Project description:Size is a fundamental feature of biological systems that affects physiology at all levels. For example, the dynamic, microtubule-based spindle that mediates chromosome segregation scales to a wide range of cell sizes across different organisms and cell types. Xenopus frog species possess a variety of egg and meiotic spindle sizes, and differences in activities or levels of microtubule-associated proteins in the egg cytoplasm between Xenopus laevis and Xenopus tropicalis have been shown to account for spindle scaling [1]. Increased activity of the microtubule severing protein katanin scales the X. tropicalis spindle smaller compared to X. laevis [2], as do elevated levels of TPX2, a protein that enriches the cross-linking kinesin-5 motor Eg5 at spindle poles [3]. To examine the conservation of spindle scaling mechanisms more broadly across frog species, we have utilized the tiny, distantly related Pipid frog Hymenochirus boettgeri. We find that egg extracts from H. boettgeri form meiotic spindles similar in size to X. tropicalis but that TPX2 and katanin-mediated scaling is not conserved. Instead, the microtubule depolymerizing motor protein kif2a functions to modulate spindle size. H. boettgeri kif2a possesses an activating phosphorylation site that is absent from X. laevis. Comparison of katanin and kif2a phosphorylation sites across a variety of species revealed strong evolutionary conservation, with X. laevis and X. tropicalis possessing distinct and unique alterations. Our study highlights the diversity and complexity of spindle assembly and scaling mechanisms, indicating that there is more than one way to assemble a spindle of a particular size.
Project description:THE skin secretion of many amphibians contains peptides that are able to kill a broad range of microorganisms (antimicrobial peptides: AMPs) and potentially play a role in innate immune defense. Similar to the toxin arsenals of various animals, amphibian AMP repertoires typically show major structural variation, and previous studies have suggested that this may be the result of diversifying selection in adaptation to a diverse spectrum of pathogens. Here we report on transcriptome analyses that indicate a very different pattern in the dwarf clawed frog H. boettgeri. Our analyses reveal a diverse set of transcripts containing two to six tandem repeats, together encoding 14 distinct peptides. Five of these have recently been identified as AMPs, while three more are shown here to potently inhibit the growth of gram-negative bacteria, including multi-drug resistant strains of the medically important Pseudomonas aeruginosa. Although the number of predicted peptides is similar to the numbers of related AMPs in Xenopus and Silurana frog species, they show significantly lower structural variation. Selection analyses confirm that, in contrast to the AMPs of other amphibians, the H. boettgeri peptides did not evolve under diversifying selection. Instead, the low sequence variation among tandem repeats resulted from purifying selection, recent duplication and/or concerted gene evolution. Our study demonstrates that defense peptide repertoires of closely related taxa, after diverging from each other, may evolve under differential selective regimes, leading to contrasting patterns of structural diversity.
Project description:Model organisms are crucial in research as they can provide key insights applicable to other species. This study proposes the use of the amphibian species Hymenochirus boettgeri, widely available through the aquarium trade, as a model organism for the study of chytridiomycosis, a disease caused by the fungus Batrachochytrium dendrobatidis (Bd) and linked to amphibian decline and extinction globally. Currently, no model organisms are used in the study of chytridiomycosis, particularly because of the lack of availability and nonstandardized methods. Thus, laboratories around the world use wild local species to conduct Bd infection experiments, which prevents comparisons between studies and reduces reproducibility. Here, we performed a series of Bd infection assays that showed that H. boettgeri has a dose- and genotype-dependent response, can generalize previous findings on virulence estimates in other species, and can generate reproducible results in replicated experimental conditions. We also provided valuable information regarding H. boettgeri husbandry, including care, housing, reproduction, and heat treatment to eliminate previous Bd infections. Together, our results indicate that H. boettgeri is a powerful and low-ecological-impact system for studying Bd pathogenicity and virulence.