potential drug for the treatment of leukemia.

As we conclude this Special Issue on fungal biology and interactions, it is only appropriate to reflect on the remarkable progress our scientific community has made in unraveling the mysteries of the fungal kingdom [...]

Background Trichoderma reesei is an organism extensively used in the bioethanol industry, owing to its capability to produce enzymes capable of breaking down holocellulose into simple sugars. The uptake of carbohydrates generated from cellulose breakdown is crucial to induce the signaling cascade that triggers cellulase production. However, the sugar transporters involved in this process in T. reesei remain poorly identified and characterized. Results To address this gap, this study used temporal membrane proteomics analysis to identify five known and nine putative sugar transporters that may be involved in cellulose degradation by T. reesei . Docking analysis pointed out potential ligands for the putative sugar transporter Tr44175. Further functional validation of this transporter was carried out in Saccharomyces cerevisiae . The results showed that Tr44175 transports a variety of sugar molecules, including cellobiose, cellotriose, cellotetraose, and sophorose. Conclusion This study has unveiled a transporter Tr44175 capable of transporting cellobiose, cellotriose, cellotetraose, and sophorose. Our study represents the first inventory of T. reesei sugar transportome once exposed to cellulose, offering promising potential targets for strain engineering in the context of bioethanol production.

Several spp. are well known for their ability to: (i) act as important biocontrol agents against phytopathogenic fungi; (ii) function as biofertilizers; (iii) increase the tolerance of plants to biotic and abiotic stresses; and (iv) induce plant defense responses via the production and secretion of elicitor molecules. In this study, we analyzed the gene-regulation effects of Epl-1 protein during the interactions of mutant Δ or wild-type strains with: (a) the phytopathogen and (b) with tomato plants, on short (24 h hydroponic cultures) and long periods (4-weeks old plants) after inoculation. Our results indicate that Epl-1 protein affects the expression of virulence genes, especially those involved in the botrydial biosynthesis ( genes), during the mycoparasitism interaction. The tomato defense-related genes were also affected, indicating that Epl-1 is involved in the elicitation of the salicylic acid pathway. Moreover, Epl-1 also regulates the priming effect in host tomato plants and contributes to enhance the interaction with the host tomato plant during the early stage of root colonization.

Thefilamentous fungus Trichoderma reesei is renowned for its exceptional ability to secrete cellulolytic enzymes, which play a crucial role in the hydrolysis of lignocellulose biomass. The expression of fungal cellulases is meticulously regulated at the transcriptional level, depending on the carbon source available in the medium. To obtain new insights into the transcriptional network controlling cellulase expression in T. reesei, we analyzed RNA-Seq data and identified and characterized a new transcription factor, Rme1, that regulates the expression of cellulolytic genes. Combining functional genomics and protein-DNA interaction assay, we showed that Rme1 acts as a repressor of cellulase production in T. reesei by directly regulating two critical genes involved in cellulose degradation: the cellobiohydrolase cel7a and the carbon catabolite repressor cre1. This is the first report of a transcription factor regulating Cre1. This study contributes to a better understanding of the complex regulation of the cellulolytic system of T. reesei and may be useful for the genetic modification of strains for the biorefinery industry.

The fungus Trichoderma reesei is an essential producer of enzymes that degrade lignocellulosic biomass to produce value-added bioproducts. The cellulolytic system of T. reesei is controlled by several transcription factors (TFs) that efficiently regulate the production of these enzymes. Recently, a new TF named Azf1 was identified as a positive regulator of cellulase expression. Here, we investigated novel regulatory functions of Azf1 by its overexpression. In the mutant strain OEazf1, overexpression of azf1 was achieved under both repression and induction conditions. Although azf1 was more abundant in transcript and protein, overexpression of this TF did not activate transcription of the cellulase gene in the presence of the repressor glucose, suggesting that Azf1 may be subject to posttranslational regulation. In cellulose, the expression of swo, encoding the accessory protein swollenin, and the β-glucosidases cel1a, cel1b, cel3b, and cel3g increases in the early stages of cultivation. The increased production of these β-glucosidases increases the hydrolysis rate of cellobiose and sophorose, which activates carbon catabolite repression (CCR) and causes repression of cellulase genes and the regulator Xyr1 in the later stages of cultivation. Moreover, overexpression of azf1 led to increased cellulase activity in T. reesei during long-term cultivation in cellulose and sugarcane bagasse. Our results provide new insights into the mechanisms regulating Azf1 and novel genes that are important targets of this TF. This work contributes to a better understanding of the complex mechanisms regulating cellulase expression in T. reesei. It will contribute to the development of strains with higher production of these essential enzymes.

The ascomycete Trichoderma reesei is one of the most well studied cellulolytic microorganisms. This fungus is widely used in the biotechnology industry, mainly in the production of biofuels. Due to its importance, its genome was sequenced in 2008, opening new avenues to study this microorganism. In this ‘post-genomic’ era, a transcriptomic and proteomic era has emerged. Here, we present an overview of new findings in the gene expression regulation network of T. reesei. We also discuss new rational strategies to obtain mutants that produce hydrolytic enzymes with a higher yield, using metabolic engineering. Finally, we present how synthetic biology strategies can be used to create engineered promoters to efficiently synthesize enzymes for biomass degradation to produce bioethanol. Synthetic construction of regulatory elements such as transcriptional factors and promoters for T. reesei expands the toolbox for fungal engineering. The identification of novel cis-regulatory elements using systems biology approaches has begun to provide building blocks for constructing complex circuits in T. reesei. The use of synthetic biology standards, such as the BioBricks assembly format, for constructing modular systems for T. reesei has been reported, and its generalization should have a strong impact in the field. While the simultaneous implementation of multiple expression systems in T. reesei has been reported, there is still a pressing need to develop novel tools and techniques that could provide an efficient strain engineering platform for this organism.

In hematologic cancers, including leukemia, cells depend on amino acids for rapid growth. Anti-metabolites that prevent their synthesis or promote their degradation are considered potential cancer treatment agents. Amino acid deprivation triggers proliferation inhibition, autophagy, and programmed cell death. l-lysine, an essential amino acid, is required for tumor growth and has been investigated for its potential as a target for cancer treatment. l-lysine [alpha]-oxidase, a flavoenzyme that degrades l-lysine, has been studied for its ability to induce apoptosis and prevent cancer cell proliferation. In this study, we describe the use of l-lysine [alpha]-oxidase (LO) from the filamentous fungus Trichoderma harzianum for cancer treatment. The study identified and characterized a novel LO from T. harzianum and demonstrated that the recombinant protein (rLO) has potent and selective cytotoxic effects on leukemic cells by triggering the apoptotic cascade through mitochondrial dysfunction. The results support future translational studies using the recombinant LO as a potential drug for the treatment of leukemia.