A "CAT" family of repetitive DNA sequences in Saccharomyces cerevisiae

An oligonucleotide probe was used to isolate yeast genomic clones containing DNA sequences with repetitive elements consisting primarily of a tandemly arranged trinucleotide, CAT. Hybridization analyses estimate that the yeast genome contains 40-50 CAT clusters, representing the first repetitive DNA sequence family found in yeast. Sequence analyses show short spacers between the CAT repeats consisting of closely related trinucleotides, primarily CGT. Some of the CAT clusters are located in longer repeating elements with lengths of 7 nucleotides or more. In one case a three-times-repeated 27-nucleotide sequence bears striking homology to the 21-base pair repeat region of the mammalian simian virus 40 promoter element. Hybridization studies further suggest that the "CAT" sequences may be widely dispersed in many diverse organisms including Escherichia coli, Drosophila, and man.     

Xenobiotic responsive element in the 5'-upstream region of the human P-450c gene.

The nucleotide sequence of the 5'-upstream region up to about -4.1 kb of the human P-450c gene was determined. Two kinds of repetitive sequences were located; one was the Alu sequence which was inserted at three positions (-3127 to -3038, -3017 to -2770, and -2167 to -1851), and the other was the SINE-R element located just upstream of the most distal Alu sequences. The region other than the two repeated sequences showed an overall similarity of 70% to that of the rat P-450c gene. Survey of XRE or its homologues, responsible for the inducible expression of the rat P-450c gene, revealed eight XRE core sequences in this region of the human P-450c gene. Three of them were carried in the Alu sequences. A fusion gene which was constructed by ligating the upstream region of the human P-450c gene to the chloramphenicol acetyltransferase (CAT) gene expressed the CAT activity in response to the inducer, methylcholanthrene, when transfected into Hepa-1 cells. Stepwise decrease in CAT activity in three regions was observed as the 5'-upstream sequence containing XRE motifs was removed. However, the XRE core sequence in the Alu sequences seemed inactive, because elimination of the three elements in the Alu sequences did not affect the expressed CAT activity. In accordance with this observation, competition experiments using gel mobility shift assay showed that XRE core sequences in the Alu sequences could not compete with the XRE sequence for the inducer-bound receptor.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evidence for transposition of dispersed repetitive DNA families in yeast.

Dispersed repetitive DNA sequences from yeast (Saccharomyces cerevisiae) nuclear DNA have been isolated as molecular hybrids in lambdagt. Related S. cerevisiae strains show marked alterations in the size of the restriction fragments containing these repetitive DNAs. "Ty1" is one such family of repeated sequences in yeast and consists of a 5.6 kilobase (kb) sequence including a noninverted 0.25 kb sequence of another repetitious family, "delta", on each end. There are about 35 copies of Ty1 and at least 100 copies of delta (not always associated with Ty1) in the haploid genome. A few Ty1 elements are tandem and/or circular, but most are disperse and show (along with delta) some sequence divergence between repeat units. Sequence alterations involving Ty1 elements have been found during the continual propagation of a single yeast clone over the course of a month. One region with a large number of delta sequences (SUP4) also shows a high frequency of sequence alterations when different strains are compared. One of the differences between two such strains involves the presence or absence of a Ty1 element. The novel joint is at one inverted pair of delta sequences.     

Identification and characterization of the centromere from chromosome XIV in Saccharomyces cerevisiae.

A functional centromere located on a small DNA restriction fragment from Saccharomyces cerevisiae was identified as CEN14 by integrating centromere-adjacent DNA plus the URA3 gene by homologous recombination into the yeast genome and then by localizing the URA3 gene to chromosome XIV by standard tetrad analysis. DNA sequence analysis revealed that CEN14 possesses sequences (elements I, II, and III) that are characteristic of other yeast centromeres. Mitotic and meiotic analyses indicated that the CEN14 function resides on a 259-base-pair (bp) RsaI-EcoRV restriction fragment, containing sequences that extend only 27 bp to the right of the element I to III region. In conjunction with previous findings on CEN3 and CEN11, these results indicate that the specific DNA sequences required in cis for yeast centromere function are contained within a region about 150 bp in length.     

Isolation of cloned DNA sequences containing ribosomal protein genes from Saccharomyces cerevisiae.

Yeast mRNA enriched for ribosomal protein mRNA was obtained by isolating poly(A)+ small mRNA from small polysomes. A comparison of cell-free translation of this small mRNA and total mRNA, and electrophoresis of the products on two-dimensional gels which resolve most yeast ribosomal proteins, demonstrated that a 5-10 fold enrichment for ribosomal protein mRNA was obtained. One hundred different recombinant DNA molecules possibly containing ribosomal protein genes were selected by differential colony hybridization of this enriched mRNA and unfractionated mRNA to a bank of yeast pMB9 hybrid plasmids. After screening twenty-five of these candidates, five different clones were found which contain yeast ribosomal protein gene sequences. The yeast mRNAs complementary to these five plasmids code for 35S-methionine-labeled polypeptides which co-migrate on two-dimensional gels with yeast ribosomal proteins. Consistent with previous studies on ribosomal protein mRNAs, the amounts of mRNA complementary to three of these cloned genes are controlled by the RNA2 locus. Although two of the five clones contain more than one yeast gene, none contain more than one identifiable ribosomal protein gene. Thus there is no evidence for "tight" linkage of yeast ribosomal protein genes. Two of the cloned ribosomal protein genes are single-copy genes, whereas two other cloned sequences contain two different copies of the same ribosomal protein gene. The fifth plasmid contains sequences which are repeated in the yeast genome, but it is not known whether any or all of the ribosomal protein gene on this clone contains repetitive DNA.     

Cloning of the fibroin gene from the oak silkworm, Antheraea yamamai and its complete sequence

The nucleotide sequences containing an entire genomic region and 5 upstream region of Antheraea yamamai fibroin gene have been determined. The gene consists of an initial exon encoding 14 amino acids, an intron (150 bp), and a long second exon coding for 2641 amino acids. The fibroin coding sequence shows a specialized organization with a highly repetitive region flanked by non repetitive 5 and 3 ends. Northern blot analyses confirmed that fibroin gene is actively expressed in the posterior silk gland of the final instar larvae of Antheraea yamamai.     

The construction of Arabidopsis expressed sequence tag assemblies. A new resource to facilitate gene identification.

The generation of large numbers of partial cDNA sequences, or expressed sequence tags (ESTs), has provided a method with which to sample a large number of genes from an organism. More than 25,000 Arabidopsis thaliana ESTs have been deposited in public databases, producing the largest collection of ESTs for any plant species. We describe here the application of a method of reducing redundancy and increasing information content in this collection by grouping overlapping ESTs representing the same gene into a "contig" or assembly. The increased information content of these assemblies allows more putative identifications to be assigned based on the results of similarity searches with nucleotide and protein databases. The results of this analysis indicate that sequence information is available for approximately 12,600 nonoverlapping ESTs from Arabidopsis. Comparison of the assemblies with 953 Arabidopsis coding sequences indicates that up to 57% of all Arabidopsis genes are represented by an EST. Clustering analysis of these sequences suggests that between 300 and 700 gene families are represented by between 700 and 2000 sequences in the EST database. A database of the assembled sequences, their putative identifications, and cellular roles is available through the World Wide Web.