Polypyrimidine segments in drosophila melanogaster DNA: I. Detection of a cryptic satellite containing polypyrimidine/polypurine DNA

Very long runs of pyrimidine nucleotides (polypyrimidines), previously detected in DNA from Drosophila melanogaster, have now been localized to a “cryptic” satellite. These polypyrimidines have an average length of 750 nucleotides and account for about 3% of the thymine residues in total DNA. The buoyant density of the DNA component which contains the polypyrimidines was detected by centrifuging native DNA to equilibrium in a CsCI gradient, and then assaying each fraction for its content of polypyrimidines. A peak was detected at a density of about 1.707 gm/cm³, distinctly heavier than the main band of DNA (1.702 gm/cm³). The buoyant density of polypyrimidine-containing molecules was little affected by differences in the molecular weight of the starting DNA in the range 10⁵-10⁷ daltons (single-stranded). Thus polypyrimidines (and their complementary polypurines) appear to form all or part of a “cryptic” satellite.Polypyrimidines have been isolated and characterized with respect to composition and buoyant density. Direct nucleoside analysis of unlabeled material indicated 34.5% deoxycytidine, 65.5% thymidine. Their banding position in neutral and alkaline CsCI gradients was consistent with a single-stranded DNA polymer of this composition.     

The drosophila ras oncogenes: Structure and nucleotide sequence

Three Drosophila genes homologous to the Ha-ras probe were isolated and mapped to positions 85D, 64B, and 62B on chromosome 3. Two of these genes (termed Dras1 and Dras2) were sequenced. In the case of Dras1, which contains multiple introns, a cDNA clone was isolated and sequenced. In the case of Dras2, the nucleotide sequence of the genomic clone was determined. Each gene codes for a protein with a predicted molecular weight of 21.6 kd. Alignment of the amino acid sequence of Dras1 with the vertebrate Ha-ras protein shows that at the amino terminus and central portion (residues 1–121 and 137–164) the two proteins are remarkably similar, and have an overall homology of 75%. The Dras2 gene lacks significant homology to the vertebrate counterpart at the extreme amino terminus and is homologous only between positions 28–120 and 139–161 (overall homology of 50%). This result suggests that the N terminus of p21 forms a distinct regulatory or functional domain. At the carboxy terminus, the major region of variability among the vertebrate ras proteins, the two Drosophila sequences also display considerable variability. However, both appear to be more similar to exon 4B of the Ki-ras gene.     

Nucleotide sequence analysis of DNA. II. Complete nucleotide sequence of the cohesive ends of bacteriophage lambda DNA

At the ends of bacteriophage λ DNA, the 5′-terminated strands are 12 nucleotides longer than the 3′-terminated strands. The complete sequence of deoxynucleotides in both the protruding 5′-terminated single strands of λ DNA has been determined by partial repair and by complete repair followed by sequencing of isolated oligonucleotides. Starting from the 5′-end of the left-hand cohesive end, the 12 nucleotides are in the sequence dpGpGpGpCpGpGpCpGpApCpCpT. The sequence from the right-hand cohesive end is exactly complementary to that from the left-hand end.     

Chromosome structure: DNA nucleotide sequence elements of a subset of the minichromosomes of the protozoan Trypanosoma brucei.

The genome of the protozoan Trypanosoma brucei contains a set of about 100 minichromosomes of about 50 to 150 kb in size. The small size of these chromosomes, their involvement in antigenic variation, and their mitotic stability make them ideal candidates for a structural analysis of protozoan chromosomes and their telomeres. We show that a subset of the minichromosomes is composed predominantly of simple-sequence DNA, with over 90% of the length of the minichromosome consisting of a tandem array of 177-bp repeats, indicating that these molecules have limited protein-coding capacity. Proceeding from the tip of the telomere to a chromosome internal position, a subset of the minichromosomes contained the GGGTTA telomere repeat, a 29-bp telomere-derived repeat, a region containing 74-bp G + C-rich direct repeats separated by approximately 155 bp of A + T-rich DNA that has a bent character, and 50 to 150 kb of the 177-bp repeat. Several of the minichromosome-derived telomeres did not encode protein-coding genes, indicating that the repertoire of telomeric variant cell surface glycoprotein genes is restricted to some telomeres only. The telomere organization in trypanosomes shares striking similarities to the organization of telomeres and subtelomeres in humans, yeasts, and plasmodia. An electron microscopic analysis of the minichromosomes showed that they are linear molecules without abnormal structures in the main body of the chromosome. The structure of replicating molecules indicated that minichromosomes probably have a single bidirectional origin of replication located in the body of the chromosome. We propose a model for the structure of the trypanosome minichromosomes.     

The DNA polymerase-primase from drosophila melanogaster embryos. Rate and fidelity of polymerization on single-stranded DNA templates

The DNA polymerase activity of the near homogeneous, multisubunit DNA polymerase-primase from Drosophila melanogaster embryos has been compared to Escherichia coli DNA polymerase III core, DNA polymerase III, and DNA polymerase III holoenzyme. The rate of deoxynucleotide incorporation by the Drosophila polymerase on singly primed phi X174 DNA is similar to that observed with equivalent levels of DNA polymerase III holoenzyme in the absence of E. coli single-stranded DNA binding protein. However, analysis of the DNA products indicates that the Drosophila polymerase is less processive than DNA polymerase III holoenzyme, and closely resembles DNA polymerase III. The Drosophila polymerase-primase contains neither 3'-5' exonuclease nor RNase H-like activities, and catalyzes no significant pyrophosphate exchange. There is a low level of DNA-dependent ATPase activity which can be eliminated by a second glycerol gradient sedimentation (Kaguni, L.S., Rossignol, J.-M., Conaway, R.C., and Lehman, I.R. (1983) Proc. Natl. Acad. Sci. U. S. A. 80, 2221-2225). Although lacking a 3'-5' exonuclease, the replication fidelity of the D. melanogaster polymerase is similar to that of E. coli DNA polymerase III holoenzyme which possesses such an activity.     

The nucleotide sequence of the initiator tRNA from Drosophila melanogaster.

The nucleotide sequence of Drosophila melanogaster methionine tRNAi was determined to be: pA-G-C-A-G-A-G-U-m1G-m2G-C-G-C-A-G-U-G-G-A-A-G-C-G-U-m2G-C-U-G-G-G-C-C-C-A-U-t6A-A-C-C-C-A-G-A-G-m7G-D-m5C-C-C-G-A-G-G-A-U-C-G-m1A-A-A-C-C-U-U-G-C-U-C-U-G-C-U-A-C-C-A(OH). It differs from vertebrate initiator tRNAs in only 6 out of 75 positions.     

The nucleotide sequence of histidine tRNA gamma of Drosophila melanogaster.

The nucleotide sequence of D. melanogaster histidine tRNA gamma was determined to be: pG-G-C-C-G-U-G-A-U-C-G-U-C-psi-A-G-D-G-G-D-D-A-G-G-A-C-C-C-C-A-C-G-psi-U-G-U-G- m1G-C-C-G-U-G-G-U-A-A-C-C-m5C-A-G-G-U-psi-C-G-m1A-A-U-C-C-U-G-G-U-C-A-C-G-G-m5C -A-C-C-AOH. An additional unpaired G is found at the 5' end, and the T in the TpsiC loop is replaced by a U.     

The nucleotide sequence of glutamate tRNA4 of Drosophila melanogaster.

The nucleotide sequence of Drosophila melanogaster glutamate tRNA4 was determined to be: pU-C-C-C-A-U-A-U-G-G-U-C-psi-A-G-D-G-G-C-D-A-G-G-A-U-A-U-C-U-G-G-C (m) -U-U-U-C-A-C-C-A-G-A-A-G-G-C-C-C-G-G-G-T-psi-U-C-G-A-U-U-C-C-C-G-G-U-A-U-G-G-G-A-A-C-C-AOH. A partial modified C is found at position 32 in the anticodon loop.     

The alpha-amylase gene in Drosophila melanogaster: nucleotide sequence, gene structure and expression motifs.

We present the complete nucleotide sequence of a Drosophila alpha-amylase gene and its flanking regions, as determined by cDNA and genomic sequence analysis. This gene, unlike its mammalian counterparts, contains no introns. Nevertheless the insect and mammalian genes share extensive nucleotide similarity and the insect protein contains the four amino acid sequence blocks common to all alpha-amylases. In Drosophila melanogaster, there are two closely-linked copies of the alpha-amylase gene and they are divergently transcribed. In the 5'-regions of the two gene-copies we find high sequence divergence, yet the typical eukaryotic gene expression motifs have been maintained. The 5'-terminus of the alpha-amylase mRNA, as determined by primer extension analysis, maps to a characteristic Drosophila sequence motif. Additional conserved elements upstream of both genes may also be involved in amylase gene expression which is known to be under complex controls that include glucose repression.     

The nucleotide sequence of the fission yeast DNA topoisomerase II gene: structural and functional relationships to other DNA topoisomerases.

We have determined the complete nucleotide sequence of a 5.3-kb long genomic DNA fragment of the fission yeast Schizosaccharomyces pombe that encodes DNA topoisomerase II. It contains a 4293 bp long single open reading frame. The predicted polypeptide has 1431 residues (mol. wt 162,000) and shows three characteristic domains; the large C-terminal region, which consists of alternating acidic-basic stretches and might be a chromatin-binding domain, the NH2 half domain homologous to the ATP-binding gyrB subunit of bacterial gyrase and the central-to-latter part which is homologous to the NH2 domain of the catalytic gyrA subunit, suggesting a possible evolutionary consequence of the gene fusion of the bacterial gyrase subunits into the eucaryotic DNA topoisomerase II gene. We have found that the cloned fission yeast TOP2 gene can complement the budding yeast top2 mutation, although the fission yeast TOP2 protein sequence is only 50% homologous to the recently determined sequence of budding yeast (J.C. Wang, personal communication). Conversely, the budding yeast TOP2 gene can complement the fission yeast top2 mutations, indicating that their DNA topoisomerase II genes are functionally exchangeable.     

Chromosomal location and nucleotide sequence of the Escherichia coli dapA gene.

Restriction fragments hybridizing to phage HP1c1 DNA were identified in digests of DNA from lysogenic strains of Haemophilus influenzae. The results showed that the cohesive ends of the mature phage DNA were joined in lysogens and that the phage genome was covalently linked to the host DNA, indicating that lysogeny involves recombination between specific sites on the phage and host chromosomes. The site on the phage chromosome at which this recombination occurred was between 110 and 750 base pairs of the left end on the mature phage genome.     

Comparative morphology of nucleolar DNA in Drosophila. II. Drosophila fulvimaculoides and drosophila tumiditarsus

This paper reports the demonstration, using fluorescence microscopy, of nucleolar DNA in two species of Drosophila. In Drosophila fulvimaculoides, the nucleolar DNA presents a variable morphology, suggestive of puffing activity. This material, which sometimes shows a banded structure like that of the polytene chromosomes, is shown not to be coextensive with the Y chromosome. Nucleolar DNA is demonstrated in Drosophila tumiditarsus also, and previous reports of an association of the dot chromosome with the nucleolus in this species are confirmed. The special usefulness of these two species for various sorts of investigation in pointed out.     

The nucleotide sequence of oocyte 5S DNA in Xenopus laevis. II. The GC-rich region

The primary sequence of the GC-rich half of the repeating unit in X. laevis 5S DNA has been determined in both a single plasmid-cloned repeating unit and in the total population of repeatig units. The GC-rich half of the repeating unit contains a single long duplication of 174 nucleotides. The duplicated segment commences 73 nucleotides preceding the 5' end of the gene and terminates at nucleotide 101 of the gene. The duplicated portion of the gene, termed the pseudogene, differs by 10 nucleotides from the corresponding portion of the gene, and the remaining duplicated sequence of 73 nucleotides differs by 13 nucleotides. The plasmid-cloned repeating unit differs from the dominant sequence in the total population repeating units by 6 nucleotides in the GC-rich region. Evidence is provided that most of the CpG dinucleotides in 5S DNA are at least partially methylated.     

Chromosome and nucleotide sequence differentiation in genomes of polyploid Triticum species

Summary The nature of genome change during polyploid evolution was studied by analysing selected species within the tribe Triticeae. The levels of genome changes examined included structural alterations (translocations, inversions), heterochromatinization, and nucleotide sequence change in the rDNA regions. These analyses provided data for evaluating models of genome evolution in polyploids in the genus Triticum, postulated on the basis of chromosome pairing at metaphase I in interspecies hybrids.The significance of structural chromosome alterations with respect to reduced MI chromosome pairing in interspecific hybrids was assayed by determining the incidence of heterozygosity for translocations and paracentric inversions in the A and B genomes of T. timopheevii ssp. araraticum (referred to as T. araraticum) represented by two lines, 1760 and 2541, and T. aestivum cv. Chinese Spring. Line 1760 differed from Chinese Spring by translocations in chromosomes 1A, 3A, 4A, 6A, 7A, 3B, 4B, 7B and possibly 2B. Line 2541 differed from Chinese Spring by translocations in chromosomes 3A, 6A, 6B and possibly 2B. Line 1760 also differed from Chinese Spring by paracentric inversions in arms 1AL and 4AL whereas line 2541 differed by inversions in 1BL and 4AL (not all chromosomes arms were assayed). The incidence of structural changes in the A and B genomes did not coincide with the more extensive differentiation of the B genomes relative to the A genomes as reflected by chromosome pairing studies.To assay changing degrees of heterochromatinization among species of the genus Triticum, all the diploid and polyploid species were C-banded. No general agreement was observed between the amount of heterochromatin and the ability of the respective chromosomes to pair with chromosomes of the ancestral species. Marked changes in the amount of heterochromatin were found to have occurred during the evolution of some of the polyploids.The analysis of the rDNA region provided evidence for rapid fixation of new repeated sequences at two levels, namely, among the 130 bp repeated sequences of the spacer and at the level of the repeated arrays of the 9 kb rDNA units. These occurred both within a given rDNA region and between rDNA regions on nonhomologous chromosomes. The levels of change in the rDNA regions provided good precedent for expecting extensive nucleotide sequence changes associated with differentiation of Triticum genomes and these processes are argued to be the principal cause of genome differentiation as revealed by chromosome pairing studies.