Molecular Characterization of Xyloglucanase cel74a from Trichoderma reesei.

BACKGROUND: The filamentous fungus Trichoderma reesei is used on an industrial scale to produce enzymes of biotechnological interest. This fungus has a complex cellulolytic system involved in the degradation of lignocellulosic biomass. However, several aspects related to the regulation of the expression of holocellulolytic genes and the production of cellulases by this fungus are still understood. METHODS: Here, we constructed a null mutant strain for the xyloglucanase cel74a gene and performed the characterization of the Δcel74a strain to evaluate the genetic regulation of the holocellulases during sugarcane bagasse (SCB) cultivation. RESULTS: Our results demonstrate that the deletion of xyloglucanase cel74a may impact the regulation of holocellulase expression during SCB cultivation. The expression of cellulases cel7a, cel7b, and cel6a was reduced in Δcel74a strain, while the hemicellulases xyn1 and xyn2 were increased in the presence of SCB. The cel74a mutation also affected the xyloglucan hydrolysis patterns. In addition, CEL74A activity was modulated in the presence of calcium, suggesting that this ion may be required for efficient degradation of xyloglucan. CONCLUSIONS: CEL74A affects the regulation of holocellulolytic genes and the efficient degradation of SCB in T. reesei. This data makes a significant contribution to our understanding of the carbon utilization of fungal strains as a whole.

CRZ1 regulator and calcium cooperatively modulate holocellulases gene expression in Trichoderma reesei QM6a.

Trichoderma reesei is the main filamentous fungus used in industry to produce cellulases. Here we investigated the role of CRZ1 and Ca2+signaling in the fungus T. reesei QM6a concerning holocellulases production. For this, we first searched for potential CRZ1 binding sites in promoter regions of key genes coding holocellulases, as well as transcriptional regulators and sugar and calcium transporters. Using a nearly constructed T. reeseiAcrz1 strain, we demonstrated that most of the genes expected to be regulated by CRZ1 were affected in the mutant strain induced with sugarcane bagasse (SCB) and cellulose. In particular, our data demonstrate that Ca2+ acts synergistically with CRZ1 to modulate gene expression, but also exerts CRZ1-independent regulatory role in gene expression in T. reesei, highlighting the role of the major regulator Ca2+ on the signaling for holocellulases transcriptional control in the most part of cellulases genes here investigated. This work presents new evidence on the regulatory role of CRZ1 and Ca2+ sensing in the regulation of cellulolytic enzymes in T. reesei, evidencing significant and previously unknown function of this Ca2+sensing system in the control key transcriptional regulators (XYR1 and CRE1) and on the expression of genes related to sugar and Ca2+ transport.

New Genomic Approaches to Enhance Biomass Degradation by the Industrial Fungus Trichoderma reesei.

The filamentous fungi Trichoderma reesei is one of the most well-studied cellulolytic microorganisms. It is the most important fungus for the industrial production of enzymes to biomass deconstruction being widely used in the biotechnology industry, mainly in the production of biofuels. Here, we performed an analytic review of the holocellulolytic system presented by T. reesei as well as the transcriptional and signaling mechanisms involved with holocellulase expression in this fungus. We also discuss new perspectives about control of secretion and cellulase expression based on RNA-seq and functional characterization data of T. reesei growth in different carbon sources, which comprise glucose, cellulose, sophorose, and sugarcane bagasse.

The Duality of the MAPK Signaling Pathway in the Control of Metabolic Processes and Cellulase Production in Trichoderma reesei.

In this study, through global transcriptional analysis by RNA-Sequencing, we identified the main changes in gene expression that occurred in two functional mutants of the MAPK genes tmk1 and tmk2 in Trichoderma reesei during sugarcane bagasse degradation. We found that the proteins encoded by these genes regulated independent processes, sometimes in a cross-talk manner, to modulate gene expression in T. reesei. In the Δtmk2 strain, growth in sugarcane bagasse modulated the expression of genes involved in carbohydrate metabolism, cell growth and development, and G-protein-coupled receptor-mediated cell signaling. On the other hand, deletion of tmk1 led to decreased expression of the major genes for cellulases and xylanases. Furthermore, TMK1 found to be involved in the regulation of the expression of major facilitator superfamily transporters. Our results revealed that the MAPK signaling pathway in T. reesei regulates many important processes that allow the fungus to recognize, transport, and metabolize different carbon sources during plant cell wall degradation.

Defence responses in rice plants in prior and simultaneous applications of Cladosporium sp. during leaf blast suppression.

An alternative method to control rice blast (Magnaporthe oryzae) is to include biological agent in the disease management strategy. The objective of this study was to assess the leaf blast-suppressing effects of rice phylloplane fungi. One Cladosporium sp. phylloplane fungus was shown to possess biocontrolling traits based on its morphological characteristics and an analysis of its 18S ribosomal DNA. Experiments aimed at determining the optimal time to apply the bioagent and the mechanisms involved in its rice blast-suppressing activities were performed under controlled greenhouse conditions. We used foliar spraying to apply the Cladosporium sp. 48 h prior to applying the pathogen, and we found that this increased the enzymatic activity. Furthermore, in vitro tests performed using isolate C24 showed that it possessed the ability to secrete endoxylanases and endoglucanases. When Cladosporium sp. was applied either prior to or simultaneous with the pathogen, we observed a significant increase in defence enzyme activity, and rice blast was suppressed by 84.0 and 78.6 %, respectively. However, some enzymes showed higher activity at 24 h while others did so at 48 h after the challenge inoculation. Cladosporium sp. is a biological agent that is capable of suppressing rice leaf blast by activating biochemical defence mechanisms in rice plants. It is highly adapted to natural field conditions and should be included in further studies aimed at developing strategies to support ecologically sustainable disease management and reduce environmental pollution by the judicious use of fungicidal sprays.