Unraveling Fungal Radiation Resistance Regulatory Networks through the Genome-Wide Transcriptome and Genetic Analyses of Cryptococcus neoformans

Abstract

The basidiomycetous fungus Cryptococcus neoformans has been known to be highly radiation resistant and has been found in fatal radioactive environments such as the damaged nuclear reactor at Chernobyl. To elucidate the mechanisms underlying the radiation resistance phenotype of C. neoformans, we identified genes affected by gamma radiation through genome-wide transcriptome analysis and characterized their functions. We found that genes involved in DNA damage repair systems were upregulated in response to gamma radiation. Particularly, deletion of recombinase RAD51 and two DNA-dependent ATPase genes, RAD54 and RDH54, increased cellular susceptibility to both gamma radiation and DNA-damaging agents. A variety of oxidative stress response genes were also upregulated. Among them, sulfiredoxin contributed to gamma radiation resistance in a peroxiredoxin/thioredoxin-independent manner. Furthermore, we found that genes involved in molecular chaperone expression, ubiquitination systems, and autophagy were induced, whereas genes involved in the biosynthesis of proteins and fatty acids/sterols were downregulated. Most importantly, we discovered a number of novel C. neoformans genes, the expression of which was modulated by gamma radiation exposure, and their deletion rendered cells susceptible to gamma radiation exposure, as well as DNA damage insults. Among these genes, we found that a unique transcription factor containing the basic leucine zipper domain, named Bdr1, served as a regulator of the gamma radiation resistance of C. neoformans by controlling expression of DNA repair genes, and its expression was regulated by the evolutionarily conserved DNA damage response protein kinase Rad53. Taken together, the current transcriptome and functional analyses contribute to the understanding of the unique molecular mechanism of the radiation-resistant fungus C. neoformans.

Transcriptomic analysis reveals the relationship of melanization to growth and resistance to gamma radiation in Cryptococcus neoformans

“…For genes in which the function is known, the vast majority were predicted to encode proteins involved in DNA repair or other processes associated with DNA processing, which included homologues of classical radiation‐associated proteins of the rad group, identified in S. cerevisiae , such as Rad7, 8, 51, 52 and 54 (Game and Mortimer, ; Bennett et al ., ). One notable difference from the previous transcriptomic study done in C. neoformans was that we observed few genes that were downregulated by radiation, whereas the majority of differentially expressed genes were downregulated after recovering for 1 h from 3000 Gy (Jung et al ., ). We suggest that radiation dose could greatly affect transcription results: in the present experiment, the dose resulted in much less death, so repair and recovery occurred in most of the cells.…”

“…The comparison presented here of the transcriptomes of melanized and non‐melanized, irradiated and non‐irradiated cells revealed patterns of differentially expressed genes that largely agree with these previous fungal transcriptome analyses (Watson et al ., ; Kimura et al ., ; Jung et al ., ). For genes in which the function is known, the vast majority were predicted to encode proteins involved in DNA repair or other processes associated with DNA processing, which included homologues of classical radiation‐associated proteins of the rad group, identified in S. cerevisiae , such as Rad7, 8, 51, 52 and 54 (Game and Mortimer, ; Bennett et al ., ).…”

“…The ability of fungi to occupy these niches suggests that they are resistant to various types of damaging electromagnetic radiation, including ionizing radiation (IR). This resistance has been recapitulated several times in laboratory studies: strains of Aureobasidium pullulans ‐ a dimorphic ascomycete – have been shown to withstand doses of γ radiation exceeding 10 kGy (Liu et al ., ; Jiang et al ., ), which is more than 1000‐fold higher than the commonly reported D 10 dose for mammalian cells (Anno et al ., ; Jung et al ., ). The maize pathogen Ustilago maydis , additionally, is extremely resistant to DNA damage inflicted by various types of radiation (Holloman et al ., ).…”

“…Studies in two model organisms, the budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe (Watson et al, 2004;Kimura et al, 2006) suggest that complicated networks of regulatory pathways are involved in coordinating the gene expression responses to IR, with the majority of the regulated genes associated with DNA damage checkpoints, chromosome partisioning, the cell cycle, and the general stress response. More recently, a transcriptome analysis of C. neoformans revealed additional genes and metabolic pathways that contribute to the radioresistance in this fungus (Jung et al, 2016). DNA repair enzymes, predictably, were upregulated and played a significant role in survival to high doses of γ-radiation.…”

The response of the melanized yeast Exophiala dermatitidis to gamma radiation exposure

“…This places it among the most resistant fungi known, similar to C . neoformans (332.5 [Jung et al ., ; Pacelli et al ., ; Schultzhaus et al ., )] and U . maydis [500 (Lee and Yarranton, ; Holloman et al ., ; Milisavljevic et al ., )].…”