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Biomedical subjects

A A Travers

Publications and source records attributed to A A Travers.

At least 37 records · Page 2Linked to original sources
PubMed1995

The homeobox gene repo is required for the differentiation and maintenance of glia function in the embryonic nervous system of Drosophila melanogaster.

We describe the cloning, expression and phenotypic characterisation of repo, a gene from Drosophila melanogaster that is essential for the differentiation and maintenance of glia function. It is not, however, required for the initial determination of glial cells. In the embryo, the gene, which encodes a homeodomain protein, is expressed exclusively in all developing glia and closely related cells in both the central and peripheral nervous systems. The only observed exceptions in the CNS are the midline glia derived from the mesectoderm and two of three segmental nerve root glial cells. Using a polyclonal antibody we traced the spatial and temporal pattern of the protein expression in detail. Embryos homozygous for null alleles of the protein exhibit late developmental defects in the nervous system, including a reduction in the number of glial cells, disrupted fasciculation of axons, and the inhibition of ventral nerve cord condensation. The expression of an early glial-specific marker is unaffected in such homozygotes. By contrast, the expression of late glial-specific markers is either substantially reduced or absent. The specificity of expression is also observed in the locust Schistocerca gregaria and is thus evolutionarily conserved.

Animals↗
PubMed1994

Differential association of linker histones H1 and H5 with telomeric nucleosomes in chicken erythrocytes.

Rat liver telomeric DNA is organised into nucleosomes characterised by a shorter and more homogeneous average nucleosomal repeat than bulk chromatin as shown by Makarov et al. (1). The latter authors were unable to detect the association of any linker histone with the telomeric DNA. We have confirmed these observations but show that in sharp contrast chicken erythrocyte telomeric DNA is organised into nucleosomes whose spacing length and heterogeneity are indistinguishable from those of bulk chromatin. We further show that chicken erythrocyte telomeric chromatin contains chromatosomes which are preferentially associated with histone H1 relative to histone H5. This contrasts with bulk chromatin where histone H5 is the more abundant species. This observation strongly suggests that telomeric DNA condensed into nucleosome core particles has a higher affinity for H1 than H5. We discuss the origin of the discrimination of the lysine rich histones in terms of DNA sequence preferences, telomere nucleosome preferences and particular constraints of the higher order chromatin structure of telomeres.

Animals↗
PubMed1994

The solution structure and dynamics of the DNA-binding domain of HMG-D from Drosophila melanogaster.

BACKGROUND: The HMG-box is a conserved DNA-binding motif that has been identified in many high mobility group (HMG) proteins. HMG-D is a non-histone chromosomal protein from Drosophila melanogaster that is closely related to the mammalian HMG-box proteins HMG-1 and HMG-2. Previous structures determined for an HMG-box domain from rat and hamster exhibit the same global topology, but differ significantly in detail. It has been suggested that these differences may arise from hinge motions which allow the protein to adapt to the shape of its target DNA. RESULTS: We present the solution structure of HMG-D determined by NMR spectroscopy to an overall precision of 0.85 A root mean squared deviation (rmsd) for the backbone atoms. The protein consists of an extended amino-terminal region and three alpha-helices that fold into a characteristic 'L' shape. The central core region of the molecule is highly stable and maintains an angle of approximately 80 degrees between the axes of helices 2 and 3. The backbone dynamics determined from 15N NMR relaxation measurements show a high correlation with the mean residue rmsd determined from the calculated structures. CONCLUSIONS: The structure determined for the HMG-box motif from HMG-D is essentially identical to the structure determined for the B-domain of mammalian HMG-1. Since these proteins have significantly different sequences our results indicate that the global fold and the mode of interaction with DNA are also likely to be conserved in all eukaryotes.

Amino Acid Sequence↗
PubMed1994

DNA transcription. Keeping the writhe.

A new model system has been used to demonstrate how a nucleosome may remain associated with an actively transcribed gene and yet maintain DNA compaction during transcription.

Chromatin↗
PubMed1994

HMG-D, the Drosophila melanogaster homologue of HMG 1 protein, is associated with early embryonic chromatin in the absence of histone H1.

We show that HMG-D, an abundant chromosomal protein, is associated with condensed chromatin structures during the first six nuclear cleavage cycles of the developing Drosophila embryo and that histone H1 is absent from these same structures. As H1 accumulates from nuclear division 7 onwards, the nuclei become more compact and transcriptionally active. This compaction is paralleled by a reduction in size of mitotic chromatin. In addition, we find a striking correlation between the switch in HMG-D:H1 ratios and the changes that occur between nuclear cycles 8 and 13 that are collectively termed the mid-blastula transition. This transition is characterized by an increase in the nuclear cycle times, a change in the nucleo-cytoplasmic ratio, and a 5- to 20-fold decrease in nuclear volume. We propose that this is a direct consequence of a re-organization of chromatin from a less condensed state with HMG-D to a more condensed state with H1. We argue that HMG-D, either by itself or in conjunction with other chromosomal proteins, induces a condensed state of chromatin that is distinct from, and less compact than the H1-containing 30 nm fibre and that this state of chromatin could facilitate rapid nuclear cycles.

Animals↗
PubMed1994

DNA sequence organization in chromatosomes.

The chromatosome is a structural unit of chromatin which contains a histone octamer and one linker histone molecule (H5, H1a or H1b in chicken erythrocytes) bound to 168 bp of DNA (= core particle DNA extended by 22 bp). We have cloned and sequenced 280 DNA fragments of 163 to 173 bp in length isolated from chicken erythrocyte chromatosomes. We have analysed both this set and a subset of 171 clones whose lengths varied between 166 and 170 bp. The periodic modulation of the frequency of occurrence of trinucleotide sequences is neither as regular nor as pronounced for core particle DNA, even for the trinucleotide ApApA/TpTpT. Nevertheless for this trinucleotide the congruence of the preferential and avoided locations between the two sets is remarkable. We conclude that the rotational positioning of the DNA bound to the histone octamer is essentially the same in chromatosomes and nucleosome core particles. This conclusion is confirmed by Fourier analysis, which shows that within the central 145 bp the average rotational periods of about 10-20 bp and phases are very similar to those in core DNA. However the amplitude of the modulations is less, indicating that the binding of the linker histone can overcome to a certain extent the sequence-dependent bending preferences of DNA. Although the rotational signals are largely conserved the sequence organization of the chromatosomal DNA differs in significant ways from that of core particle DNA. The major difference is the preferential occurrence of short G+C-rich instead of A+T-rich sequences at the midpoint of the binding site. The second apparent difference is the exclusion of short oligo(dA).(dT) tracts from positions about 40 to 43 bp on either side of the midpoint in chromatosomal, but not in core particle DNA. Finally, we show that particular base-steps are preferentially enriched close to the termini of chromatosomal DNA and propose that these sequences may constitute one of the two DNA binding sites for the bivalent globular domain of histones H5 or H1. The implications for the positioning of GH5 relative to the path of the DNA superhelix are discussed.

Animals↗
PubMed1993

dHMG-Z, a second HMG-1-related protein in Drosophila melanogaster.

We report the identification of dHMG-Z, a gene related to dHMG-D and encoding a second invertebrate homologue of HMG 1 protein. The encoded proteins is 65% identical to dHMG-D protein, and also contains a single HMG-box as the DNA recognition motif. Analogous to dHMG-D, two transcripts are observed for dHMG-Z which are differentially regulated, and are the product of zygotic transcription unlike the dHMG-D transcripts which arise from both maternal and zygotic transcription. The genes for dHMG-D and dHMG-Z are located on adjacent loci in the genome and each contains two introns. The position of the second intron in the coding region is conserved between the two genes suggesting a common origin via gene duplication.

Animals↗
PubMed1993

The Escherichia coli FIS protein is not required for the activation of tyrT transcription on entry into exponential growth.

The Escherichia coli DNA bending protein factor for inversion stimulation (FIS), is neither necessary nor responsible for the stimulation of transcription from the wild type promoter for the tyrT operon (encoding a species of tyrosine tRNA) that occurs upon resumption of exponential growth. This conclusion is unexpected given that the regulatory element required for optimal transcription of tyrT contains three binding sites for FIS protein. In addition, it is in apparent conflict with reports from other laboratories which have described FIS-dependent activation of the stable RNA promoters rrnB P1 and thrU(tufB) in vivo. However, tyrT transcription is stimulated in a FIS-dependent manner both in vivo and in vitro when promoter function is impaired by mutation of the promoter itself or by the addition of the polymerase effector guanosine 5'-diphosphate 3'-diphosphate. These conditions, which expose a requirement for activation of stable RNA synthesis by FIS, suggest that FIS serves an adaptive role permitting high levels of stable RNA transcription on nutritional shift-up when RNA polymerase levels are depleted. In principle such a mechanism could confer a significant selective advantage thus accounting for the conservation of FIS binding sites in the regulatory regions of stable RNA promoters.

Bacterial Proteins↗
PubMed1992

Sequence-specific DNA binding by a two zinc-finger peptide from the Drosophila melanogaster Tramtrack protein.

We show that the DNA-binding domain of the Drosophila melanogaster regulatory protein Tramtrack consists of a 66 amino acid sequence containing two zinc-finger motifs and a short sequence N-terminal to the first finger motif. This short N-terminal sequence is essential for DNA binding and we suggest it is involved in maintaining the three-dimensional structure of the first finger domain, as has been seen in the nuclear magnetic resonance structure of one of the zinc-finger domains of the yeast transcription factor SW15. The characterization of the DNA-binding activity of this 66 residue peptide (delta 911zf) shows that it binds in a sequence-specific manner, as a monomer, to a natural target site with an apparent KD approximately 4 x 10(-7) M. The shortest delta 911zf binding site, which retains full affinity, consists of an 11 base-pair sequence with a one nucleotide overhang at each 5' end. DNase I, hydroxyl radical and methylation protection footprinting studies show that, in common with other zinc-finger proteins, delta 911zf binds in the major groove of DNA. The data presented are consistent with the zinc-fingers of Tramtrack contacting both strands of the DNA, and thus the binding differs in detail to that observed in the crystal structure of the three zinc-fingers of Zif268 complexed to their target DNA.

Amino Acid Sequence↗
PubMed1992

DNA curvature does not require bifurcated hydrogen bonds or pyrimidine methyl groups.

Short tracts of the homopolymer dA.dT confer intrinsic curvature on the axis of the DNA double helix. This phenomenon is assumed to be a consequence of such tracts adopting a stable B'-DNA conformation that is distinct from B-form structure normally assumed by other DNA sequences. The more stable B' structure of dA.dT tracts has been attributed to several possible stabilizing factors: (1) optimal base stacking interactions consequent upon the high propeller twist, (2) bifurcated hydrogen bonds between adjacent dA.dT base-pairs, (3) stacking interactions involving the dT methyl groups, and finally (4) a putative spine of ordered water molecules in the minor groove. DNA oligodeoxynucleotides have been synthesized that enable these hypotheses to be tested; of particular interest is the combination of effects due to bifurcation (2) and methylation of the pyrimidines nucleotides (3). The data indicate that neither bifurcated hydrogen bonds nor pyrimidine methyl groups nor both are essential for DNA curvature. The data further suggest that the influence of the minor groove spine of hydration on the B'-formation is small. The experiments favor the hypothesis that base stacking interactions are the dominant force in stabilizing the B'-form structure.

Base Sequence↗
PubMed1992

DNA deformation in nucleoprotein complexes between RNA polymerase, cAMP receptor protein and the lac UV5 promoter probed by singlet oxygen.

Singlet oxygen (1O2), generated by exciting an eosin-Tris complex with a high intensity beam of radiation at 532 nm, was used to chemically modify bases in fragments of DNA containing the lac UV5 promoter in the presence of the DNA binding proteins, RNA polymerase and CRP (cAMP receptor protein). Subsequent treatment with piperidine selectively cleaved the DNA at specific modified bases in the sequence. Using this technique we show first that the reactivity of DNA bound by CRP differs in the presence and absence of RNA polymerase. Hence the local conformation of CRP-bound DNA must change during the transition to the open complex. However, no reactivity is observed at the sites of the 40 degrees kinks described in the cocrystal structure (Steitz, 1990). Secondly we show that there is unique CRP-dependent reactivity at a specific site (position -46 on the upper strand) in the open complex. Finally, in the open complex, 1O2 also reacts with sites 90 bp upstream from the transcription start point. This reactivity is qualitatively CRP-independent. We infer that 1O2 reacts at sites where the promoter DNA is significantly distorted, and suggest that the pattern observed reflects the functional orientation of an active transcriptional complex in which the DNA is bent to form an extended loop.

Cyclic AMP Receptor Protein↗