Camacho, E. | Search bioRxiv

bioRxiv2019

The structural unit of melanin in the cell wall of the fungal pathogen Cryptococcus neoformans

Melanins are synthesized macromolecules that are found in all biological kingdoms. These pigments have a myriad of roles that range from microbial virulence to key components of the innate immune response in invertebrates. Melanins also exhibit unique properties with potential applications in physics and material sciences, ranging from electrical batteries to novel therapeutics. In the fungi, melanins such as eumelanins, are components of the cell wall that provide protection against biotic and abiotic elements. Elucidation of the smallest fungal cell wall-asociated melanin unit that serves as a building block is critical to understand the architecture of these polymers, its interaction with surrounding components, and their functional versatility. In this study, we used isopycnic gradient sedimentation, NMR, EPR, high-resolution microscopy, and proteomics to analyze the melanin in the cell wall of the human pathogenic fungus Cryptococcus neoformans. We observed that melanin is assembled into the cryptococcal cell wall in spherical structures of [~]200 nm in diameter, termed melanin granules, which are in turn composed of nanospheres of [~]30 nm in diameter, the fungal melanosomes. We noted that melanin granules are closely associated with proteins that may play critical roles in the fungal melanogenesis and the supramolecular structure of this polymer. Using this structural information, we propose a model for C. neoformans melanization that is similar to the process used in animal melanization and is consistent with the phylogenetic relatedness of the fungal and animal kingdoms.

microbiology↗

bioRxiv2019

The enigmatic role of fungal annexins: the case of Cryptococcus neoformans

Annexins are multifunctional proteins that bind to phospholipid membranes in a calcium-dependent manner. Annexins play a myriad of critical and well-characterized roles in mammals, ranging from membrane repair to vesicular secretion. The role of annexins in the kingdoms of bacteria, protozoa and fungi have been largely overlooked. The fact that there is no known homologue of annexins in the model organism may contribute to this gap in knowledge. However, annexins are found in most medically important fungal pathogens, with the notable exception of Candida albicans. In this study we evaluated the function of the one annexin gene in Cryptococcus neoformans, a causative agent of cryptococcosis. This gene CNAG_02415, is annotated in the C. neoformans genome as a target of calcineurin through its transcription factor Crz1, and we propose to update its name to cryptococcal annexin, AnnexinC1. C. neoformans strains deleted for AnnexinC1 revealed no difference in survival after exposure to various chemical stressor relative the wild type, as well as no major alteration in virulence or mating. The only alteration observed in strains deleted for AnnexinC1 was a small increase in the titan cells formation in vitro. The preservation of annexins in many different fungal species suggests an important function, and therefore the lack of a strong phenotype for annexin-deficient C. neoformans is suggestive of either redundant genes that can compensate for the absence of AnnexinC1 function or novel functions not revealed by standard assays of cell function and pathogenicity. ImportanceCryptococcus neoformans is the deadliest human fungal pathogen, causing almost 200,000 deaths each year. Treatment of this lethal infection is lengthy, and in some patients therapy is not curative and patients require lifelong therapy. Fundamental research in this yeast is needed so that we can understand mechanisms of infection and disease and ultimately devise better therapies. In this work we investigated a fungal representative of the annexin family of proteins, specifically in the context of virulence and mating. We find that the cryptococcal annexin does not seem to be involved in virulence or mating but affects generation of titan cells, enlarged yeast cells that are detected in the lungs of mammalian hosts. Our data provides new knowledge in an unexplored area of fungal biology.

microbiology↗