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S M Burgess

Publications and source records attributed to S M Burgess.

15 recordsLinked to original sources
PubMed2004

Integration target site selection for retroviruses and transposable elements.

When a retrovirus infects a cell, its RNA genome is reverse transcribed into a double-stranded DNA, which is then permanently integrated into the host chromosome. Integration is one of the essential steps in the retroviral life cycle. Many transposable elements also move around and integrate into the host genome as part of their life cycle, some through RNA intermediates and some through 'cut and paste' mechanisms. Integration of retroviruses and transposable elements into 'sensitive areas' of the genome can cause irreparable damage. On the other hand, because of their ability to integrate permanently, and the relatively efficient rates of transgenesis, retroviruses and transposable elements are widely used as gene delivery tools in basic research and gene therapy trials. Recent events in gene therapy treatments for X-linked severe combined immunity deficiencies (X-SCID) have highlighted both the promise and some of the risks involved with utilizing retroviruses. Nine of 11 children were successfully treated for X-SCID using a retrovirus carrying the gene mutated in this disease. However, later two of these children developed leukemias because of retroviral integrations in the putative oncogene LMO2 [1]. A third child has also been demonstrated to have an integration in LMO2, but is as of yet nonsymptomatic [2]. It is a bit difficult to explain the high frequency of integrations into the same gene using a random model of retroviral integration, and there has been evidence for decades that retroviral integrations may not be random. But the data were somewhat limited in their power to determine the precise nature of the integration biases. The completion of the human genome sequence coupled with sensitive polymerase chain reaction techniques and an ever-decreasing cost of sequencing has given a powerful new tool to the study of integration site selection. In this review, we describe the findings from several recent global surveys of target site selection by retroviruses and transposable elements, and discuss the possible ramifications of these findings to both mechanisms of action and to the use of these elements as gene therapy vectors.

Animals↗
PubMed2001

Substrate specificity and excision kinetics of Escherichia coli endonuclease VIII (Nei) for modified bases in DNA damaged by free radicals.

Endonuclease VIII (Nei) is one of three enzymes in Escherichia coli that are involved in base-excision repair of oxidative damage to DNA. We investigated the substrate specificity and excision kinetics of this DNA glycosylase for bases in DNA that have been damaged by free radicals. Two different DNA substrates were prepared by gamma-irradiation of DNA solutions under N(2)O or air, such that they contained a multiplicity of modified bases. Although previous studies on the substrate specificity of Nei had demonstrated activity on several pyrimidine derivatives, this present study demonstrates excision of additional pyrimidine derivatives and a purine-derived lesion, 4,6-diamino-5-formamidopyrimidine, from DNA containing multiple modified bases. Excision was dependent on enzyme concentration, incubation time, and substrate concentration, and followed Michaelis-Menten kinetics. The kinetic parameters also depended on the identity of the individual modified base being removed. Substrates and excision kinetics of Nei and a naturally arising mutant form involving Leu-90-->Ser (L90S-Nei) were compared to those of Escherichia coli endonuclease III (Nth), which had previously been determined under experimental conditions similar to those in this study. This comparison showed that Nei and Nth significantly differ from each other in terms of excision rates, although they have common substrates. The present work extends the substrate specificity of Nei and shows the effect of a single mutation in the nei gene on the specificity of Nei.

DNA Damage↗
PubMed1999

Collisions between yeast chromosomal loci in vivo are governed by three layers of organization.

The relative probabilities that different pairs of chromosomal loci will collide with one another in vegetatively growing diploid yeast cells have been assessed using a genetic assay for Cre/loxP site-specific recombination. Recombination rates have been determined for 18 different pairs of loxP sites representing diverse pairs of positions within the genome. Overall, relative collision probabilities vary over an eightfold range. Within this range, a hierarchy comprising three levels of organization can be discerned. First, collisions between loci on nonhomologous chromosomes are governed by nonspecific centromere clustering. Second, a sequence is closer to allelic or nearby sequences on its homolog than to sequences on nonhomologous chromosomes, an effect most simply attributed to homolog pairing. Third, a sequence can be closer to other sequences nearby on the same chromosome than to sequences on other chromosomes. These findings provide a framework for assessing the role of chromosome disposition in cellular processes such as DNA repair and gene expression. Also the possibility is raised that genome-wide coalignment of homologs is not the fundamental raison d'etre of the somatic pairing process. We suggest instead that pairing may exist to promote juxtaposition of homologous regions within irregular genome complements.

Base Sequence↗
PubMed1999

Somatic pairing of homologs in budding yeast: existence and modulation.

FISH analysis of well-spread chromosomes reveals that homologs are paired in vegetatively growing budding yeast diploid cells, via multiple interstitial interactions, and independent of recA homologs and mating type heterozygosity. Pairing is present during G1 and G2, and in cells arrested at G1 by mating pheromone, but is disrupted during S phase. Thus, somatic pairing is qualitatively analogous to premeiotic and early meiotic pairing. S-phase pairing disruption occurs by a complex intranuclear program involving regional, nucleus-wide, and temporal determinants. Pairing is also disrupted in two G2-arrest conditions (cdc13ts and nocodazole). Together these findings suggest that cell cycle signals may provoke pairing disruption by modulating underlying chromosome and/or chromatin structure. Whether the cell chooses to disrupt pairing contacts or not (e.g., S phase and G2 arrest, but not G1 arrest or normal G1 or G2), could be dictated by functional considerations involving homolog/sister discrimination.

Animals↗
PubMed1999

GCN5-dependent histone H3 acetylation and RPD3-dependent histone H4 deacetylation have distinct, opposing effects on IME2 transcription, during meiosis and during vegetative growth, in budding yeast.

Diploid yeast undergo meiosis under certain conditions of nutrient limitation, which trigger a transcriptional cascade involving two key regulatory genes. IME1 is a positive activator of IME2, which activates downstream genes. We report that Gcn5, a histone H3 acetylase, plays a central role in initiation of meiosis via effects on IME2 expression. An allele, gcn5-21, was isolated as a mutant defective in spore formation. gcn5-21 fails to carry out meiotic DNA replication, recombination, or meiotic divisions. This mutant also fails to induce IME2 transcription; IME1 transcription, however, is essentially normal. Further investigation shows that during wild-type meiosis the IME2 promoter undergoes an increase in the level of bound acetylated histone H3. This increase is contemporaneous with meiotic induction of IME2 transcription and is absent in gcn5-21. In contrast, the RPD3 gene, which encodes a histone H4 deacetylase and is known to be required for repression of basal IME2 transcription in growing yeast cells, is not involved in induction of IME2 transcription or IME2 histone acetlyation during meiosis. These and other results suggest that Gcn5 and Rpd3 play distinct roles, modulating transcription initiation in opposite directions under two different cellular conditions. These roles are implemented via opposing effects of the two gene products on acetylation of two different histones. Finally, we find that gcn5 and rpd3 single mutants are not defective in meiosis if acetate is absent and respiration is promoted by a metabolically unrelated carbon source. Perhaps intracellular acetate levels regulate meiosis by controlling histone acetylation patterns.

Acetylation↗
PubMed1999

The cytomorphology of papillary serous carcinoma of the endometrium in cervical smears.

BACKGROUND: An investigation into the determination of cytomorphologic criteria that may distinguish papillary serous carcinoma of the endometrium (PSC) from typical endometrioid carcinoma (TEC) in cervical smears was undertaken. Preoperative identification of this poor prognostic variant of endometrial carcinoma may influence the surgical management of these cases and the choice of adjuvant therapy. METHODS: The cervical smears of 12 cases of histologically confirmed PSC; 12 cases of TEC, including 2 villoglandular/papillary variants, and 6 cases of mixed PSC and papillary endometrioid carcinoma were reviewed. In all cases an initial diagnosis of malignancy had been made on the cervical smears. Twenty-seven criteria were evaluated and the relation between the cytologic characteristics and the type of adenocarcinoma, the degree of association, and the intergroup homogeneity were tested. RESULTS: Features strongly associated with PSC were hypercellular smears with a background tumor diathesis, papillae, bare nuclei, and cells with large pleomorphic nuclei and bulky dense cytoplasm. In contrast, TEC showed a relatively monomorphic population of cells with moderately enlarged oval nuclei and delicate cytoplasm. In the mixed tumors, the features were similar to those of PSC, suggesting preferential exfoliation of the PSC component of the tumor. CONCLUSIONS: Assessment of the cytomorphology of PSC and TEC of the endometrium in Papanicolaou stained cervical smears is possible using statistically significant diagnostic criteria.

Carcinoma, Endometrioid↗
PubMed1998

Early health-related behaviours and their impact on later life chances: evidence from the US.

This paper uses evidence from the US to examine the impact of adolescent illegal consumption and violent behaviour on later life chances. Specifically, we look at the effect of such behaviour by young men in late adolescence on productivity and household formation 10 years on. We find that alcohol and soft drug consumption have no harmful effects on economic prospects in later life. In contrast, hard drug consumption and violent behaviour in adolescence are both associated with lower productivity even by the time the individuals are in their late twenties. These effects are substantial and affect earnings levels and earnings growth. These results are robust to the inclusion of a rich set of additional controls measuring aspects of the individuals' backgrounds. However, we find no evidence of any of these behaviours significantly affecting household formation.

Adolescent↗
PubMed1998

not really finished is crucial for development of the zebrafish outer retina and encodes a transcription factor highly homologous to human Nuclear Respiratory Factor-1 and avian Initiation Binding Repressor.

Not really finished (nrf), a larval-lethal mutation in zebrafish generated by retroviral insertion, causes specific retinal defects. Analysis of mutant retinae reveals an extensive loss of photoreceptors and their precursors around the onset of visual function. These neurons undergo apoptosis during differentiation, affecting all classes of photoreceptors, suggesting an essential function of nrf for the development of all types of photoreceptors. In the mutant, some photoreceptors escape cell death, are functional and, as judged by opsin expression, belong to at least three classes of cones and one class of rods. The protein encoded by nrf is a close homologue of human Nuclear Respiratory Factor 1 and avian Initiation Binding Repressor, transcriptional regulators binding the upstream consensus sequence RCGCRYGCGY. At 24 hours of development, prior to neuronal differentiation, nrf is expressed ubiquitously throughout the developing retina and central nervous system. At 48 hours of development, expression of nrf is detected in the ganglion cell layer, in the neurons of the inner nuclear layer, and in the optic nerve and optic tracts, and, at 72 hours of development, is no longer detectable by in situ hybridization. Mutants contain no detectable nrf mRNA and die within 2 weeks postfertilization as larvae with reduced brain size. On the basis of its similarity with NRF-1 and IBR, nrf is likely involved in transcriptional regulation of multiple target genes, including those that encode mitochondrial proteins, growth factor receptors and other transcription factors. This demonstrates the power of insertional mutagenesis as a means for characterizing novel genes necessary for vertebrate retinal development.

Amino Acid Sequence↗
PubMed1994

MMM1 encodes a mitochondrial outer membrane protein essential for establishing and maintaining the structure of yeast mitochondria.

In the yeast Saccharomyces cerevisiae, mitochondria are elongated organelles which form a reticulum around the cell periphery. To determine the mechanism by which mitochondrial shape is established and maintained, we screened yeast mutants for those defective in mitochondrial morphology. One of these mutants, mmm1, is temperature-sensitive for the external shape of its mitochondria. At the restrictive temperature, elongated mitochondria appear to quickly collapse into large, spherical organelles. Upon return to the permissive temperature, wild-type mitochondrial structure is restored. The morphology of other cellular organelles is not affected in mmm1 mutants, and mmm1 does not disrupt normal actin or tubulin organization. Cells disrupted in the MMM1 gene are inviable when grown on nonfermentable carbon sources and show abnormal mitochondrial morphology at all temperatures. The lethality of mmm1 mutants appears to result from the inability to segregate the aberrant-shaped mitochondria into daughter cells. Mitochondrial structure is therefore important for normal cell function. Mmm1p is located in the mitochondrial outer membrane, with a large carboxyl-terminal domain facing the cytosol. We propose that Mmm1p maintains mitochondria in an elongated shape by attaching the mitochondrion to an external framework, such as the cytoskeleton.

Amino Acid Sequence↗
PubMed1993

A mechanism to enhance mRNA splicing fidelity: the RNA-dependent ATPase Prp16 governs usage of a discard pathway for aberrant lariat intermediates.

PRP16 encodes an RNA-dependent ATPase required for the second step of mRNA splicing in S. cerevisiae. We have isolated seven alleles of PRP16 that, like the original allele prp16-1, allow splicing of introns with a mutant branch site (UACUAAC to UACUACC), by forming lariat intermediates at the mutant C nucleotide. Every suppressor mutation maps to the region of PRP16 common to RNA-dependent ATPases. We purified three of the mutant proteins and found that all exhibit reduced ATPase activity, as does Prp16-1. An in vivo analysis of the steady-state levels of the splicing intermediates and products provides evidence for a pathway, under the genetic control of PRP16, to discard incorrectly branched substrates. We propose that decreasing the rate of ATP hydrolysis by Prp16 allows aberrantly formed lariat intermediates more time to proceed through the productive rather than the discard branch of this pathway.

Adenosine Triphosphatases↗
PubMed1993

Beat the clock: paradigms for NTPases in the maintenance of biological fidelity.

Kinetic proofreading is a strategy used by cells to reduce the number of errors made when replicating and expressing genetic information. Recent advances in mRNA splicing suggest a variation on the theme of previously described kinetic proofreading mechanisms, which may apply to other multicomponent assembly processes in the cell.

Acid Anhydride Hydrolases↗
PubMed1986

beta-Glucuronidase from Escherichia coli as a gene-fusion marker.

We have developed a gene-fusion system based on the Escherichia coli beta-glucuronidase gene (uidA). The uidA gene has been cloned from E. coli K-12 and its entire nucleotide sequence has been determined. beta-Glucuronidase has been purified to homogeneity and characterized. The enzyme has a subunit molecular weight of 68,200, is very stable, and is easily and sensitively assayed using commercially available substrates. We have constructed gene fusions of the E. coli lacZ promoter and coding region with the coding region of the uidA gene that show beta-glucuronidase activity under lac control. Plasmid vectors have been constructed to facilitate the transfer of the beta-glucuronidase coding region to heterologous control regions, using many different restriction endonuclease cleavage sites. There are several biological systems in which uidA-encoded beta-glucuronidase may be an attractive alternative or complement to previously described gene-fusion markers such as beta-galactosidase or chloramphenicol acetyltransferase.

Base Sequence↗
PubMed1977

The developmental changes in the placenta of the guinea-pig.

The placental disc and the yolk sac endoderm of intact guinea-pigs from day 14 of gestation to term, and of ovariectomized and fetectomized animals at 35 days of pregnancy, were observed by histological means. The chorionic giant cells were the first to attain maximal development (between days 17-35) and they had begun to degenerate by day 40. The spongy zone syncytiotrophoblast was well established and vascularized by day 18, and the maximal size of the zone was maintained between days 30 and 64. The spongy zone then dwindled in size just before parturition. Labyrinthine tissue appeared on day 18, but rapid growth and complete vascularization was not attained until as late as day 40. However, its continued increase in size almost up to term closely paralleled the growth of the fetus. If the placenta secretes progesterone from day 18 to parturition, the spongy zone is the only tissue sufficiently well established to carry out this activity throughout this period. The compensatory hypertrophy of the spongy zone syncytiotrophoblast in 35 day pregnant animals after ovariectomy and fetectomy supports the hypothesis that the spongy tissue secretes progesterone.

Animals↗