Steven D. Harris

Polarisome meets spitzenkörper: microscopy, genetics, and genomics converge.

The impact of filamentous fungi on human welfare has never been greater. Fungi are acknowledged as the most economically devastating plant pathogens ([1][1]) and are attaining increasing notoriety for their ability to cause life-threatening infections in humans ([57][2], [71][3]), and fungal

Farnesol‐induced apoptosis in Aspergillus nidulans reveals a possible mechanism for antagonistic interactions between fungi

The dimorphic fungus Candida albicans secretes farnesol, which acts as a quorum‐sensing molecule and prevents the yeast to mycelium conversion. In this study we examined the effect of farnesol in the filamentous fungus Aspergillus nidulans. We show that externally added farnesol has no effect on hyphal morphogenesis; instead, it triggers morphological features characteristic of apoptosis. Additional experiments suggest that mitochondria and reactive oxygen species (ROS) participate in farnesol‐induced apoptosis. Moreover, the effects of farnesol appear to be mediated by the FadA heterotrimeric G protein complex. Because A. nidulans does not secrete detectable amounts of farnesol, we propose that it responds to farnesol produced by other fungi. In agreement with this notion, growth and development were impaired in a farnesol‐dependent manner when A. nidulans was co‐cultivated with C. albicans. Taken together, our data suggest that farnesol, in addition to its quorum‐sensing function that regulates morphogenesis, is also employed by C. albicansTo reduce competition from other microbes.

Regulation of Apical Dominance in Aspergillus nidulans Hyphae by Reactive Oxygen Species

In fungal hyphae, apical dominance refers to the suppression of secondary polarity axes in the general vicinity of a growing hyphal tip. The mechanisms underlying apical dominance remain largely undefined, although calcium signaling may play a role. Here, we describe the localized accumulation of reactive oxygen species (ROS) in the apical region of Aspergillus nidulans hyphae. Our analysis of atmA (ATM) and prpA (PARP) mutants reveals a correlation between localized production of ROS and enforcement of apical dominance. We also provide evidence that NADPH oxidase (Nox) or related flavoproteins are responsible for the generation of ROS at hyphal tips and characterize the roles of the potential Nox regulators NoxR, Rac1, and Cdc42 in this process. Notably, our genetic analyses suggest that Rac1 activates Nox, whereas NoxR and Cdc42 may function together in a parallel pathway that regulates Nox localization. Moreover, the latter pathway may also include Bem1, which we propose represents a p40phox analog in fungi. Collectively, our results support a model whereby localized Nox activity generates a pool of ROS that defines a dominant polarity axis at hyphal tips.

Structure and Biosynthesis of Heat-Stable Antifungal Factor (HSAF), a Broad-Spectrum Antimycotic with a Novel Mode of Action

ABSTRACT A screen for antifungal compounds from Lysobacter enzymogenes strain C3, a bacterial biological control agent of fungal diseases, has previously led to the isolation of heat-stable antifungal factor (HSAF). HSAF exhibits inhibitory activities against a wide range of fungal species and shows a novel mode of antifungal action by disrupting the biosynthesis of a distinct group of sphingolipids. We have now determined the chemical structure of HSAF, which is identical to that of dihydromaltophilin, an antifungal metabolite with a unique macrocyclic lactam system containing a tetramic acid moiety and a 5,5,6-tricyclic skeleton. We have also identified the genetic locus responsible for the biosynthesis of HSAF in strain C3. DNA sequencing of this locus revealed genes for a hybrid polyketide synthase-nonribosomal peptide synthetase (PKS-NRPS), a sterol desaturase, a ferredoxin reductase, and an arginase. The disruption of the PKS-NRPS gene generated C3 mutants that lost the ability to produce HSAF and to inhibit fungal growth, demonstrating a hybrid PKS-NRPS that catalyzed the biosynthesis of the unique macrolactam system that is found in many biologically active natural products isolated from marine organisms. In addition, we have generated mutants with disrupted sterol desaturase, ferredoxin reductase, and arginase and examined the metabolites produced in these mutants. The work represents the first study of the genetic basis for the biosynthesis of the tetramic acid-containing macrolactams. The elucidation of the chemical structure of HSAF and the identification of the genetic locus for its biosynthesis establish the foundation for future exploitation of this group of compounds as new fungicides or antifungal drugs.

Cell Polarity in Filamentous Fungi: Shaping the Mold

The formation of highly polarized hyphae that grow by apical extension is a defining feature of the filamentous fungi. High-resolution microscopy and mathematical modeling have revealed the importance of the cytoskeleton and the Spitzenkorper (an apical vesicle cluster) in hyphal morphogenesis. However, the underlying molecular mechanisms remain poorly characterized. In this review, the pathways and functions known to be involved in polarized hyphal growth are summarized. A central theme is the notion that the polarized growth of hyphae is more complex than in yeast, though similar sets of core pathways are likely utilized. In addition, a model for the establishment and maintenance of hyphal polarity is presented. Key features of the model include the idea that polarity establishment is a stochastic process that occurs independent of internal landmarks. Moreover, the stabilization of nascent polarity axes may be the critical step that permits the emergence of a new hypha.

Autophagy in filamentous fungi

Autophagy is a ubiquitous, non-selective degradation process in eukaryotic cells that is conserved from yeast to man. Autophagy research has increased significantly in the last ten years, as autophagy has been connected with cancer, neurodegenerative disease and various human developmental processes. Autophagy also appears to play an important role in filamentous fungi, impacting growth, morphology and development. In this review, an autophagy model developed for the yeast Saccharomyces cerevisiae is used as an intellectual framework to discuss autophagy in filamentous fungi. Studies imply that, similar to yeast, fungal autophagy is characterized by the presence of autophagosomes and controlled by Tor kinase. In addition, fungal autophagy is apparently involved in protection against cell death and has significant effects on cellular growth and development. However, the only putative autophagy proteins characterized in filamentous fungi are Atg1 and Atg8. We discuss various strategies used to study and monitor fungal autophagy as well as the possible relationship between autophagy, physiology, and morphological development.

Regulation of hyphal morphogenesis by cdc42 and rac1 homologues in Aspergillus nidulans

The ability of filamentous fungi to form hyphae requires the establishment and maintenance of a stable polarity axis. Based on studies in yeasts and animals, the GTPases Cdc42 and Rac1 are presumed to play a central role in organizing the morphogenetic machinery to enable axis formation and stabilization. Here, we report that Cdc42 (ModA) and Rac1 (RacA) share an overlapping function required for polarity establishment in Aspergillus nidulans. Nevertheless, Cdc42 appears to have a more important role in hyphal morphogenesis in that it alone is required for the timely formation of lateral branches. In addition, we provide genetic evidence suggesting that the polarisome components SepA and SpaA function downstream of Cdc42 in a pathway that may regulate microfilament formation. Finally, we show that microtubules become essential for the establishment of hyphal polarity when the function of either Cdc42 or SepA is compromised. Our results are consistent with the action of parallel Cdc42 and microtubule‐based pathways in regulating the formation of a stable axis of hyphal polarity in A. nidulans.

Branching of fungal hyphae: regulation, mechanisms and comparison with other branching systems

The ability of rapidly growing hyphae to generate new polarity axes that result in the formation of a branch represents one of the most important yet least understood aspects of fungal cell biology. Branching is central to the development of mycelial colonies and also appears to play a key role in fungal interactions with other organisms. This review presents a description of the two major patterns of hyphal branching, apical and lateral, and highlights the roles of internal and external factors in the induction of branch formation. In addition, potential mechanisms underlying branch site selection are outlined, and the possible roles of multiple signaling pathways (i.e., G protein alpha, Cdc42, NDR kinases) and subcellular structures (i.e., the Spitzenkorper, septins) are discussed. Finally, other forms of branching in the plant and animal kingdoms are briefly summarized and compared to hyphal branching.

Septum formation in Aspergillus nidulans

Abstract Filamentous fungi form multicellular hyphae that are partitioned by septa. In A. nidulans, septum formation requires the assembly of a septal band following the completion of mitosis. Recent observations show that this band is a dynamic structure composed of actin, a septin and a formin. In addition, assembly is dependent upon a conserved protein kinase cascade that regulates mitotic exit and septation in yeast. Hyphal differentiation may reflect the regulation of this cascade by cyclin-dependent kinase activity. In this review, the dynamics and regulation underlying the assembly of the septal band are discussed.

A 2-stage ovarian cancer screening strategy using the Risk of Ovarian Cancer Algorithm (ROCA) identifies early-stage incident cancers and demonstrates high positive predictive value

A 2‐stage ovarian cancer screening strategy was evaluated that incorporates change of carbohydrate antigen 125 (CA125) levels over time and age to estimate risk of ovarian cancer. Women with high‐risk scores were referred for transvaginal ultrasound (TVS).