Histon deacetylases in fungi: novel functions

Acetylation of distinct lysine residues of core-histones is linked to transcriptional activation and histone deacetylases (HDACs) in many cases represent gene repressors. Inhibition of these enzymes by specific HDAC- inhibitors can induce growth arrest, differentiation, or apoptosis and therefore these compounds are being explored as therapeutic agents for the treatment of certain types of cancer. During the last decade, we have studied HDACs in different filamentous fungi and identified important biological functions of these enzymes not only in our model organism Aspergillus nidulans, but also in pathogenic species like Cochliobolus carbonum and Aspergillus fumigatus. In addition to their significance as pathogens, filamentous fungi are playing an outstanding role in pharmaceutical production, biotechnology, waste management, and food production. For a better control of these processes and an optimization of biotechnological production, the understanding of transcriptional regulation in these organisms, in particular on the chromatin level, is of crucial importance. Our results demonstrate, that distinct fungal HDACs like HosA and HdaA are involved in the regulation of secondary metabolites (SMs) that have attracted attention by virtue of their biotechnological and pharmaceutical applications. In addition to important pharmaceuticals such as penicillin, cephalosporin or cyclosporin, also toxins such as aflatoxins and ergot-alkaloids are part of the metabolic portfolio of filamentous fungi and many other SMs are still to be discovered. To elucidate the role of the class 1 HDAC HosA in the regulation of SM production in Aspergillus nidulans and other filamentous fungi is a major goal of this project proposal. The second class 1 enzyme, RpdA, turned out to be essential for Aspergillus nidulans and is distinguished by a fungal specific sequence motif that cannot be deleted without affecting the viability of the fungus. In the light of this finding, (fungal-specific) HDAC inhibitors might serve as therapeutic tools against fungal infections. Indeed, a first HDAC inhibitor, MGCD290, with intrinsic antifungal property was presented recently and is currently tested in clinical trails. In addition, there are strong indications that HOS2, the HosA orthologue in yeast, is involved in the regulation of fungal drug resistance. Thus, HDAC inhibitors might also have potential as additives used in combination with drugs administered within classical antifungal therapy regimes. Actually, synergy between azole- antifungals and HDAC inhibitors was recently observed in yeast and several higher fungi. The use of different HDAC mutant strains should contribute to our understanding of the mechanisms, by which fungal HDACs exert their catalytic activity in regulating fungal metabolism, drug resistance, and physiological functions essential for growth and development of these organisms.

In addition to their indispensable ecological role in the recycling of organic material, filamentous fungi are producers of a variety of medically important bioactive small molecules such as antibiotics or cholesterol-lowering drugs. As pathogens, however, some fungal species are also responsible for invasive infections with high mortality rates in immunosuppressed patients. As in higher eukaryotes, DNA of fungi is organized together with histones and other proteins in a more or less tightly packed structure called chromatin. The grade of chromatin condensation thereby determines the accessibility of genes or whole gene clusters for transcription factors. Histone modifying enzymes such as histone deacetylases (HDACs) contribute considerably to structural changes of chromatin and therefore are regulating the readout of genetic information. In this project, we were able to demonstrate that the class 1 HDAC HosA significantly represses the production of several bioactive small molecules in the filamentous fungus and model organism Aspergillus nidulans. Consequently, hosA deletion strains show a remarkably increased production of (partly novel) secondary metabolites that are not produced by wild type strains under growth conditions usually used in laboratories. Depletion of HosA activity thus might support the mining for bioactive molecules for novel (medical) applications in Aspergilli and other fungal species. On the other hand, however, synthesis of other fungal metabolites, most notably the antibiotic penicillin, is strictly dependent on HosA activity. In addition to its classical function as repressor, HosA obviously has activating functions as well - an unexpected and exciting regulatory mechanism that remains to be studied in detail. In a previous project, we were able to demonstrate that RpdA, another fungal class 1 HDAC, is required for optimal growth and development of Aspergillus nidulans. Investigations of this project confirmed these earlier results and revealed, that activity of RpdA-type enzymes is also crucial for growth but also germination of many fungal species including human pathogens. Its essential role turns RpdA into a potential target for histone deacetylase inhibitors with antifungal activity. The fact, that several of these substances have been already approved for the treatment of certain types of cancer, facilitates their use for novel therapeutic applications. Since our investigations revealed, that RpdA-type proteins comprise several structural peculiarities when compared to homologous enzymes of higher eukaryotes, the search for novel RpdA-specific inhibitors seems to be promising. Indeed, numerous novel HDAC inhibitors were identified during this project that exhibit a remarkable antifungal efficacy via inhibition of RpdA activity. In the light of a rising resistance of fungal pathogens to conventional therapies, RpdA inhibition via specific inhibitors might represent a promising new strategy to combat life-threatening fungal infections, alone or in combination with classical antifungal therapies.