A mutation in C. neoformans mitochondrial NADH dehydrogenase results in increased virulence in mice

Abstract

Cryptococcus neoformans: (H99W) was serially passaged in the invertebrate wax moth Galleria mellonella fifteen times to study how fungal virulence evolves under selection and whether those adaptations affect virulence. The G. mellonella passaged strain (P15) and the pre-passage H99W strains were used to infect three different host models of C. neoformans: C. elegans, G. mellonella , and Balb/c mice. While there was no difference in survival in the invertebrate models, P15 killed mice faster than H99W through both intratracheal and intravenous routes of infection and mice infected intravenously with P15 showed higher fungal burden in the brain. Characterization of the major virulence factors of C. neoformans found that P15 had increased capsule size, GXM release, and melanization. Whole genome sequencing of P15 and H99W revealed two mutations in P15, an insertion in the promoter region of NADH dehydrogenase (CNAG_09000) and an insertion in the LMP1 gene (CNAG_06765). Both ATP production and metabolic rate were higher in P15 compared to H99W. Quantitative RT-PCR suggested that the increased ATP was due to increased RNA levels of NADH dehydrogenase. Thus, adaptation to growth in hemocytes resulted in increased production of ATP, increased metabolic rate, and increased virulence in mice. This was likely due to differential expression of virulence factors, which skewed the host immune response to a less efficient Th2 response, with higher levels of IL-4, IL-10, and TNF-α in the brain. Overall, serial passage experiments have increased our understanding of how this yeast evolves under innate immune selection pressure.

The interplay between electron transport chain function and iron regulatory factors influences melanin formation inCryptococcus neoformans

“…Given that melanin formation is influenced by a mutation in the gene for mitochondrial NADH dehydrogenase, a component of complex I of the ETC (19), we hypothesized that mitochondrial functions influence laccase expression, trafficking, and/or activity. In particular, inhibition of ETC complexes I and III generates ROS including superoxide anion radical and hydrogen peroxide, and the subsequent influence on intracellular redox conditions may impact melanin formation (16–18, 22, 43).…”

“…A number of additional studies demonstrate the role of mitochondria the virulence of C. neoformans . For example, a mutation in the promoter of the gene encoding mitochondrial complex I protein NADH dehydrogenase increases capsule and melanin formation (19). Mutations in other regulatory factors also influence mitochondrial functions and virulence.…”

Succinate Dehydrogenase, Succinate, and Superoxides: A Genetic, Epigenetic, Metabolic, Environmental Explosive Crossroad

“…This makes the long-term use of each SDHI problematic and results in a faster turnover of SDHIs of the last generation that were shown to interfere with other ubiquinone-binding proteins, such as those involved in complex III [ 152 ]. On the other hand, the widespread use of SDHIs increases the risk of the appearance of dangerous drug-resistant microorganisms, including fungi [ 156 , 158 , 159 ]. These, with potential growth advantages, could in the future become multi-drug resistant and have uncontrollable broadening targets, from plants to other living organisms.…”

Succinate Dehydrogenase, Succinate, and Superoxides: A Genetic, Epigenetic, Metabolic, Environmental Explosive Crossroad

“…This has implied a consistently faster turnover of SDHI molecules of the last generation, additionally interfering with other ubiquinone-binding proteins such as those involved in complex III. On the other hand, the widespread use of SDHI increases the risk of appearance of dangerous drug-resistant microorganisms, including fungi [138][139][140]. These, with potential growth advantages, could in the future become multi-drug resistant and have uncontrollable broadening targets, from plant to other living organisms.…”