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T Moss

Publications and source records attributed to T Moss.

At least 19 recordsLinked to original sources

Variants of the Xenopus laevis ribosomal transcription factor xUBF are developmentally regulated by differential splicing.

XUBF is a Xenopus ribosomal transcription factor of the HMG-box family which contains five tandemly disposed homologies to the HMG1 & 2 DNA binding domains. XUBF has been isolated as a protein doublet and two cDNAs encoding the two molecular weight variants have been characterised. The major two forms of xUBF identified differ by the presence or absence of a 22 amino acid segment lying between HMG-boxes 3 and 4. Here we show that the mRNAs for these two forms of xUBF are regulated during development and differentiation over a range of nearly 20 fold. By isolating two of the xUBF genes, it was possible to show that both encoded the variable 22 amino acid segment in exon 12. Oocyte splicing assays and the sequencing of PCR-generated cDNA fragments, demonstrated that the transcripts from one of these genes were differentially spliced in a developmentally regulated manner. Transcripts from the second gene were found to be predominantly or exclusively spliced to produce the lower molecular weight form of xUBF. Expression of a high molecular weight form from yet a third gene was also detected. Although the intron-exon structures of the Xenopus and mouse UBF genes were found to be essentially identical, the differential splicing of exon 8 found in mammals, was not detected in Xenopus.

Amino Acid Sequence

High resolution studies of the Xenopus laevis ribosomal gene promoter in vivo and in vitro.

The first high resolution maps of the Xenopus laevis ribosomal promoter and its flanking regions (-179 to +14) have been created by assaying point mutants both in oocyte and in vitro. Within the promoter boundaries (-141(-145) to +3(+4)), domains analogous to the Core Promoter and "Upstream Control Element" (UCE) were clearly detected. The base pairs at -133, within the UCE, and -20, -10, -7, and +3, within the Core, were all shown to be especially important for promotion. Between the Core and UCE, two central promoter elements (CPEs) were also resolved. Surprisingly, these CPEs did not correspond to the highly conserved enhancer homology (approximately -70 to -110), but to CCCGGCC motifs immediately flanking it. Although xUBF was shown to be a limiting component for in vitro transcription, none of the point mutations was found to affect the interaction of this factor with the promoter.

Animals

Readthrough enhancement and promoter occlusion on the ribosomal genes of Xenopus laevis.

An RNA polymerase I termination site is found just upstream of the ribosomal gene promoter in mammals and amphibia. It has been suggested that this termination site may actively enhance ribosomal transcription in a process known as readthrough enhancement or that it may simply prevent the disruption of initiation complexes or promoter occlusion. There is, however, a consensus of opinion that the terminator is important for efficient ribosomal transcription. Here we have quantitatively investigated the relative importance of readthrough enhancement and promoter occlusion on the transcription of the microinjected Xenopus laevis ribosomal gene. The results show that, in this system, promoter occlusion is limited and terminator mutations predominantly affect readthrough enhancement. The terminator is shown to be unnecessary for the enhancer activity of the rest of the ribosomal spacer. Model calculations suggest that readthrough enhancement could be explained by polymerase recycling and that it may be unnecessary to postulate a specific mechanism of polymerase handover.

Animals

Heterogeneity in the Xenopus ribosomal transcription factor xUBF has a molecular basis distinct from that in mammals.

The Xenopus polymerase I transcription factor xUBF is an HMG-box protein which has been purified as two polypeptides of approximately 82 and 85 kDa. Recently a cDNA sequence predicted an xUBF protein (xUBF1) of 677 amino acids (79 kDa) containing five tandem HMG-boxes. Here a second and distinct xUBF cDNA has been isolated and characterised. This cDNA codes an xUBF protein (xUBF2) of 701 amino acids (82 kDa), having 93% homology with xUBF1 but containing an insertion of 22 amino acids between HMG-boxes 3 and 4. In vitro translation of synthetic mRNAs derived from the xUBF1 and 2 cDNAs was used to show that the electrophoretic mobility of the gene products accounted for the major xUBF molecular weight heterogeneity noted in vivo. It is also shown that the Xenopus laevis genome contains 3 or 4 distinct loci which hybridise with xUBF coding sequences, leaving open the possibility of yet further unrecognised heterogeneity in xUBF.

Amino Acid Sequence

The RNA polymerase I transcription factor xUBF contains 5 tandemly repeated HMG homology boxes.

The RNA polymerase I transcription factor UBF has been identified in human, mouse, rat and Xenopus and the primary structure of the human protein has been determined. Human UBF was shown to contain four tandem homologies to the folding domains of the HMG1 and 2 proteins and hence to belong to a previously unrecognised family of 'HMG-box' transcription factors. Here, cDNA clones encoding the Xenopus laevis UBF (xUBF) have been isolated and sequenced. Northern and Southern blots revealed that in tissue culture cells, xUBF is coded on a single major mRNA size species by a small number of genes. The deduced primary structure of xUBF is highly homologous with the human protein except for a central deletion which removes most of one HMG-box. This explains the major size difference between the X. laevis and human proteins and may well explain their different transcriptional specificities. It is shown that xUBF contains 5 tandemly repeated HMG-boxes and that by analogy the human protein contains 6.

Amino Acid Sequence

The value of autopsies in neurosurgery.

Many previous studies have reported the value of autopsy in assessing clinical diagnostic accuracy. None of them however, assessed the value of autopsies in a specific clinical speciality. The authors reviewed the findings of 123 consecutive neurosurgical autopsies with reference to the premortem clinical diagnoses. The study showed that 7% of cases had a wrong clinical diagnosis and in 9% of cases the clinical diagnosis was incomplete. Only in 5% of all cases knowledge of the autopsy findings would have led to a change in management and outcome. The autopsies also confirmed that 11% of cases died following a surgical complication and in 3% of cases the primary cause of death was non-neurosurgical. The latter was a previously unrecognised finding in 8% of autopsies. The autopsy will remain a valuable means of clinical audit and the increasing financial pressures to reduce the number of autopsies should be resisted.

Adolescent

Immunocytochemical evidence of lymphocytic derivation of neoplastic cells in malignant angioendotheliomatosis.

Neoplastic angioendotheliomatosis is a rare disorder usually characterized by primarily cutaneous or neurological symptoms. Approximately 40 cases of malignant angioendotheliomatosis with primary central nervous system (CNS) symptoms have been reported. Some investigators have postulated a hematopoietic origin for this neoplasm. Most of the literature, however, has perpetuated the idea that the often bizarre symptoms seen with this entity result from neoplastic endothelial cell proliferation within the small vessels of affected organs, including the brain and spinal cord. This report describes the immunohistochemical examination and confirmation of the cell of origin of this neoplasm based on five previously unpublished cases of malignant angioendotheliomatosis with primarily CNS symptoms. It includes the first documentation of a T-cell lymphoma presenting as malignant angioendotheliomatosis. All cases include autopsy findings, and in four cases the diagnosis was made postmortem. One case was proven by stereotactic biopsy, but the patient succumbed as a result of severe intracranial bleeding that occurred at the time of biopsy. Tissues were studied with avidin-biotin peroxidase immunohistochemical techniques using a panel of monoclonal antibodies directed against the leukocyte common antigen, LN-1, LN-2, and anti-Factor VIII, and also using Ulex europaeus agglutinin 1. Based on the results obtained, the authors conclude that the proliferative cells seen within the vessel lumina are of lymphocytic origin and agree that the condition should more properly be designated intravascular lymphomatosis. The therapeutic implications of this conclusion point to the possible administration of chemotherapy and radiotherapy in an effort to achieve remissions in an otherwise relentlessly progressive neurological disorder.

Adult

Shed tumor gangliosides and progression of human neuroblastoma.

Shedding of membrane gangliosides is characteristic of human and experimental tumors. Because some shed tumor gangliosides have potent tumor-enhancing properties, significant ganglioside shedding could influence tumor progression. We examined this possibility in a human tumor, neuroblastoma. Ganglioside shedding, measured as circulating tumor-derived GD2 ganglioside, and the outcome of 74 patients with advanced stage (III and IV) disease were studied. Progression-free survival (PFS) was inversely related to circulating GD2 levels at the time of diagnosis (P = .018). By Kaplan-Meier analysis, the quartile of patients having the highest circulating GD2 levels (greater than or equal to 568 pmol/mL) had a strikingly different outcome from the quartile of patients with the lowest (less than or equal to 103 pmol/mL) GD2 levels (P = .013): median PFS was shorter (9 v 28 months), and the long-term survival rate lower (2-year PFS of 24% v 70%). We conclude that more rapid disease progression and lower survival rate are associated with high circulating GD2 levels at diagnosis and speculate that shed neuroblastoma tumor gangliosides play a role in accelerating tumor progression.

Biomarkers, Tumor

Point mutation analysis of the Xenopus laevis RNA polymerase I core promoter.

The core region of the Xenopus laevis pre-ribosomal RNA promoter was subjected to point mutation analysis. A total of 27 point mutants within a 78 base pair region from -64 to +14, (relative to the start of transcription at +1), were assayed by oocyte microinjection. The results locate the 3' boundary of the core promoter at +4 and the 5' boundary at between -33 and -39 and suggest that this region of the Xenopus promoter is generally similar in organisation to mammalian core promoters. In particular, the conserved guanidine nucleotides at -7 and -16 are clearly essential for promoter function. The data suggest that interactions between the transcription machinery and the promoter occur in four distinct regions around +2 to +4, -7, -17 to -20 and -28 to -33. This particular periodicity of functionally important nucleotides is consistent with a model in which all protein-DNA interactions take place from predominantly one side of the DNA helix.

Animals

Comparison of three silver stains for demonstrating neurofibrillary tangles and neuritic plaques in brain tissue stored for long periods.

Three methods were compared to find a reliable method for demonstrating neurofibrillary tangles (NFTs) and neuritic plaques (NPs) in brain tissue stored for long periods in formalin or as paraffin blocks. The short-term fixation of tissue, e.g. up to 6 months in formalin does not usually present a problem using any of the three methods tried, e.g. Gallyas, modified Palmgren, or modified Bielschowsky, but once the time lengthens to 6 years or more the demonstration of NFTs and NPs is not so reliable using the first two methods. The modified Bielschowsky method, however, demonstrates well both NFTs and NPs in material stored in formalin or as paraffin blocks for long periods, e.g. 7 years, and also compares favourably with the other methods on freshly processed material and fixed tissue, stored for shorter periods. We also noted as a consistent trend, irrespective of the staining technique employed, the detection of fewer plaques and tangles in material stored in formalin, as opposed to that stored as blocks embeded in paraffin wax.

Aged

Coordinate replication of dispersed repetitive sequences in Physarum polycephalum.

The synchronous macroplasmodial growth phase of the slime mould Physarum polycephalum was used to study the in vivo replication of large chromosomal DNA segments. Newly replicated DNA was isolated at various points in S-phase by its preferential association with the nuclear matrix. This DNA was then used to probe cosmid clones of the Physarum genome. The results indicate that certain dispersed repetitive sequences in the genome are coordinately replicated. The observed pattern of replication may be due either to the presence of a replication origin within each repetitive sequence or to the systematic arrangement of these sequences around a replication origin. The latter appears more likely since the repetitive sequences are probably not randomly scattered within the genome.

Cell Nucleus

Cerebral distribution of immunoconjugate after treatment for neoplastic meningitis using an intrathecal radiolabeled monoclonal antibody.

A detailed autopsy and autoradiographic study was performed after the death of a patient undergoing intrathecal, antibody-guided irradiation for carcinomatous meningitis. The results demonstrated tumor cells infiltrating the surface meninges and a severe astrocytic reaction associated with oedema in the periventricular and brain stem subpial white matter. This was not seen in cortical or other gray matter structures. Autoradiographic examination correlated well, demonstrating isotope within the oedematous areas of the white matter in addition to the expected concentration in the leptomeningeal layers. These findings are discussed in the context of antibody binding to tumor tissue and the possible benefits conferred in the treatment of infiltrating tumor cells.

Antibodies, Monoclonal

The Xenopus laevis ribosomal gene terminator contains sequences that both enhance and repress ribosomal transcription.

A DNA segment approximately 200 base pairs upstream of the Xenopus laevis ribosomal promoter acts both as an upstream promoter element that augments transcription and as a transcription terminator. It is, however, unclear to what extent these two activities are related. A segment of the X. laevis ribosomal DNA, containing the terminator and the upstream promoter element, was subjected to point mutation, and the effects of the resulting mutations were investigated by oocyte microinjection. Analysis of 26 point mutants revealed not only sequences that augment 40S transcription but also those that repress it. The sequences that augmented transcription lay within the T3 homology box and also near the site of 3'-end formation. These sequences also played a role in termination. The sequences that repressed transcription lay within the G+C-rich DNA flanking the T3 box. It can be concluded that termination is probably essential but may not be sufficient for the activity of the upstream promoter element.

Animals

The enhancement of ribosomal transcription by the recycling of RNA polymerase I.

It has been suggested that the tandemly repeated ribosomal genes of eukaryotes may be subject to a special mechanism of transcriptional enhancement, called Readthrough Enhancement, in which transcription factors are recycled. Recent experiments with the mouse ribosomal genes, although consistent with this possibility, were unable to distinguish between true Readthrough Enhancement and promoter occlusion. To test directly for Readthrough Enhancement, the pre-ribosomal RNA of Xenopus laevis was prematurely terminated within the 18S gene on a circular template. This premature termination was found to reduce the efficiency of pre-ribosomal RNA promotion in cis by 80 to 90%. Since the pre-ribosomal RNA is normally terminated only 213 base pairs upstream of its own initiation site, the results strongly suggest that the recycling of RNA polymerase, or Readthrough Enhancement, is a means by which ribosomal transcription is enhanced in Xenopus laevis.

Animals

A complex array of sequences enhances ribosomal transcription in Xenopus laevis.

The ribosomal DNA spacer in Xenopus laevis was shown in previous studies to be involved in regulating the expression of the ribosomal genes. Here transcription enhancement by this spacer has been studied in some detail, to fully identify the sequences involved and to determine their relative importance in this phenomenon. It is shown that the 60/81 base-pair (bp) repeats, which were reported to be enhancer elements, act as part of a mode of enhancement whose effect is amplified by the spacer promoters or Bam islands. The "Bam super repeat", a combination of spacer promoter and 60/81 bp elements, is the major enhancer unit. Within a Bam super repeat, a near linear correlation between the number of 60/81 bp elements and enhancer activity is observed. Thus, there is no significant co-operativity in the binding of transcription factors to an array of these elements. Multiple Bam super repeats do not act additively and may actually interfere with each others action. Surprisingly this effect is observed both in the presence and absence of active spacer promoters. Sequences between the 3' end of the 28 S coding region and the first spacer promoter may also be involved in enhancement but only in a very minor fashion. In confirmation of recent studies, the presence of the unique ribosomal termination sequence, 213 bp upstream from the pre-rRNA initiation site, is essential for efficient promotion, as deletion of this sequence virtually abolishes pre-rRNA- transcription. These data are discussed in terms of the possible mechanisms of transcription enhancement.

Animals

X-linked bulbo-spinal neuronopathy: a family study of three patients.

The clinical features of two brothers and one nephew with X-linked recessive bulbo-spinal neuronopathy are described. The neurophysiological investigations and sural nerve biopsy, previously unreported, confirmed that both motor and sensory nerves are affected. Because of the genetic implications, the importance is stressed of recognising this disorder as a separate entity which should not be classified with the spinal muscular atrophies.

Adult