A universal mark of centromeric chromatin is its packaging by a variant of histone H3 known as centromeric H3 (CenH3). The mechanism by which CenH3s are incorporated specifically into centromere DNA or the specialized function they serve there is not known. In a genetic approach to identify factors involved in CenH3 deposition, we screened for dosage suppressors of a temperaturesensitive cse4 allele in Saccharomyces cerevisiae (Cse4 is the S. cerevisiae CenH3). Independent screens yielded ORF YDL139C, which we named SCM3. Dosage suppression by SCM3 was specific for alleles affecting the histone fold domain of Cse4. Copurification and two-hybrid studies showed that Scm3 and Cse4 interact in vivo, and chromatin immunoprecipitation revealed that Scm3, like Cse4, is found associated with centromere DNA. Scm3 contains two essential protein domains, a Leu-rich nuclear export signal and a heptad repeat domain that is widely conserved in fungi. A conditional scm3 allele was generated to allow us to deplete Scm3. Upon Scm3 depletion, cells undergo a Mad2-dependent G 2/M arrest, and centromere localization of Cse4 is perturbed. We suggest that S. cerevisiae Scm3 defines a previously undescribed family of fungal kinetochore proteins important for CenH3 localization.centromeric H3 ͉ chromosome segregation ͉ kinetochore ͉ yeast ͉ CENP-A
Centromere DNA element II (CDEII) of budding yeast centromeres is an AT-rich sequence essential for centromere (CEN) function. Sequence analysis of Saccharomyces cerevisiae CDEIIs revealed that A 5-7 /T 5-7 tracts are statistically overrepresented at the expense of AA/TT and alternating AT. To test the hypothesis that this nonrandom sequence organization is functionally important, a CEN library in which the CDEII sequences were randomized was generated. The library was screened for functional and nonfunctional members following centromere replacement in vivo. Functional CENs contained CDEIIs with the highly biased A n /T n run distribution of native centromeres, while nonfunctional CDEIIs resembled those picked from the library at random. Run content, defined as the fraction of residues present in runs of four or more nucleotides, of the functional and nonfunctional CDEII populations differed significantly (P , 0.001). Computer searches of the genome for regions with an A 1 T content comparable to CDEIIs revealed that such loci are not unique to centromeres, but for 14 of the 16 chromosomes the AT-rich locus with the highest A n$4 1 T n$4 run content was the centromere. Thus, the distinctive and nonrandom sequence organization of CDEII is important for centromere function and possesses informational content that could contribute to the determination of centromere identity.
This article is the second in a series that examines the intelligibility of a person with congenital aglossia (PWCA). Specific factors examined in this study included (a) intelligibility for meaningful words versus nonsense words, (b) intelligibility for consonant-vowel-consonant words (CVCs) as a function of phonemic segment types, and (c) whether there is a correlation between intelligibility for these factors and the acoustic properties of the speech samples. Results revealed greater intelligibility for meaningful versus nonsense CVCs, greater intelligibility for back, low, and high-back versus front vowels embedded in CVCs, and greater intelligibility for productions as a function of phonemic variables, which demonstrated the following hierarchy: initial consonant > consonant vowel > vowel consonant > final consonant. Further results suggest that consonant recognition was consistently affected by “vowel context.” This suggests that movement sequencing appeared to be of importance for speech perception in productions of a PWCA.
Centromere H3 proteins (CenH3's) are variants of histone H3 specialized for packaging centromere DNA. Unlike canonical H3, which is among the most conserved of eukaryotic proteins, CenH3's are rapidly evolving, raising questions about orthology and conservation of function across species. To gain insight on CenH3 evolution and function, a phylogenetic analysis was undertaken on CenH3 proteins drawn from a single, ancient lineage, the Fungi. Using maximum-likelihood methods, a credible phylogeny was derived for the conserved histone fold domain (HFD) of 25 fungal CenH3's. The collection consisted mostly of hemiascomycetous yeasts, but also included basidiomycetes, euascomycetes, and an archaeascomycete. The HFD phylogeny closely recapitulated known evolutionary relationships between the species, supporting CenH3 orthology. The fungal CenH3's lacked significant homology in their N termini except for those of the Saccharomyces/Kluyveromyces clade that all contained a region homologous to the essential N-terminal domain found in Saccharomyces cerevisiae Cse4. The ability of several heterologous CenH3's to function in S. cerevisiae was tested and found to correlate with evolutionary distance. Domain swapping between S. cerevisiae Cse4 and the noncomplementing Pichia angusta ortholog showed that species specificity could not be explained by the presence or absence of any recognized secondary structural element of the HFD.
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