Myosin-II proteins are involved in the growth, morphogenesis, and virulence of the human pathogenic fungus Mucor circinelloides

Abstract

Mucormycosis is an emerging lethal invasive fungal infection. The infection caused by fungi belonging to the order Mucorales has been reported recently as one of the most common fungal infections among COVID-19 patients. The lack of understanding of pathogens, particularly at the molecular level, is one of the reasons for the difficulties in the management of the infection. Myosin is a diverse superfamily of actin-based motor proteins that have various cellular roles. Four families of myosin motors have been found in filamentous fungi, including myosin I, II, V, and fungus-specific chitin synthase with myosin motor domains. Our previous study on Mucor circinelloides, a common pathogen of mucormycosis, showed that the Myo5 protein (ID 51513) belonging to the myosin type V family had a critical impact on the growth and virulence of this fungus. In this study, to investigate the roles of myosin II proteins in M. circinelloides, silencing phenotypes and null mutants corresponding to myosin II encoding genes, designated mcmyo2A (ID 149958) and mcmyo2B (ID 136314), respectively, were generated. Those mutant strains featured a significantly reduced growth rate and impaired sporulation in comparison with the wild-type strain. Notably, the disruption of mcmyo2A led to an almost complete lack of sporulation. Both mutant strains displayed abnormally short, septate, and inflated hyphae with the presence of yeast-like cells and an unusual accumulation of pigment-filled vesicles. In vivo virulence assays of myosin-II mutant strains performed in the invertebrate model Galleria mellonella indicated that the mcmyo2A-knockout strain was avirulent, while the pathogenesis of the mcmyo2B null mutant was unaltered despite the low growth rate and impaired sporulation. The findings provide suggestions for critical contributions of the myosin II proteins to the polarity growth, septation, morphology, pigment transportation, and pathogenesis of M. circinelloides. The findings also implicate the myosin family as a potential target for future therapy to treat mucormycosis.

Alternative in-vivo models of mucormycosis

“…The same group recently demonstrated that myosin II plays an important role in growth, sporulation and hyphal development in Mucor. Interestingly, only disruption of one myosin II gene, mcmyo2A , led to abrogation of pathogenicity, while mcmyo2B deletion had no impact on virulence, although the mutant was similarly attenuated in growth ( Trieu et al., 2022 ). Iron acquisition is crucial for pathogenicity also in Mucorales, as M. lusitanicus that is not able to produce rhizoferrin, the mucormycete siderophore, was shown to be avirulent.…”

“…Therefore, this review will focus on alternative model systems that have been exploited so far for mucormycosis studies, highlighting novel alternative model animals or adapted methods established in the last three to four years to avoid repetition of what has already been reviewed in detail by Jacobsen (2019) . The alternative animal models presented in the review have significantly contributed to fostering our understanding of previously undescribed virulence factors of Mucorales, some of which are unique to this group of fungi ( Patiño-Medina et al., 2018 ; Patiño-Medina et al., 2019a ; Wurster et al., 2020 ; Alejandre-Castañeda et al., 2022 ; Itabangi et al., 2022 ; Trieu et al., 2022 ; Szebenyi et al., 2023 ). Moreover, these models have unveiled pathogen recognition and immune evasion strategies of Mucoralean fungi that are relevant for interactions with the human host ( Chamilos et al., 2008a ; Hoffmann et al., 2013 ; Inglesfield et al., 2018 ; López-Muñoz et al., 2018 ; Hassan et al., 2021 ; Itabangi et al., 2022 ).…”

Experimental Models to Study the Pathogenesis and Treatment of Mucormycosis

“…For experiments requiring drug treatment, dosage is determined by measuring the hemolymph volume of larvae of various weights and calculating the mean hemolymph volume/kg weight by linear regression [90]. The sporangiospore inoculum (usually 10 6 spores), any drugs being tested, and vehicles serving as infection or treatment controls, are injected diluted in 5-20 µL of insect physiological saline into the larval hemocoel via a hind pro-leg [89][90][91]. Infected larvae are incubated in the dark at 30 • C and monitored daily for survival.…”

Construction of the RNAi plasmids to suppress the expression of chitin synthase-encoding genes (<i>chs</i>) in fungus <i>Mucor lusitanicus</i>

“…This fungus is also one of the agents that cause mucormycosis. Our study analyzes the structures and functions of some genes and gene families involved in controlling the filamentous phenotype and pathogenicity of this fungus, such as the gene families encoding myosin II and myosin V (Trieu et al, 2017(Trieu et al, , 2022. These studies showed that the virulence of M. lusitanicus is closely related to some of its phenotypic characteristics.…”

“…These studies showed that the virulence of M. lusitanicus is closely related to some of its phenotypic characteristics. Specifically, growth rate, spore production, and yeast-like growth are signs that reduce the virulence of the fungus (Trieu et al, 2017(Trieu et al, , 2022.…”