A Mucoralean White Collar-1 Photoreceptor Controls Virulence by Regulating an Intricate Gene Network during Host Interactions

Abstract

Mucolares are an ancient group of fungi encompassing the causal agents for the lethal infection mucormycosis. The high lethality rates, the emerging character of this disease, and the broad antifungal resistance of its causal agents are mucormycosis features that are alarming clinicians and researchers. Thus, the research field around mucormycosis is currently focused on finding specific weaknesses and targets in Mucorales for developing new treatments. In this work, we tested the role of the white-collar genes family in the virulence potential of Mucor lusitanicus. Study of the three genes of this family, mcwc-1a, mcwc-1b, and mcwc-1c, resulted in a marked functional specialization, as only mcwc-1a was essential to maintain the virulence potential of M. lusitanicus. The traditional role of wc-1 genes regulating light-dependent responses is a thoroughly studied field, whereas their role in virulence remains uncharacterized. In this work, we investigated the mechanism involving mcwc-1a in virulence from an integrated transcriptomic and functional approach during the host–pathogen interaction. Our results revealed mcwc-1a as a master regulator controlling an extensive gene network. Further dissection of this gene network clustering its components by type of regulation and functional criteria disclosed a multifunctional mechanism depending on diverse pathways. In the absence of phagocytic cells, mcwc-1a controlled pathways related to cell motility and the cytoskeleton that could be associated with the essential tropism during tissue invasion. After phagocytosis, several oxidative response pathways dependent on mcwc-1a were activated during the germination of M. lusitanicus spores inside phagocytic cells, which is the first stage of the infection. The third relevant group of genes involved in virulence and regulated by mcwc-1a belonged to the “unknown function,” indicating that new and hidden pathways are involved in virulence. The unknown function category is especially pertinent in the study of mucormycosis, as it is highly enriched in specific fungal genes that represent the most promising targets for developing new antifungal compounds. These results unveil a complex multifunctional mechanism used by wc-1 genes to regulate the pathogenic potential in Mucorales that could also apply to other fungal pathogens.

Stable and reproducible homologous recombination enables CRISPR-based engineering in the fungus Rhizopus microsporus

“…Consequently, other derived genetic tools were also developed in this fungus, such as gene overexpression ( Pérez-Arques et al., 2021a ), complementation ( Trieu et al., 2015 ), and protein tagging ( Navarro-Mendoza et al., 2019 ). During the last decade, this fungus has served as the unique model with a wide repertoire of genetic engineering tools in the urgent search for new target genes that are involved in Mucorales pathogenesis ( Lax et al., 2020 ; Li et al., 2011 ; Trieu et al., 2017 ; Navarro-Mendoza et al., 2018 ; Pérez-Arques et al., 2019 , 2021b ; López-Fernández et al., 2018 ; Patiño-Medina et al., 2018 , 2019 ). However, M. lusitanicus shows reduced virulence, and survival assays can only be done in the specific mouse strains following rigorous immunosuppression ( Li et al., 2011 ).…”

“…In addition, Mucorales can rapidly acquire new antifungal drug resistance through an exclusive RNAi-based mechanism to quickly and temporally generate resistant epimutants ( Calo et al., 2014 ). In this sense, most of the current studies in Mucorales are focused on investigating new genes, pathways, methodologies, and virulence factors that might be the targets for future antifungal developments against mucormycosis ( Binder et al., 2018 ; Gebremariam et al., 2019 ; Lax et al., 2020 ; López-Fernández et al., 2018 ; López-Muñoz et al., 2018 ; Navarro-Mendoza et al., 2018 ; Pérez-Arques et al., 2019 , 2021a , 2021b ; Trieu et al., 2017 ).…”

Mucorales and Mucormycosis: Recent Insights and Future Prospects

“…Structural genes are carB and carRP (mutants showing an albino phenotype), and regulatory genes are crgA (mutant showing a carotene overproducing phenotype). Additionally, the white-collar family ( mcwc1a , mcwc1b , and mcwc1c: mutants showing altered phototropism, crgA -dependent regulation, and light-defective induction; respectively) were functionally studied, proposing a complex model to explain the regulation of the light responses in Mucorales [ 84 , 90 , 91 , 92 , 93 , 94 , 95 ]. Later, the RNAi machinery of M. lusitanicus was thoroughly studied, dissecting all the steps of the canonical pathway: the triggering and amplification (genes rdrp1 and rdrp2 , mutants showing RNAi-defective phenotypes) [ 64 , 69 ], the production of siRNAs (genes dicer1 and dicer2 , mutants showing RNAi-defective phenotypes) [ 65 , 89 ], and the targeting machinery (genes ago1 , ago2 , and ago3 , mutants showing RNAi-defective phenotypes)) [ 66 ].…”

Genetic Manipulation in Mucorales and New Developments to Study Mucormycosis

“…Besides their natural high antifungal drug resistance, Mucorales can rapidly acquire new antifungal drug resistances through an exclusive RNAi-based mechanism to fast and temporally generate resistant epimutants [ 22 ]. In this sense, most of the current studies in Mucorales are focused on investigating new genes, pathways, methodologies, and virulence factors that might be the targets for future antifungal developments against mucormycosis [ 23 , 24 , 25 , 26 , 27 , 28 , 29 , 30 , 31 , 32 ]. However, this renewed interest in mucormycosis studies is still hampered by the few modern genetic tools available in Mucorales.…”