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D Stott

Publications and source records attributed to D Stott.

31 records · Page 2Linked to original sources
PubMed1996

The human homolog T of the mouse T(Brachyury) gene; gene structure, cDNA sequence, and assignment to chromosome 6q27.

We have cloned the human gene encoding the transcription factor T. T protein is vital for the formation of posterior mesoderm and axial development in all vertebrates. Brachyury mutant mice, which lack T protein, die in utero with abnormal notochord, posterior somites, and allantois. We have identified human T genomic clones and derived the mRNA sequence and gene structure. There is 91% amino acid identity between human and mouse T proteins overall and complete identity across 77 amino acids of the T-box motif within the DNA-binding domain. Human T expression is very similar to that found for T in other vertebrate species and is confined to cells derived from the notochord. The human T gene maps to chromosome 6q27 and is only the second human member of the T-box gene family to be described.

Amino Acid Sequence↗
PubMed1993

Rescue of the tail defect of Brachyury mice.

The mouse Brachyury (T) gene is required for normal development of axial structures. Embryos homozygous for the T mutation show severe deficiencies in mesoderm formation. They lack the notochord and allantois, have abnormal somites, and die at approximately 10 days postcoitum probably as a result of the allantois defect. Mice heterozygous for the T mutation exhibit a variable short-tailed phenotype. The T gene has been cloned and shown to be expressed in the tissues most strongly affected by the mutation. In this paper, we show that a single-copy transgene representing the wild-type T allele is able to rescue the T-associated tail phenotype. In addition, we show that increasing dosage of the T gene in Tc/+ mice causes an increased extension of the axis. These data show the correlation of the level of T product with the extension of the anteroposterior axis, directly demonstrating the involvement of the T product in this process.

Allantois↗
PubMed1991

Multiple spatially specific enhancers are required to reconstruct the pattern of Hox-2.6 gene expression.

Murine Hox genes are organized into four clusters that share many features with the homeotic clusters of Drosophila. This evolutionary conservation and the clear relationships between the position of a gene within a cluster and its expression pattern have led to the suggestion that the structure of the cluster is essential for proper regulation. Using a Hox-2.6-lacZ reporter gene in transgenic mice we have shown that the overall expression pattern of the endogenous Hox-2.6 gene can be reconstructed when it is isolated from the complex. The transgene was expressed in the proper tissues, with the correct spatial distribution and temporal pattern. Furthermore, direct comparison by in situ hybridization revealed that the levels of transgene expression are similar to those of the endogenous gene. This has allowed us to define three elements that regulate particular aspects of the Hox-2.6 pattern, two of which act as spatially specific enhancers. One enhancer, region A, directed expression only in the neural tube, whereas the other, region C, specified the majority of the Hox-2.6 pattern. Both were also capable of imposing the correct boundaries of expression on heterologous promoters. The definition of such elements will allow the characterization of the trans-acting factors that mediate spatial regulation in the mammalian embryo.

Animals↗
PubMed1991

Sequences and factors required for the F9 embryonal carcinoma stem cell E1a-like activity.

F9 embryonal carcinoma (EC) stem cells contain an E1a-like activity that is absent from differentiated derivatives. We have previously characterized proteins present in F9 EC cell extracts that bind to the E1a-dependent E2A promoter and have shown that two of them, TF68 and DRTF1, are required for efficient transcription in vitro (N. B. La Thangue, B. Thimmapaya, and P. W. J. Rigby, Nucleic Acids Res. 18:2929-2938, 1990). We now show that the E1a-like activity is detectable in transient transfection assays. Deletion mutations show that a distal sequence element, which includes the ATF/CREB consensus, is required for expression in both cell types, although it does not mediate the down-regulation of promoter activity that accompanies differentiation. A series of point mutations generated by in vitro mutagenesis confirm this and show that sequences around -60 are necessary for efficient expression in stem cells but not in differentiated derivatives. These sequences bind DRTF1, the activity of which is strongly down-regulated during differentiation. Surprisingly, mutations in a previously uncharacterized region of the promoter restore activity to a promoter carrying the -60 mutation and lead to the formation of a new DNA-protein complex.

Adenovirus Early Proteins↗
PubMed1990

Regulation of RNA polymerase III transcription in response to Simian virus 40 transformation.

Transcription by RNA polymerase III of the B2 family of middle-repetitive elements is activated in response to transformation by a variety of agents, including DNA tumour viruses, RNA tumour viruses and chemical carcinogens. We have investigated the mechanism of activation in SV40-transformed cells and we find that the effect is due to an increase in the activity of the general class III transcription factor TFIIIC, achieved both by an increase in factor abundance and by a change in its phosphorylation state. SV40 transformation also stimulates transcription of other genes by RNA polymerase III but the effect may be balanced by compensatory post-transcriptional changes. TFIIIC may mediate the stimulation of polymerase III transcription by a range of transforming viruses.

Animals↗
PubMed1989

Regulation of RNA polymerase III transcription in response to F9 embryonal carcinoma stem cell differentiation.

B2 genes are rodent-specific middle repetitive elements transcribed by RNA polymerase III. They are expressed in the ectoderm and mesoderm but not in the embryonic or extraembryonic endoderm of early mouse embryos. This tissue specificity is mimicked in vitro by embryonal carcinoma and embryonic stem cell lines. Nuclear run-on experiments show that the down-regulation of B2 genes during F9 embryonal carcinoma cell differentiation into endoderm occurs at the transcriptional level and that other class III genes, including those encoding tRNA, show a similar response. We have used cell-free extracts to investigate the molecular mechanisms responsible. The specific down-regulation of transcription by RNA polymerase III during F9 cell differentiation is due to a reduction in the activity of the general class III transcription factor TFIIIB.

Animals↗
PubMed1987

Studies on the migration of mouse germ cells.

Primordial germ cells in the mouse embryo migrate from their site of origin to the gonad where they differentiate, giving rise eventually to the gametes of the mature adult animal. The migratory phase is transient and therefore permits analysis of factors regulating the motile activity of cells in tissues. Germ cells can be isolated during migration and cultured on feeder cells of an established cell line (STO). In this system the molecular interactions mediating germ cell adhesion can be probed using antibodies to both germ cell surface determinants and to extracellular matrix components.

Animals↗
PubMed1986

Migratory and postmigratory mouse primordial germ cells behave differently in culture.

In all vertebrate groups, the progenitors of the germ line, the primordial germ cells (PGCs) arise extragonadally and move to the developing gonad early in embryonic development. We have examined the behavior of isolated pregonadal and gonadal PGCs in vitro on feeder layers of an embryo-derived cell line. Histochemically and serologically identified pregonadal germ cells are found to be actively motile in vitro and, furthermore, show behavior characteristic of invasive cells. PGCs isolated from the developing gonad, however, show little locomotory activity and are not invasive on the same cellular substrate. These observations suggest that PGCs undergo a major change in phenotype at the time of their entry into the gonad anlagen.

Alkaline Phosphatase↗
PubMed1986

Invasive behaviour of mouse primordial germ cells in vitro.

We have isolated migrating primordial germ cells (PGCs) from 10.5-day mouse embryos and studied their behaviour when cultured on a mouse embryo fibroblast (STO) cell line. Living and fixed PGCs were identified by fluorescent labelling with a monoclonal antibody specific for PGCs in the culture system used. The behaviour of the cells was studied using interference reflexion microscopy (IRM) and time-lapse video cinematography. The IRM pattern displayed by PGCs is typical of highly motile cell types, the cells lack focal contacts and possess large areas of close contacts indicative of weak membrane to substrate interaction. The PGCs exhibit relatively high rates of translocation and lack contact inhibition. They were observed to underlap STO cells in subconfluent monolayers and to penetrate between the cells of confluent monolayers, becoming located between the monolayer and its substrate. These observations support the hypothesis that migrating mouse PGCs are inherently motile and are able transiently to disrupt the adhesion of surrounding cells. These results suggest that PGCs actively migrate to the developing gonad in vivo.

Animals↗
PubMed1984

Detection and identification of serum monoclonal immunoglobulin by immunoisoelectric focusing. Limits of sensitivity and use during relapse of multiple myeloma.

The limits of detection of four classes of monoclonal immunoglobulin and free light chain in serum by isoelectric focusing and immunoisoelectric focusing have been determined and the sensitivity of these techniques compared with that obtained using immunoelectrophoresis and zonal electrophoresis with immunofixation. Immunoisoelectric focusing was 10-40 times more sensitive than immunoelectrophoresis and could be used to detect concentrations of monoclonal immunoglobulin that were undetectable by zonal electrophoresis with immunofixation. The relevance of this work in monitoring multiple myeloma during treatment and relapse is discussed.

Antibodies, Monoclonal↗
PubMed1974

IgM-producing tumors in the BALB-c mouse: a model for B-cell maturation.

Five adjuvant induced BALB/c tumors producing IgM-McPc 1748, W 3469, TEPC 183, McPc 774, and Y 5781-were characterized morphologically by electron microscopy, analysis of the distribution of surface-bound and intracytoplasmic IgM using immunofluorescence, and by biochemical study of IgM synthesis, turnover, and secretion. The cells of different tumors appear to represent different stages in B-cell maturation when compared to normal, lipopolysaccharide-stimulated B cells. Thus, McPc 1748 tumor cells resemble 10-25-h stimulated normal B cells, 3469 cells resemble 20-35-h stimulated B cells, TEPC 183 cells resemble 45-65-h stimulated B cells, Y 5781 cells resemble 80-110-h stimulated B cells, and McPc 774 cells resemble 100-130-h stimulated B cells.

Animals↗