Functional genomic analysis in Trichoderma reesei

Life of most organisms is dominated by nutrient availability, the need for reproduction and the rotation of the earth which results in daily changes between light and darkness. In the course of the proposed project we intend to dissect the molecular machinery for environmental sensing of the biotechnological workhorse Trichoderma reesei (Hypocrea jecorina) using cellulase gene expression and sexual development as model output pathways under controlled light conditions. Crucial factors for these processes are on the one hand genes involved in signal perception, most prominently represented by the G-protein coupled receptors, and on the other hand kinases and phosphatases, which represent signal transmitters to the respective target promotors. One crucial aim of our proposal is the elucidation of the connection of nutrient signaling with recognition of a potential mating partner and initiation of sexual development. Although it is well known that a deteriorating environment - especially lack of appropriate nutrient conditions - is beneficial for initiation of sexual development, this interrelationship has not been investigated with respect to interconnections to signaling pathways in that much detail. This joint project will provide the first comprehensive set of gene knock out strains of a specific group of genes along with an initial functional characterization in T. reesei. Investigation of the influence of signaling genes on sexual development as well as including genes known to be involved in sexual development in T. reesei will allow us to identify crossroads and checkpoints of the interrelationship between sexual development and environmental signaling. Consequently, we will not only apply high throughput gene knock-out for the first time in T. reesei at a large scale. With our case study we will also provide crucial insights into physiological relevance of signal transducing proteins at a broad range. The obtained data will also represent an invaluable basis and resource for research with signal transduction in fungi as well as for the industrial application of these organisms.

Life of most organisms is dominated by nutrient availability, the need for reproduction and the rotation of the earth which results in daily changes between light and darkness. In the course of the proposed project we intended to dissect the molecular machinery for environmental sensing of the biotechnological workhorse Trichoderma reesei (Hypocrea jecorina) using cellulase gene expression and sexual development as model output pathways under controlled light conditions. Crucial factors for these processes are on the one hand genes involved in signal perception, most prominently represented by the G-protein coupled receptors (GPCRs), and on the other hand kinases and phosphatases, which represent signal transmitters to the respective target promotors. In the course of this project, we created a knock out library of signalling components that now facilitates efficient investigation of the relevance of kinases, phosphatases and G-protein coupled receptors in physiology of T. reesei. Phenotypic analysis of groups of these genes revealed diverse functions in growth, sexual and asexual development, regulation of cellulase gene expression, protease production and secondary metabolism. Evaluation of the role of G-protein coupled receptors in surface sensing of natural substrates enabled us to assign the function of cellulose sensing to class XIII GPCRs. These receptors were found to sense glucose, a function which appears to be required also for the formation of attachment structures on natural cellulosic substrates. Our results highlight the major importance of kinases, phosphatases and GPCRs for utilization of cellulosic plant biomass by fungi. Consequently, the resources and knowledge established in the course of this project are highly interesting for industrial strain improvement of T. reesei in order to achieve sustainable, environmentally friendly green fermentation processes for facilitation of decarbonization of conventional chemical processes from biofuel production to multistep chemical conversion pathways using enzymes.