Biology subjects

Prostak, S. M.

Publications and source records attributed to Prostak, S. M..

4 recordsLinked to original sources

bioRxiv2025

Homology-mediated transformation of frog-killing fungus Batrachochytrium dendrobatidis illuminates chytrid development and pathogenesis

The chytrid fungus Batrachochytrium dendrobatidis (Bd) infects amphibians and causes chytridiomycosis, a disease linked to global amphibian decline. Despite its ecological importance, Bd has lacked robust tools for genetic manipulation, limiting molecular insights into its development and pathogenicity. Here, we establish homologous recombination as a system for stable transformation of Bd, enabling targeted chromosomal integration of exogenous DNA, endogenous protein tagging, and targeted gene deletion. We use this system to visualize Bd infection in live amphibians to enhance understanding of host invasion and pathogenesis. We also use this system to test a previous hypothesis regarding the role of chitin synthases in Bd development by tagging the endogenous chitin synthase Myo17D and observing its rapid relocalization to the plasma membrane during de novo cell wall assembly. Finally, we use our homologous recombination approach for targeted gene deletion by knocking out the URA3 locus, and confirm the resulting genotype and phenotype via sequencing and drug resistance assays. This genetic transformation system offers a foundational tool for molecular studies of Bd, advancing our capacity to dissect molecular mechanisms of chytrid pathogenesis.

cell biology↗

bioRxiv2025

Frog-killing chytrid fungi deploy different strategies to regulate intracellular pressure in cell types that have or lack a cell wall

Cell morphogenesis is crucial for the physiology of animals and fungi alike. While animals typically shape their cells using the actin cytoskeleton, fungi control cell shape through polarized deposition of new cell wall material, which is inflated by intracellular osmotic "turgor" pressure. Understanding where and when these mechanisms evolved is essential for understanding the evolution of cell morphogenesis. To this end, we study chytrid fungi, which have a cell type that lacks a cell wall (the "zoospore") and a cell type that has a cell wall (the "sporangium"). While chytrid sporangia rely on polarized cell wall growth to control shape, we previously showed that the "frog-killing" chytrid fungus Batrachochytrium dendrobatidis (Bd) uses actin to control zoospore shape. Whether either zoospores or sporangia also use intracellular pressure regulation in cell shape control remains an open question. Here, we use live-cell imaging, environmental perturbations, and small molecule inhibitors to show that Bd sporangia generate and maintain turgor pressure, while Bd zoospores use specialized organelles called contractile vacuoles to pump water out of the cell, thereby keeping internal pressure low. Because chytrid fungi diverged prior to the evolution of the Dikarya--the fungal group comprising yeast, mushrooms, and filamentous fungi--these findings suggest that turgor pressure evolved early, and that cell morphogenesis underwent a major transition during early fungal evolution. We also suggest that the last common fungal ancestor may have, like chytrid fungi, employed stage-specific strategies for cell shape control--illustrating how developmental flexibility in cellular mechanisms can serve as a wellspring of evolutionary innovation.

cell biology↗

bioRxiv2022

Amphibian mucus triggers a developmental transition in the frog-killing chytrid fungus

The frog-killing fungus Batrachochytrium dendrobatidis (Bd) grows within the mucus-coated skin of amphibians. Each lifecycle, Bd transitions from its motile, dispersal form to its sessile growth form in a complex process called encystation. Although encystation is critical to Bd growth, whether and how this developmental transition is triggered by external signals was previously unknown. We discovered that exposure to amphibian mucus triggers rapid and reproducible encystation within minutes. This response can be recapitulated with purified mucin, the bulk component of mucus, but not by similarly-viscous methylcellulose or simple sugars. Mucin-induced encystation does not require gene expression, but does require surface adhesion, calcium signaling, and modulation of the actin cytoskeleton. Mucus-induced encystation may represent a key mechanism for synchronizing Bd development with arrival at the host.

cell biology↗

bioRxiv2020

The actin networks of chytrid fungi reveal evolutionary loss of cytoskeletal complexity in the fungal kingdom

Cells from across the eukaryotic tree use actin polymers and a number of conserved regulators for a wide variety of functions including endocytosis, cytokinesis, and cell migration. Despite this conservation, the actin cytoskeleton has undergone significant evolution and diversification, highlighted by the differences in the actin cytoskeletal networks of mammalian cells and yeast. Chytrid fungi diverged before the emergence of the Dikarya (multicellular fungi and yeast), and therefore provide a unique opportunity to study the evolution of the actin cytoskeleton. Chytrids have two life stages: zoospore cells that can swim with a flagellum, and sessile sporangial cells that, like multicellular fungi, are encased in a chitinous cell wall. Here we show that zoospores of the amphibian-killing chytrid Batrachochytrium dendrobatidis (Bd) build dynamic actin structures that resemble those of animal cells, including pseudopods, an actin cortex, and filopodia-like actin spikes. In contrast, Bd sporangia assemble actin patches similar to those of yeast, as well as perinuclear actin shells. Our identification of actin cytoskeletal elements in the genomes of five species of chytrid fungi indicate that these actin structures are controlled by both fungal-specific components as well as actin regulators and myosin motors found in animals but not other fungal lineages. The use of specific small molecule inhibitors indicate that nearly all of Bds actin structures are dynamic and use distinct nucleators: while pseudopods and actin patches are Arp2/3-dependent, the actin cortex appears formin-dependent, and actin spikes require both nucleators. The presence of animal- and yeast-like actin cytoskeletal components in the genome combined with the intermediate actin phenotypes in Bd suggests that the simplicity of the yeast cytoskeleton may be due to evolutionary loss.

cell biology↗