Microevolution of Cryptococcus neoformans Driven by Massive Tandem Gene Amplification

Abstract

The subtelomeric regions of organisms ranging from protists to fungi undergo a much higher rate of rearrangement than is observed in the rest of the genome. While characterizing these ~40-kb regions of the human fungal pathogen Cryptococcus neoformans, we have identified a recent gene amplification event near the right telomere of chromosome 3 that involves a gene encoding an arsenite efflux transporter (ARR3). The 3,177-bp amplicon exists in a tandem array of 2-15 copies and is present exclusively in strains with the C. neoformans var. grubii subclade VNI A5 MLST profile. Strains bearing the amplification display dramatically enhanced resistance to arsenite that correlates with the copy number of the repeat; the origin of increased resistance was verified as transport-related by functional complementation of an arsenite transporter mutant of Saccharomyces cerevisiae. Subsequent experimental evolution in the presence of increasing concentrations of arsenite yielded highly resistant strains with the ARR3 amplicon further amplified to over 50 copies, accounting for up to ~1% of the whole genome and making the copy number of this repeat as high as that seen for the ribosomal DNA. The example described here therefore represents a rare evolutionary intermediate-an array that is currently in a state of dynamic flux, in dramatic contrast to relatively common, static relics of past tandem duplications that are unable to further amplify due to nucleotide divergence. Beyond identifying and engineering fungal isolates that are highly resistant to arsenite and describing the first reported instance of microevolution via massive gene amplification in C. neoformans, these results suggest that adaptation through gene amplification may be an important mechanism that C. neoformans employs in response to environmental stresses, perhaps including those encountered during infection. More importantly, the ARR3 array will serve as an ideal model for further molecular genetic analyses of how tandem gene duplications arise and expand.

Sirtuins in the phylum Basidiomycota: A role in virulence in Cryptococcus neoformans

“…C. neoformans telomeres are very different to those seen in brewing yeast as they are not gene-poor, and there has been no reported evidence of silencing46. Consistent with this, Sir2 does not interact with the telomeric regions in this pathogen; the closest binding sites in the ChIP-seq data were a minimum of ~29 kb away from the ends of the chromosomes.…”

Product Binding Enforces the Genomic Specificity of a Yeast Polycomb Repressive Complex

“…By generating modification-specific antibodies, we found that H3K27me3 is deposited by the EZH2/E(z) ortholog across subtelomeric domains that repress the expression of underlying genes and noncoding transcripts. Polycomb heterochromatin may therefore have a specialized role in the regulation of subtelomeres, which in fungi are enriched for rapidly-evolving genes involved in niche adaptation and specialized metabolic functions (Brown et al, 2010; Chow et al, 2012). Consistent with this idea, the 41 kb average size of subtelomeric H3K27me3 domains closely matches a computational prediction of C. neoformans subtelomere size based on enrichment for metabolism-related gene products (~40 kb) (Chow et al, 2012).…”

“…Polycomb heterochromatin may therefore have a specialized role in the regulation of subtelomeres, which in fungi are enriched for rapidly-evolving genes involved in niche adaptation and specialized metabolic functions (Brown et al, 2010; Chow et al, 2012). Consistent with this idea, the 41 kb average size of subtelomeric H3K27me3 domains closely matches a computational prediction of C. neoformans subtelomere size based on enrichment for metabolism-related gene products (~40 kb) (Chow et al, 2012). Intriguingly, subtelomeric genes in other pathogenic fungi are silenced by domains of repressive chromatin and respond to environmental changes during the process of host infection, raising the possibility that Polycomb heterochromatin could contribute to the pathogenicity of C. neoformans (Domergue et al, 2005; Las Peñas et al, 2003; McDonagh et al, 2008).…”

The ace-1 Locus Is Amplified in All Resistant Anopheles gambiae Mosquitoes: Fitness Consequences of Homogeneous and Heterogeneous Duplications

“…They can confer a quantitative advantage in situations in which having larger amounts of the corresponding protein is advantageous. Examples of such coding-gene duplications abound for proteins involved in functions directly related to the environment (e.g., resistance to xenobiotics through higher levels of detoxification, [6–8]; greater amylase production providing adaptation to a starch-rich diet in humans and dogs, [9,10]; higher hexose transporter levels for adaptation to an environment in which resources are limited, [11]). However, homogeneous duplications are also often associated with deleterious pleiotropic effects (or selective costs), probably due to the disruption of biochemical balance or overproduction costs ([12], e.g., in Culex pipiens , esterases may account for up to 12% of total protein, by weight, [13]).…”