Omics and epigenetic investigation of radiation resistance mechanism in melanized fungi

Zheng Wang 1 Zachary Schultzhaus 1 Amy Chen 1 Seongwon Kim 1 Greg Ellis 1 Melody Chiang 2 Dasha Leary 1 Igor Shuryak 3
1Center for Bio/Molecular Science and Engineering, Naval Research Laboratory, Washington, DC, USA
2., Thomas Jefferson High School for Science and Technology, Alexandria, Virginia, USA
3Center for Radiological Research, Columbia University, New York, NY, USA

The proliferation of melanized fungi under cosmic radiation on spacecraft, as well as the presence of numerous melanized fungal species in the damaged nuclear reactor at Chernobyl suggest that these fungi are resistant to ionizing radiation (IR). However, the molecular mechanisms that contribute to radiation resistance in melanized fungi are not well understood. The DNA repair machinery for these organisms, for example, is not extraordinary. To identify novel responses to IR in fungi, then, we have investigated two radiation-resistant, melanized yeasts: the ascomycete Wangiella dermatitidis and the basidiomycete Cryptococcus neoformans. In both organisms, unmelanized cells were only moderately susceptible to acute IR exposure. Initial investigations of the epigenetic response of W. dermatitidis to IR stress suggest that survival mechanisms do not strictly involve DNA methylation, but may include alteration of histone modifications. Additionally, we completed an -omics investigation of the C. neoformans radiation response. RNA-seq analysis of melanized and unmelanized C. neoformans treated with a sublethal, acute dose of gamma radiation (200Gy) revealed IR-responding genes including DNA repair enzymes, cell cycle proteins, fatty acid catabolic enzymes, and antioxidants. The most highly regulated genes, however, encoded novel hypothetical proteins containing various motifs associated with mitochondrial RNA stability, DNA binding, and protein secretion. Deleting these genes resulted in susceptibility to IR. In contrast to previous reports, moreover, approximately 800 genes were significantly regulated by L-DOPA, a substrate for melanization in C. neoformans. Analysis of transcriptome data using a machine learning approach verified that expression levels of a distinct cluster of 102 genes were lower in the melanized cells than in the non-melanized cells. This cluster of genes was negatively regulated in melanized cells but positively regulated in non-melanized cells upon exposure to IR. This radiation modulation was additionally correlated to a shotgun metabolomics analysis.