CaMtw1, a member of the evolutionarily conserved Mis12 kinetochore protein family, is required for efficient inner kinetochore assembly in the pathogenic yeast <i>Candida albicans</i>

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A fungus-specific outer kinetochore complex, the Dam1 complex, is essential in Saccharomyces cerevisiae, nonessential in fission yeast, and absent from metazoans. The reason for the reductive evolution of the functionality of this complex remains unknown. Both Candida albicans and Schizosaccharomyces pombe have regional centromeres as opposed to the short-point centromeres of S. cerevisiae. The interaction of one microtubule per kinetochore is established both in S. cerevisiae and C. albicans early during the cell cycle, which is in contrast to the multiple microtubules that bind to a kinetochore only during mitosis in S. pombe. Moreover, the Dam1 complex is associated with the kinetochore throughout the cell cycle in S. cerevisiae and C. albicans but only during mitosis in S. pombe. Here, we show that the Dam1 complex is essential for viability and indispensable for proper mitotic chromosome segregation in C. albicans. The kinetochore localization of the Dam1 complex is independent of the kinetochoremicrotubule interaction, but the function of this complex is monitored by a spindle assembly checkpoint. Strikingly, the Dam1 complex is required to prevent precocious spindle elongation in premitotic phases. Thus, constitutive kinetochore localization associated with a one-microtubule-one kinetochore type of interaction, but not the length of a centromere, is correlated with the essentiality of the Dam1 complex.The separation of sister chromatids during mitosis is driven by a dynamic interaction between two macromolecular structures: (i) the kinetochore (KT), a complex proteinaceous structure built on the centromere DNA (10, 47), and (ii) the mitotic spindle formed by microtubules (MTs). The process of chromosome segregation has been studied extensively in a wide range of organisms, from simple unicellular yeasts to humans. While the major sequence of events required for chromosome segregation remains the same, a few species-specific differences exist. The microtubule organizing centers (MTOCs) in budding yeasts, called spindle pole bodies (SPBs), are embedded in the nuclear envelope and assemble an intranuclear mitotic spindle (5). Thus, the interaction of spindle MTs with KTs does not require the breakdown of the nuclear envelope during closed mitosis in budding yeasts, and the KT-MT interaction is established early in the cell cycle (12, 23). The fission yeast Saccharomyces pombe also undergoes closed mitosis (23), but SPBs remain outside the nuclear envelope during interphase (27) and spindle MTs are nucleated and interact with KTs only when the duplicated SPBs enter the nuclear membrane following mitotic initiation (14). Metazoan cells, in contrast, undergo open mitosis (22,45) in which the MTs, originating from MTOCs, gain access to KTs only when the nuclear envelope breaks down during mitosis. Finally, the number of MTs that bind to a single chromosome differs in these organisms: in Saccharomyces cerevisiae only one MT binds per KT (52), in S. pombe two or three MTs bind per KT (13), and in metazoans multipl...

supporting

confidence: 50%

supporting

confidence: 50%

mentioning

confidence: 99%

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The Essentiality of the Fungus-Specific Dam1 Complex Is Correlated with a One-Kinetochore-One-Microtubule Interaction Present throughout the Cell Cycle, Independent of the Nature of a Centromere

Thakur

Sanyal

2011

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“…In S. cerevisiae, CENP-A/Cse4 proteolysis, which is mediated by Psh1, an E3 ubiquitin ligase, restricts its localization primarily to the centromere (30,56,102). Overexpression of CENP-A/CaCse4 results in recruitment of more CENP-A/CaCse4 molecules along with other kinetochore proteins such as Mtw1 at the centromere in C. albicans (17,106). Posttranslational modifications associated with the N-terminal histone tail are related to different functional states of chromatin-like repression (silencing) or activation of transcription.…”

Section: Cenp-a: the Universal Component Of Centromeric Chromatinmentioning

confidence: 99%

Diversity in Requirement of Genetic and Epigenetic Factors for Centromere Function in Fungi

Roy

Sanyal

2011

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A centromere is a chromosomal region on which several proteins assemble to form the kinetochore. The centromere-kinetochore complex helps in the attachment of chromosomes to spindle microtubules to mediate segregation of chromosomes to daughter cells during mitosis and meiosis. In several budding yeast species, the centromere forms in a DNA sequence-dependent manner, whereas in most other fungi, factors other than the DNA sequence also determine the centromere location, as centromeres were able to form on nonnative sequences (neocentromeres) when native centromeres were deleted in engineered strains. Thus, in the absence of a common DNA sequence, the cues that have facilitated centromere formation on a specific DNA sequence for millions of years remain a mystery. Kinetochore formation is facilitated by binding of a centromere-specific histone protein member of the centromeric protein A (CENP-A) family that replaces a canonical histone H3 to form a specialized centromeric chromatin structure. However, the process of kinetochore formation on the rapidly evolving and seemingly diverse centromere DNAs in different fungal species is largely unknown. More interestingly, studies in various yeasts suggest that the factors required for de novo centromere formation (establishment) may be different from those required for maintenance (propagation) of an already established centromere. Apart from the DNA sequence and CENP-A, many other factors, such as posttranslational modification (PTM) of histones at centric and pericentric chromatin, RNA interference, and DNA methylation, are also involved in centromere formation, albeit in a species-specific manner. In this review, we discuss how several genetic and epigenetic factors influence the evolution of structure and function of centromeres in fungal species.A complete understanding of the complexities of the process of cellular differentiation requires a thorough analysis of the molecular events occurring during eukaryotic cell division. As an important part of this process, a cell has to ensure accurate segregation of duplicated chromosomes into its progeny cells. In eukaryotes, specific DNA sequences, and the factors that bind to them immediately after replication, partly dictate the state of chromatin. Apart from the genetic factors, many epigenetic phenomena also contribute to formation of specialized chromatin required for specific functions. One such specialized chromatin domain, the centromere (CEN)-kinetochore complex, plays a crucial role in high-fidelity chromosome segregation and has great implications for human health. A consequence of improper chromosome segregation is the abnormal chromosome numbers associated with most human cancers. For example, in human colorectal cancers, almost 85% of cells are aneuploid, with 60 to 90 chromosomes (128).The high-fidelity chromosome segregation that occurs during mitosis and meiosis requires a functional centromere, defined as the primary constriction on a chromosome. The centromere is a region where spindle fibers attach to ...

“…First, there was a high propensity among the heterodiploid progeny strains to lose an entire haploid set of chromosomes from a species. Centromeres are clustered in C. albicans (2,23,30,31) and C. dubliniensis (19). The 5-FOA selection performed in this study probably enriches a population of cells where centromere clustering is somehow compromised, leading to the loss of the URA3-containing chromosome.…”

Section: Discussionmentioning

confidence: 98%

A Stable Hybrid Containing Haploid Genomes of Two Obligate Diploid Candida Species

Chakraborty

Aiyaz²,

Kakade

et al. 2013

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Candida albicans and Candida dubliniensis are diploid, predominantly asexual human-pathogenic yeasts. In this study, we constructed tetraploid (4n) strains of C. albicans of the same or different lineages by spheroplast fusion. Induction of chromosome loss in the tetraploid C. albicans generated diploid or near-diploid progeny strains but did not produce any haploid progeny. We also constructed stable heterotetraploid somatic hybrid strains (2n ؉ 2n) of C. albicans and C. dubliniensis by spheroplast fusion. Heterodiploid (n ؉ n) progeny hybrids were obtained after inducing chromosome loss in a stable heterotetraploid hybrid. To identify a subset of hybrid heterodiploid progeny strains carrying at least one copy of all chromosomes of both species, unique centromere sequences of various chromosomes of each species were used as markers in PCR analysis. The reduction of chromosome content was confirmed by a comparative genome hybridization (CGH) assay. The hybrid strains were found to be stably propagated. Chromatin immunoprecipitation (ChIP) assays with antibodies against centromere-specific histones (C. albicans Cse4/C. dubliniensis Cse4) revealed that the centromere identity of chromosomes of each species is maintained in the hybrid genomes of the heterotetraploid and heterodiploid strains. Thus, our results suggest that the diploid genome content is not obligatory for the survival of either C. albicans or C. dubliniensis. In keeping with the recent discovery of the existence of haploid C. albicans strains, the heterodiploid strains of our study can be excellent tools for further species-specific genome elimination, yielding true haploid progeny of C. albicans or C. dubliniensis in future.