Functional analysis of a centromere from fission yeast: a role for centromere-specific repeated DNA sequences

1990 ◽  
Vol 10 (5) ◽  
pp. 1863-1872
Author(s):  
L Clarke ◽  
M P Baum
Keyword(s):  
Dna Sequences ◽  
Inverted Repeat ◽  
Repeated Sequence ◽  
Repeated Sequences ◽  
Central Core ◽  
Repeated Dna ◽  
Inverted Repeat Region ◽  
Repeated Dna Sequences ◽  
Centromere Function ◽  
Chromosome Ii

A circular minichromosome carrying functional centromere sequences (cen2) from Schizosaccharomyces pombe chromosome II behaves as a stable, independent genetic linkage group in S. pombe. The cen2 region was found to be organized into four large tandemly repeated sequence units which span over 80 kilobase pairs (kb) of untranscribed DNA. Two of these units occurred in a 31-kb inverted repeat that flanked a 7-kb central core of nonhomology. The inverted repeat region had centromere function, but neither the central core alone nor one arm of the inverted repeat was functional. Deletion of a portion of the repeated sequences that flank the central core had no effect on mitotic segregation functions or on meiotic segregation of a minichromosome to two of the four haploid progeny, but drastically impaired centromere-mediated maintenance of sister chromatid attachment in meiosis I. This requirement for centromere-specific repeated sequences could not be satisfied by introduction of random DNA sequences. These observations suggest a function for the heterochromatic repeated DNA sequences found in the centromere regions of higher eucaryotes.

scholarly journals Functional analysis of a centromere from fission yeast: a role for centromere-specific repeated DNA sequences.

1990 ◽  
Vol 10 (5) ◽  
pp. 1863-1872 ◽  
Author(s):  
L Clarke ◽  
M P Baum
Keyword(s):  
Dna Sequences ◽  
Inverted Repeat ◽  
Repeated Sequence ◽  
Repeated Sequences ◽  
Central Core ◽  
Repeated Dna ◽  
Inverted Repeat Region ◽  
Repeated Dna Sequences ◽  
Centromere Function ◽  
Chromosome Ii

A circular minichromosome carrying functional centromere sequences (cen2) from Schizosaccharomyces pombe chromosome II behaves as a stable, independent genetic linkage group in S. pombe. The cen2 region was found to be organized into four large tandemly repeated sequence units which span over 80 kilobase pairs (kb) of untranscribed DNA. Two of these units occurred in a 31-kb inverted repeat that flanked a 7-kb central core of nonhomology. The inverted repeat region had centromere function, but neither the central core alone nor one arm of the inverted repeat was functional. Deletion of a portion of the repeated sequences that flank the central core had no effect on mitotic segregation functions or on meiotic segregation of a minichromosome to two of the four haploid progeny, but drastically impaired centromere-mediated maintenance of sister chromatid attachment in meiosis I. This requirement for centromere-specific repeated sequences could not be satisfied by introduction of random DNA sequences. These observations suggest a function for the heterochromatic repeated DNA sequences found in the centromere regions of higher eucaryotes.

scholarly journals The chromatin structure of centromeres from fission yeast: differentiation of the central core that correlates with function.

1991 ◽  
Vol 112 (2) ◽  
pp. 191-201 ◽  
Author(s):  
C Polizzi ◽  
L Clarke
Keyword(s):  
Fission Yeast ◽  
Dna Sequences ◽  
Chromatin Structure ◽  
Chromatin Organization ◽  
Repeated Sequences ◽  
Central Core ◽  
Repeated Dna ◽  
Structural Differentiation ◽  
Yeast Saccharomyces Cerevisiae ◽  
Repeated Dna Sequences

We have examined the chromatin structure of centromere regions from the fission yeast Schizosaccharomyces pombe. The large and complex centromere regions of the S. pombe chromosomes encompass many kilobase pairs of DNA and contain several classes of tandemly repeated DNA sequences. The repeated sequences are further organized into a large inverted repeat flanking a central core, a conserved structural feature among all three centromeres in S. pombe. The nucleosomal configuration of the centromere regions is nonuniform and highly varied. Most of the centromere-specific repeated DNA sequences are packaged into nucleosomes typical of bulk chromatin. However, the central core and core-associated repeated sequences from the centromere regions of chromosomes I (cen1) and II (cen2), when present in S. pombe, show an altered chromatin structure, with little or no evidence of regular nucleosomal packaging. The atypical chromatin organization of the cen2 central core is not due to transcription, as no transcripts from this region were detected. These same DNA sequences, however, are packaged into nucleosomes typical of bulk chromatin when present in a nonfunctional environment on a minichromosome in the budding yeast Saccharomyces cerevisiae. Because the cen2 central core sequences themselves do not preclude regular nucleosomal packaging, we speculate that in S. pombe they constitute a specialized site of kinetochore protein assembly. The atypical nucleosomal pattern of the cen2 central core remains constant during the cell cycle, with only minor differences observed for some sequences. We propose that the unusual chromatin organization of the core region forms the basis of a higher order structural differentiation that distinguishes the centromere from the chromosome arms and specifies the essential structure for centromere function.

Clusters of interspersed repeated DNA sequences in the rice genome (Oryza)

Genome ◽  
1993 ◽  
Vol 36 (5) ◽  
pp. 944-953 ◽  
Author(s):  
Xinping Zhao ◽  
Gary Kochert
Keyword(s):  
Dna Sequences ◽  
Blot Hybridization ◽  
Rice Genome ◽  
Repeated Sequence ◽  
Repeated Sequences ◽  
Repeated Dna ◽  
Rice Varieties ◽  
Slot Blot Hybridization ◽  
Repeated Dna Sequence ◽  
Genus Oryza

We have characterized a repeated DNA sequence (RTL 122) from rice (Oryza sauva L.) with respect to its organization in the rice genome and its distribution among rice and other plants. The results indicate that the RTL 122 sequence is interspersed in the rice genome and limited to the genus Oryza. It is highly polymorphic and can be used to fingerprint rice varieties. A structure was observed in which several repeated sequences were clustered in DNA regions of 15–20 kb. We characterized three bacteriophage lambda clones that contained the RTL 122 sequence. Southern analysis using probes derived from restriction fragments of the three lambda clones indicated that all fragments except one are interspersed repeated sequences and belong to different repeated sequence families. Subsequent slot blot hybridization showed that most of them are only present within the genus Oryza. Some of the Oryza-specific, physically linked sequences show the same phylogenetic distribution, which suggests that these sequences might have evolved in a coordinate fashion. On the other hand, some of the repeated sequences have a different distribution even though they are physically adjacent in the genome. We speculate that such blocks of interspersed repeated sequences may serve as hotspots for rapid changes in the rice genome.Key words: rice, Oryza, repeated sequences, DNA fingerprinting, coordinated evolution.

Identification of DNA regions required for mitotic and meiotic functions within the centromere of Schizosaccharomyces pombe chromosome I

1991 ◽  
Vol 11 (4) ◽  
pp. 2206-2215
Author(s):  
K M Hahnenberger ◽  
J Carbon ◽  
L Clarke
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Sister Chromatid ◽  
Inverted Repeat ◽  
Distal Portion ◽  
Central Core ◽  
Inverted Repeat Region ◽  
Artificial Chromosomes ◽  
Centromere Function ◽  
Chromosome I ◽  
Meiosis I

We have determined the structural organization and functional roles of centromere-specific DNA sequence repeats in cen1, the centromere region from chromosome I of the fission yeast Schizosaccharomyces pombe. cen1 is composed of various classes of repeated sequences designated K', K"(dgl), L, and B', arranged in a 34-kb inverted repeat surrounding a 4- to 5-kb nonhomologous central core. Artificial chromosomes containing various portions of the cen1 region were constructed and assayed for mitotic and meiotic centromere function in S. pombe. Deleting K' and L from the distal portion of one arm of the inverted repeat had no effect on mitotic centromere function but resulted in greatly increased precocious sister chromatid separation in the first meiotic division. A centromere completely lacking K' and L, but containing the central core, one copy of B' and K" in one arm, and approximately 2.5 kb of the core-proximal portion of B' in the other arm, was also fully functional mitotically but again did not maintain sister chromatid attachment in meiosis I. However, deletion of K" from this minichromosome resulted in complete loss of centromere function. Thus, one copy of at least a portion of the K" (dgl) repeat is absolutely required but is not sufficient for S. pombe centromere function. The long centromeric inverted-repeat region must be relatively intact to maintain sister chromatid attachment in meiosis I.

scholarly journals Identification of DNA regions required for mitotic and meiotic functions within the centromere of Schizosaccharomyces pombe chromosome I.

1991 ◽  
Vol 11 (4) ◽  
pp. 2206-2215 ◽  
Author(s):  
K M Hahnenberger ◽  
J Carbon ◽  
L Clarke
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Sister Chromatid ◽  
Inverted Repeat ◽  
Distal Portion ◽  
Central Core ◽  
Inverted Repeat Region ◽  
Artificial Chromosomes ◽  
Centromere Function ◽  
Chromosome I ◽  
Meiosis I

We have determined the structural organization and functional roles of centromere-specific DNA sequence repeats in cen1, the centromere region from chromosome I of the fission yeast Schizosaccharomyces pombe. cen1 is composed of various classes of repeated sequences designated K', K"(dgl), L, and B', arranged in a 34-kb inverted repeat surrounding a 4- to 5-kb nonhomologous central core. Artificial chromosomes containing various portions of the cen1 region were constructed and assayed for mitotic and meiotic centromere function in S. pombe. Deleting K' and L from the distal portion of one arm of the inverted repeat had no effect on mitotic centromere function but resulted in greatly increased precocious sister chromatid separation in the first meiotic division. A centromere completely lacking K' and L, but containing the central core, one copy of B' and K" in one arm, and approximately 2.5 kb of the core-proximal portion of B' in the other arm, was also fully functional mitotically but again did not maintain sister chromatid attachment in meiosis I. However, deletion of K" from this minichromosome resulted in complete loss of centromere function. Thus, one copy of at least a portion of the K" (dgl) repeat is absolutely required but is not sufficient for S. pombe centromere function. The long centromeric inverted-repeat region must be relatively intact to maintain sister chromatid attachment in meiosis I.

scholarly journals A sister-strand exchange mechanism for recA-independent deletion of repeated DNA sequences in Escherichia coli.

Genetics ◽  
1993 ◽  
Vol 135 (3) ◽  
pp. 631-642 ◽  
Author(s):  
S T Lovett ◽  
P T Drapkin ◽  
V A Sutera ◽  
T J Gluckman-Peskind
Keyword(s):  
Escherichia Coli ◽  
Dna Sequences ◽  
Repeated Sequences ◽  
Repeated Dna ◽  
Repeated Dna Sequences ◽  
E Coli ◽  
Deletion Formation ◽  
Dna Strands

Abstract In the genomes of many organisms, deletions arise between tandemly repeated DNA sequences of lengths ranging from several kilobases to only a few nucleotides. Using a plasmid-based assay for deletion of a 787-bp tandem repeat, we have found that a recA-independent mechanism contributes substantially to the deletion process of even this large region of homology. No Escherichia coli recombination gene tested, including recA, had greater than a fivefold effect on deletion rates. The recA-independence of deletion formation is also observed with constructions present on the chromosome. RecA promotes synapsis and transfer of homologous DNA strands in vitro and is indispensable for intermolecular recombination events in vivo measured after conjugation. Because deletion formation in E. coli shows little or no dependence on recA, it has been assumed that homologous recombination contributes little to the deletion process. However, we have found recA-independent deletion products suggestive of reciprocal crossovers when branch migration in the cell is inhibited by a ruvA mutation. We propose a model for recA-independent crossovers between replicating sister strands, which can also explain deletion or amplification of repeated sequences. We suggest that this process may be initiated as post-replicational DNA repair; subsequent strand misalignment at repeated sequences leads to genetic rearrangements.

scholarly journals THE EVOLUTION OF RESTRICTED RECOMBINATION AND THE ACCUMULATION OF REPEATED DNA SEQUENCES

Genetics ◽  
1986 ◽  
Vol 112 (4) ◽  
pp. 947-962 ◽  
Author(s):  
Brian Charlesworth ◽  
Charles H Langley ◽  
Wolfgang Stephan
Keyword(s):  
Dna Sequences ◽  
Single Copy ◽  
Repeated Sequences ◽  
Stabilizing Selection ◽  
Crossing Over ◽  
Repeated Dna ◽  
Repeated Dna Sequences ◽  
Unequal Exchange ◽  
Single Copy Sequence ◽  
Speed Up

ABSTRACT We suggest hypotheses to account for two major features of chromosomal organization in higher eukaryotes. The first of these is the general restriction of crossing over in the neighborhood of centromeres and telomeres. We propose that this is a consequence of selection for reduced rates of unequal exchange between repeated DNA sequences for which the copy number is subject to stabilizing selection: microtubule binding sites, in the case of centromeres, and the short repeated sequences needed for terminal replication of a linear DNA molecule, in the case of telomeres. An association between proximal crossing over and nondisjunction would also favor the restriction of crossing over near the centromere. The second feature is the association between highly repeated DNA sequences of no obvious functional significance and regions of restricted crossing over. We show that highly repeated sequences are likely to persist longest (over evolutionary time) when crossing over is infrequent. This is because unequal exchange among repeated sequences generates single copy sequences, and a population that becomes fixed for a single copy sequence by drift remains in this state indefinitely (in the absence of gene amplification processes). Increased rates of exchange thus speed up the process of stochastic loss of repeated sequences.

Molecular characterization and chromosome location of repeated DNA sequences in Hordeum species and in the amphiploid tritordeum (×Tritordeum Ascherson et Graebner)

Genome ◽  
1995 ◽  
Vol 38 (5) ◽  
pp. 850-857 ◽  
Author(s):  
Esther Ferrer ◽  
Yolanda Loarce ◽  
Gregorio Hueros
Keyword(s):  
In Situ Hybridization ◽  
Dna Sequences ◽  
Diploid Species ◽  
Repeated Sequences ◽  
Hordeum Chilense ◽  
Repeated Dna ◽  
Repeated Dna Sequences ◽  
In Situ Experiments ◽  
Basic Genome

Genomic DNA from 19 species and subspecies representing the four basic genomes (H, I, X, and Y) of Hordeum was restricted with HaeIII and hybridized with two repeated DNA sequences of Hordeum chilense. The potential use of repeated sequences in ascertaining genomic affinities within the genus Hordeum was studied by comparing restriction fragment patterns. The study demonstrated the following: (i) species that shared a basic genome showed more similar hybridization fragment patterns than species with different genomes, whether with pHchl or pHch3; (ii) hybridization with pHchl revealed the presence of certain fragments limited to the species with a H genome; and (iii) the alloploid nature of species like H. jubatum was confirmed. The chromosomal distribution of the two repeated sequences was studied in species representing each basic genome and in the amphiploid tritordeum using fluorescent in situ hybridization. No interspecific differences were found between the diploid species. In situ experiments indicated the alloploid nature of H. depressum. Both sequences allow H. chilense chromatin to be distinguished from wheat chromosomes in tritordeum.Key words: repeated DNA sequences; in situ hybridization, Hordeum, tritordeum.

scholarly journals Mapping simple repeated DNA sequences in heterochromatin of Drosophila melanogaster.

Genetics ◽  
1993 ◽  
Vol 134 (4) ◽  
pp. 1149-1174 ◽  
Author(s):  
A R Lohe ◽  
A J Hilliker ◽  
P A Roberts
Keyword(s):  
Drosophila Melanogaster ◽  
Y Chromosome ◽  
Dna Sequences ◽  
Repeated Sequences ◽  
Chromosome 2 ◽  
Repeated Dna ◽  
Mitotic Chromosomes ◽  
Satellite Dnas ◽  
Repeated Dna Sequences ◽  
Chromosome Map

Abstract Heterochromatin in Drosophila has unusual genetic, cytological and molecular properties. Highly repeated DNA sequences (satellites) are the principal component of heterochromatin. Using probes from cloned satellites, we have constructed a chromosome map of 10 highly repeated, simple DNA sequences in heterochromatin of mitotic chromosomes of Drosophila melanogaster. Despite extensive sequence homology among some satellites, chromosomal locations could be distinguished by stringent in situ hybridizations for each satellite. Only two of the localizations previously determined using gradient-purified bulk satellite probes are correct. Eight new satellite localizations are presented, providing a megabase-level chromosome map of one-quarter of the genome. Five major satellites each exhibit a multi-chromosome distribution, and five minor satellites hybridize to single sites on the Y chromosome. Satellites closely related in sequence are often located near one another on the same chromosome. About 80% of Y chromosome DNA is composed of nine simple repeated sequences, in particular (AAGAC)n (8 Mb), (AAGAG)n (7 Mb) and (AATAT)n (6 Mb). Similarly, more than 70% of the DNA in chromosome 2 heterochromatin is composed of five simple repeated sequences. We have also generated a high resolution map of satellites in chromosome 2 heterochromatin, using a series of translocation chromosomes whose breakpoints in heterochromatin were ordered by N-banding. Finally, staining and banding patterns of heterochromatic regions are correlated with the locations of specific repeated DNA sequences. The basis for the cytochemical heterogeneity in banding appears to depend exclusively on the different satellite DNAs present in heterochromatin.

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