Development of a molecular genetics and cell biology toolbox for the filamentous fungus Diplodia sapinea.

Diplodia sapinea (Fr.) Fuckel is a widespread fungal pathogen affecting conifers worldwide. Infections can lead to severe symptoms, such as shoot blight, canker, tree death, or blue stain in harvested wood, especially in Pinus species.

A Reliable and Simple Method for the Production of Viable Pycnidiospores of the Pine Pathogen and a Spore-Based Infection Assay on Scots Pine.

is a globally distributed opportunistic fungal pathogen of conifers that causes severe production losses in forestry. The fungus frequently colonizes pine trees as an endophyte without causing visible symptoms but can become pathogenic when the host plant is weakened by stress, such as drought or heat. Forest damage might therefore further increase due to the effects of climate change.

Highly conserved, but highly specific: Somatic cell-cell fusion in filamentous fungi.

The development of ascomycete fungal colonies involves cell-cell fusion at different growth stages. In the model fungus Neurospora crassa, communication of two fusing cells is mediated by an unusual signaling mechanism, in which the two partners take turns in signal sending and receiving. In recent years, the molecular basis of this unusual cellular behavior has started to unfold, indicating the presence of an excitable signaling network.

A dialogue-like cell communication mechanism is conserved in filamentous ascomycete fungi and mediates interspecies interactions.

In many filamentous fungi, germinating spores cooperate by fusing into supracellular structures, which develop into the mycelial colony. In the model fungus Neurospora crassa, this social behavior is mediated by an intriguing mode of communication, in which two fusing cells take turns in signal sending and receiving. Here we show that this dialogue-like cell communication mechanism is highly conserved in distantly related fungal species and mediates interspecies interactions.

CAZymes from the thermophilic fungus Thermoascus aurantiacus are induced by C5 and C6 sugars.

Background: Filamentous fungi are excellent lignocellulose degraders, which they achieve through producing carbohydrate active enzymes (CAZymes). CAZyme production is highly orchestrated and gene expression analysis has greatly expanded understanding of this important biotechnological process. The thermophilic fungus Thermoascus aurantiacus secretes highly active thermostable enzymes that enable saccharifications at higher temperatures; however, the genome-wide measurements of gene expression in response to CAZyme induction are not understood.

The F-box protein gene - is a target for reverse engineering enzyme hypersecretion in filamentous fungi.

Carbohydrate active enzymes (CAZymes) are vital for the lignocellulose-based biorefinery. The development of hypersecreting fungal protein production hosts is therefore a major aim for both academia and industry. However, despite advances in our understanding of their regulation, the number of promising candidate genes for targeted strain engineering remains limited.

Development of genetic tools for the thermophilic filamentous fungus .

Background: Fungal enzymes are vital for industrial biotechnology, including the conversion of plant biomass to biofuels and bio-based chemicals. In recent years, there is increasing interest in using enzymes from thermophilic fungi, which often have higher reaction rates and thermal tolerance compared to currently used fungal enzymes. The thermophilic filamentous fungus produces large amounts of highly thermostable plant cell wall-degrading enzymes.

Plasma Membrane Fusion Is Specifically Impacted by the Molecular Structure of Membrane Sterols During Vegetative Development of .

Cell-to-cell fusion is crucial for the development and propagation of most eukaryotic organisms. Despite this importance, the molecular mechanisms mediating this process are only poorly understood in biological systems. In particular, the step of plasma membrane merger and the contributing proteins and physicochemical factors remain mostly unknown.

A nuclear protein NsiA from Epichloë festucae interacts with a MAP kinase MpkB and regulates the expression of genes required for symbiotic infection and hyphal cell fusion.

The endophytic fungus Epichloë festucae systemically colonizes the intercellular spaces of cool-season grasses to establish a mutualistic symbiosis. Hyphal growth of the endophyte within the host plant is tightly regulated and synchronized with the growth of the host plant. A genetic screen to identify symbiotic genes identified mutant FR405 that had an antagonistic interaction with the host plant.

Evidence of repeated horizontal transfer of sterol C-5 desaturase encoding genes among dikarya fungi.

Gene duplication and horizontal gene transfer (HGT) are two important but different forces for adaptive genome evolution. In eukaryotic organisms, gene duplication is considered to play a more important evolutionary role than HGT. However, certain fungal lineages have developed highly efficient mechanisms that avoid the occurrence of duplicated gene sequences within their genomes.

Phosphatidic acid produced by phospholipase D is required for hyphal cell-cell fusion and fungal-plant symbiosis.

Although lipid signaling has been shown to serve crucial roles in mammals and plants, little is known about this process in filamentous fungi. Here we analyze the contribution of phospholipase D (PLD) and its product phosphatidic acid (PA) in hyphal morphogenesis and growth of Epichloë festucae and Neurospora crassa, and in the establishment of a symbiotic interaction between E. festucae and Lolium perenne.

Plasma Membrane Integrity During Cell-Cell Fusion and in Response to Pore-Forming Drugs Is Promoted by the Penta-EF-Hand Protein PEF1 in .

Plasma membrane damage commonly occurs during cellular growth and development. To counteract these potentially lethal injuries, membrane repair mechanisms have evolved, which promote the integrity of the lipid bilayer. Although the membrane of fungi is the target of important clinical drugs and agricultural fungicides, the molecular mechanisms which mediate membrane repair in these organisms remain elusive.

Spatio-temporal MAPK dynamics mediate cell behavior coordination during fungal somatic cell fusion.

Mitogen-activated protein kinases (MAPKs) are conserved regulators of proliferation, differentiation and adaptation in eukaryotic cells. Their activity often involves changes in their subcellular localization, indicating an important role for these spatio-temporal dynamics in signal transmission. A striking model illustrating these dynamics is somatic cell fusion in Germinating spores of this fungus rapidly alternate between signal sending and receiving, thereby establishing a cell-cell dialog, which involves the alternating membrane recruitment of the MAPK MAK-2 in both fusion partners.

Establishment of Neurospora crassa as a host for heterologous protein production using a human antibody fragment as a model product.

Background: Filamentous fungi are commonly used as production hosts for bulk enzymes in biotechnological applications. Their robust and quick growth combined with their ability to secrete large amounts of protein directly into the culture medium makes fungi appealing organisms for the generation of novel production systems. The red bread mold Neurospora crassa has long been established as a model system in basic research.

Molecular Mechanisms Regulating Cell Fusion and Heterokaryon Formation in Filamentous Fungi.

For the majority of fungal species, the somatic body of an individual is a network of interconnected cells sharing a common cytoplasm and organelles. This syncytial organization contributes to an efficient distribution of resources, energy, and biochemical signals. Cell fusion is a fundamental process for fungal development, colony establishment, and habitat exploitation and can occur between hyphal cells of an individual colony or between colonies of genetically distinct individuals.

Accumulation of specific sterol precursors targets a MAP kinase cascade mediating cell-cell recognition and fusion.

Sterols are vital components of eukaryotic cell membranes. Defects in sterol biosynthesis, which result in the accumulation of precursor molecules, are commonly associated with cellular disorders and disease. However, the effects of these sterol precursors on the metabolism, signaling, and behavior of cells are only poorly understood.

Signal exchange and integration during self-fusion in filamentous fungi.

Growth and propagation of filamentous ascomycete fungi commonly involves vegetative cell fusion. In the red bread mold Neurospora crassa and many other ascomycete species, fusion occurs between germinating spores during colony formation and between hyphal branches in established mycelia. Both fusion processes promote the development and behavior of the fungal colony as a supra-cellular network.

The tetraspanin TSP3 of Neurospora crassa is a vacuolar membrane protein and shares characteristics with IDI proteins.

The fungal vacuole is an organelle, which adopts pleiotropic morphologies and functions. In aging and starving hyphae it is the compartment of degradation and recycling of cellular constituents. Here we identified TSP3, one of three tetraspanins present in the filamentous ascomycete fungus Neurospora crassa, as a vacuolar membrane protein.

Cell fusion in Neurospora crassa.

In recent years, the filamentous fungus Neurospora crassa has advanced as a model organism for studying eukaryotic cell-cell communication and fusion. Cell merger in this fungus employs an unusual mode of communication, in which the fusion partners appear to switch between signal sending and receiving. Many molecular factors mediating this intriguing mechanism and the subsequent membrane merger have been identified.

The AngFus3 Mitogen-Activated Protein Kinase Controls Hyphal Differentiation and Secondary Metabolism in Aspergillus niger.

Adaptation to a changing environment is essential for the survival and propagation of sessile organisms, such as plants or fungi. Filamentous fungi commonly respond to a worsening of their growth conditions by differentiation of asexually or sexually produced spores. The formation of these specialized cell types is, however, also triggered as part of the general life cycle by hyphal age or density.

A sensing role of the glutamine synthetase in the nitrogen regulation network in Fusarium fujikuroi.

In the plant pathogenic ascomycete Fusarium fujikuroi the synthesis of several economically important secondary metabolites (SM) depends on the nitrogen status of the cells. Of these SMs, gibberellin and bikaverin synthesis is subject to nitrogen catabolite repression (NCR) and is therefore only executed under nitrogen starvation conditions. How the signal of available nitrogen quantity and quality is sensed and transmitted to transcription factors is largely unknown.

HAM-2 and HAM-3 are central for the assembly of the Neurospora STRIPAK complex at the nuclear envelope and regulate nuclear accumulation of the MAP kinase MAK-1 in a MAK-2-dependent manner.

Intercellular communication and somatic cell fusion are important for fungal colony establishment, multicellular differentiation and have been associated with host colonization and virulence of pathogenic species. By a combination of genetic, biochemical and live cell imaging techniques, we characterized the Neurospora crassa STRIPAK complex that is essential for self-signalling and consists of the six proteins HAM-2/STRIP, HAM-3/striatin, HAM-4/SLMAP, MOB-3/phocein, PPG-1/PP2A-C and PP2A-A. We describe that the core STRIPAK components HAM-2 and HAM-3 are central for the assembly of the complex at the nuclear envelope, while the phosphatase PPG-1 only transiently associates with this central subcomplex.

Genetic basis of cell-cell fusion mechanisms.

Cell-cell fusion in sexually reproducing organisms is a mechanism to merge gamete genomes and, in multicellular organisms, it is a strategy to sculpt organs, such as muscle, bone, and placenta. Moreover, this mechanism has been implicated in pathological conditions, such as infection and cancer. Studies of genetic model organisms have uncovered a unifying principle: cell fusion is a genetically programmed process.

The NDR kinase scaffold HYM1/MO25 is essential for MAK2 map kinase signaling in Neurospora crassa.

Cell communication is essential for eukaryotic development, but our knowledge of molecules and mechanisms required for intercellular communication is fragmentary. In particular, the connection between signal sensing and regulation of cell polarity is poorly understood. In the filamentous ascomycete Neurospora crassa, germinating spores mutually attract each other and subsequently fuse.