Centromeres of Candida albicans form on unique and different DNA sequences but a closely related species, Candida tropicalis, possesses homogenized inverted repeat (HIR)-associated centromeres. To investigate the mechanism of centromere type transition, we improved the fragmented genome assembly and constructed a chromosome-level genome assembly of C. tropicalis by employing PacBio sequencing, chromosome conformation capture sequencing (3C-seq), chromoblot, and genetic analysis of engineered aneuploid strains. Further, we analyzed the 3D genome organization using 3C-seq data, which revealed spatial proximity among the centromeres as well as telomeres of seven chromosomes in C. tropicalis. Intriguingly, we observed evidence of inter-centromeric translocations in the common ancestor of C. albicans and C. tropicalis. Identification of putative centromeres in closely related Candida sojae, Candida viswanathii and Candida parapsilosis indicates loss of ancestral HIR-associated centromeres and establishment of evolutionary new centromeres (ENCs) in C. albicans. We propose that spatial proximity of the homologous centromere DNA sequences facilitated karyotype rearrangements and centromere type transitions in human pathogenic yeasts of the CUG-Ser1 clade.
The diploid budding yeast Candida albicans harbors unique CENPA-rich 3-to 5-kb regions that form the centromere (CEN) core on each of its eight chromosomes. The epigenetic nature of these CENs does not permit the stabilization of a functional kinetochore on an exogenously introduced CEN plasmid. The flexible nature of such centromeric chromatin is exemplified by the reversible silencing of a transgene upon its integration into the CENPA-bound region. The lack of a conventional heterochromatin machinery and the absence of defined boundaries of CENPA chromatin makes the process of CEN specification in this organism elusive. Additionally, upon native CEN deletion, C. albicans can efficiently activate neocentromeres proximal to the native CEN locus, hinting at the importance of CEN-proximal regions. In this study, we examine this CEN-proximity effect and identify factors for CEN specification in C. albicans. We exploit a counterselection assay to isolate cells that can silence a transgene when integrated into the CEN-flanking regions. We show that the frequency of reversible silencing of the transgene decreases from the central core of CEN7 to its peripheral regions. Using publicly available C. albicans high-throughput chromosome conformation capture data, we identify a 25-kb region centering on the CENPA-bound core that acts as CEN-flanking compact chromatin (CFCC). Cisand trans-chromosomal interactions associated with the CFCC spatially segregates it from bulk chromatin. We further show that neocentromere activation on chromosome 7 occurs within this specified region. Hence, this study identifies a specialized CEN-proximal domain that specifies and restricts the centromeric activity to a unique region. KEYWORDS centromere; CENPA; Hi-C; Candida albicans; neocentromere I N a majority of eukaryotes, centromere (CEN) specification is fulfilled by the assembly of the histone H3 variant, CENPA, and subsequent kinetochore stabilization. In most fungal species, CENs extend over to a region on every chromosome, ranging in sizes from 3 to 300 kb (Friedman and Freitag 2017), which are categorized as short regional or long regional CENs. The CENs in the Candida species are one of the most well-studied short regional CENs. Candida albicans has unique 3-5 kb CENPA-bound CEN DNA (Sanyal et al. 2004), as does C. dubliniensis (Padmanabhan et al. 2008) and C. lusitaniae (Kapoor et al. 2015). On the other hand, the 10-to 11-kb CENs in C. tropicalis consist of a 2-to 5-kb central core flanked by inverted repeats (Chatterjee et al. 2016). The 40-to 110-kb-long regional CENs of Schizosaccharomyces pombe contain a 10-to 14-kb-long CENPAbound region flanked by outer pericentric repeats (Clarke et al. 1986). The filamentous yeast Neurospora crassa harbors 150-300 kb of heterochromatic CENPA-rich DNA (Smith et al. 2011). Among the basidiomycetes, Cryptococcus neoformans contain 27-65 kb of transposon-rich CENPA-bound DNA sequences (Yadav et al. 2018). Evidently, regional CENs in fungi do not
50Aneuploidy is associated with drug resistance in fungal pathogens. In tropical 51 countries, Candida tropicalis is the most frequently isolated Candida species from 52 patients. To facilitate the study of genomic rearrangements in C. tropicalis, we 53 assembled its genome in seven gapless chromosomes by combining next-54 generation sequencing (NGS) technologies with chromosome conformation capture 55 sequencing (3C-seq). Our 3C-seq data revealed interchromosomal centromeric and 56 telomeric interactions in C. tropicalis, similar to a closely related fungal pathogen 57 Candida albicans. By performing a genome-wide synteny analysis between C. 58 tropicalis and C. albicans, we identified 39 interchromosomal synteny breakpoints 59 (ICSBs), which are relics of ancient translocations. Majority of ICSBs are mapped 60 within 100 kb of homogenized inverted repeat-associated (HIR) centromeres (17/39) 61 or telomere-proximal regions (7/39) in C. tropicalis. Further, we developed a genome 62 assembly of Candida sojae and used the available genome assembly of Candida 63 viswanathii, two closely related species of C. tropicalis, to identify the putative 64 centromeres. In both species, we identified the putative centromeres as HIR-65 associated loci, syntenic to the centromeres of C. tropicalis. Strikingly, a centromere-66 specific motif is conserved in these three species. Presence of similar HIR-67 associated putative centromeres in early-diverging Candida parapsilosis indicated 68 that the ancestral CUG-Ser1 clade species possessed HIR-associated centromeres. 69We propose that homology and spatial proximity-aided translocations among the 70 ancestral centromeres and loss of HIR-associated centromere DNA sequences led 71 to the emergence of evolutionary new centromeres (ENCs) on unique DNA 72 sequences. These events might have facilitated karyotype evolution and centromere-73 type transition in closely-related CUG-Ser1 clade species. 74 75 76 Significance Statement 77We assembled the genome of Candida tropicalis, a frequently isolated fungal 78 pathogen from patients in tropical countries, in seven complete chromosomes. 79 Comparative analysis of the CUG-Ser1 clade members suggests chromosomal 80 rearrangements are mediated by homogenized inverted repeat (HIR)-associated 81 centromeres present in close proximity in the nucleus as revealed by chromosome 82 conformation capture. These translocation events facilitated loss of ancestral HIR-83 associated centromeres and seeding of evolutionary new centromeres on unique 84 DNA sequences. Such karyotypic rearrangements can be a major source of genetic 85 variability in the otherwise majorly clonally propagated human fungal pathogens of 86 the CUG-Ser1 clade. The improved genome assembly will facilitate studies related to 87 aneuploidy-induced drug resistance in C. tropicalis. 88 89 Introduction 90 91 The efficient maintenance of the genetic material and its propagation to subsequent 92 generations determine the fitness of an organism. Genomic rearrangements are 93 often associat...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.