PubMed Health⌕ Search

Biomedical subjects

D S Latchman

Publications and source records attributed to D S Latchman.

At least 199 records · Page 11Linked to original sources
PubMed1995

Regulation of neurite outgrowth and SNAP-25 gene expression by the Brn-3a transcription factor.

SNAP-25 is a presynaptic nerve terminal protein which is also essential for the process of neurite outgrowth in vivo and in vitro. However the processes regulating its expression have not been characterized previously. We show that the gene encoding this protein, SNAP, is strongly activated by the Brn-3a POU (Pit-Oct-Unc) family transcription factor. Expression of both Brn-3a and SNAP-25 increases when ND7 neuronal cells are induced to extend neurite processes by serum removal. Inhibition of Brn-3a expression in these cells inhibits SNAP-25 expression and abolishes the neurite outgrowth that normally occurs in response to serum removal. These results identify Brn-3a as the first transcription factor having a role in process outgrowth in neuronal cells acting, at least in part, via the activation of SNAP-25 gene expression.

Base Sequence↗
PubMed1995

The neuronal nicotinic acetylcholine receptor alpha 2 subunit gene promoter is activated by the Brn-3b POU family transcription factor and not by Brn-3a or Brn-3c.

The regulatory region of the neuronal nicotinic acetylcholine receptor alpha 2 subunit gene, which contains six copies of the octamer-related sequence CCCCATGCAAT, is activated by the Brn-3b POU family transcription factor but not by the closely related factors Brn-3a and Brn-3c. This effect is in contrast to the previously documented inhibitory effect of Brn-3b on octamer-containing promoters that are activated by Brn-3a and Brn-3c. Activation of the alpha 2 gene by Brn-3b requires that both the POU domain and other N-terminal sequences are derived from Brn-3b and is dependent on the intactness of the alpha 2 gene regulatory region, being lost in truncated derivatives containing one, two, or four copies of the octamer-related sequence. Surprisingly, however, these truncated derivatives are activated by Brn-3c. These effects are discussed in terms of both the influence of the target sequence and its context in the promoter on activation by the various forms of Brn-3 as well as of the processes that restrict expression of the alpha 2 subunit gene to a few cells in the nervous system.

Animals↗
PubMed1995

Altered dinucleotide content within the latently transcribed regions of the DNA of alpha herpes viruses--implications for latent RNA expression and DNA structure.

The alphaherpesviruses establish latent infections in sensory neuronal cells, during which a number of latency-associated RNA transcripts (LATs) are produced. The reasons for the production of the LATs, however, and the mechanism by which the LAT region of the genome remains active during latency are unknown. Here we have analysed alphaherpesvirus sequences in an attempt to assess any differences between latently active regions of the genomes and the genomes as a whole which might be necessary for this differential expression pattern or for the LAT function, whatever that might be. We show that the LAT regions of all of the four alphaerpesviruses examined exhibit a previously unidentified and marked increase in CC + GG dinucleotides within the region and that three of the four viruses examined also show a marked decrease in CG + GC dinucleotides. The CC + GG increase was shown to occur throughout the LAT regions of these viruses, but the GC + CG decrease was shown to be confined only to LATs which are thought to be expressed as introns, and also to the intron sequences of the IE1 genes (at least for herpes simplex viruses 1 and 2), which overlap the LAT region. Bovine herpesvirus, the virus which did not show the GC + CG reduction in its LAT region but did show the GG + CC increase, is thought not to express LATs by a mechanism involving splicing, again indicating that the CG + GC reduction may be an intron-related phenomenon and also further suggesting that the two effects of CG + GC decrease and GG + CC increase may be independent of one another. Possible reasons for these unusual dinucleotide frequencies within the latently active regions of the alphaherpesviruses relating to DNA and RNA structure and to LAT function are discussed.

Base Sequence↗
PubMed1995

Activation of the herpes simplex virus immediate-early gene promoters by neuronally expressed POU family transcription factors.

Herpes simplex virus immediate-early (IE) promoters contain the TAATGARAT motif which acts as a target site for the cellular POU family transcription factors Oct-1 and Oct-2. Here we show that other members of the POU family that are expressed in sensory neurons can also affect IE promoter activity. In particular, two members of the Brn-3 family of POU proteins Brn-3a and Brn-3c can activate the IE-1 and IE-3 promoters when co-transfected into fibroblasts and neuronal cells whereas a third member Brn-3b represses IE promoter activity. In contrast, Brn-3 proteins cannot overcome the inhibitory effect of neuronal Oct-2 on IE promoter activity in co-transfections nor do they prevent transactivation of the IE promoters by the Oct-1-Vmw65 complex. The potential regulation of the IE promoters by several different neuronally expressed POU proteins during the initiation, maintenance and re-activation of latent infection in sensory neurons is discussed.

Animals↗
PubMed1995

Activation of the alpha-internexin promoter by the Brn-3a transcription factor is dependent on the N-terminal region of the protein.

The Brn-3a, Brn-3b, and Brn-3c proteins are closely related POU (Pit-Oct-Unc) family transcription factors which are expressed predominantly in neuronal cells. We have identified the alpha-internexin gene as the first reported, neuronally expressed, target gene whose promoter activity is modulated by these factors. Both the Brn-3a and Brn-3c factors can activate the alpha-internexin promoter while Brn-3b represses it and can prevent activation by Brn-3a. Using chimeric constructs containing different regions of Brn-3a or Brn-3b, we show that activation of the alpha-internexin promoter requires the N-terminal region of Brn-3a. In contrast the activation by Brn-3a but not Brn-3b of an artificial promoter containing a synthetic Brn-3 binding site can be shown using the same constructs to be dependent on the POU domain of Brn-3a. Moreover, the isolated POU domain of Brn-3a can activate this artificial promoter but not the alpha-internexin promoter. Hence Brn-3a contains two distinct transactivation domains, at the N terminus and within the POU domain, whose effect is dependent upon the target promoter. The relationship of gene transactivation by Brn-3a to its ability to transform primary cells which is also dependent on the N-terminal region of the protein is discussed.

Base Sequence↗
PubMed1995

The levels of the antagonistic POU family transcription factors Brn-3a and Brn-3b in neuronal cells are regulated in opposite directions by serum growth factors.

The Brn-3a and Brn-3b proteins are closely related POU family transcription factors with generally antagonistic effects on gene expression. We show that transfer of ND7 neuronal cells to medium containing either no foetal calf serum or low concentrations of serum results in a rise in Brn-3a mRNA levels and a fall in Brn-3b mRNA levels, although the precise serum dependence of these two effects differ. These effects can be reversed by addition of specific growth factors to the medium lacking serum, although not all growth factor treatments which suppress the rise in Brn-3a can reverse the fall in Brn-3b levels. These effects do not correlate with the effects of each treatment on cellular proliferation indicating that they are not simply a consequence of changes in proliferation. Interestingly however, treatments which produce a rise in Brn-3a levels also induce the outgrowth of neuritic processes. Hence the expression of a functionally antagonistic pair of POU factors is regulated in opposite directions by treatments with serum growth factors and this is likely to represent one means by which such growth factors modulate the gene expression patterns and ultimately the behaviour of neuronal cells.

Animals↗
PubMed1995

Nerve growth factor induces the Oct-2 transcription factor in sensory neurons with the kinetics of an immediate-early gene.

The Oct-2 transcription factor has a predominantly inhibitory effect on gene expression in neuronal cell lines. This factor and its corresponding mRNA have previously been shown to be elevated in adult rat dorsal root ganglion (DRG) neurons chronically exposed to nerve growth factor (NGF). Here we show that the Oct-2 mRNA is rapidly induced in DRG cells exposed to NGF and that such induction still occurs to a lesser extent in the presence of the protein synthesis inhibitor cycloheximide. These findings characterize Oct-2 as a novel member of the immediate-early class of NGF-induced transcription factors whose previously defined members have a predominantly stimulatory effect on the expression of other genes. Induction of the Oct-2 mRNA was also observed in DRG neurons treated with acidic fibroblast growth factor or epidermal growth factor but not with a range of other growth factors and neurotrophins. The role of the Oct-2 transcription factor in mediating the response of DRG neurons to NGF is discussed.

Animals↗
PubMed1995

Regulated splicing of the Oct-2 transcription factor RNA in neuronal cells.

The primary RNA transcript derived from the gene encoding the Oct-2 transcription factor is alternatively spliced to yield a number of different mRNAs which encode different isoforms of this protein. The mRNAs encoding two such isoforms Oct-2c and mini Oct-2 were originally detected in neuronal cells. We show here that the mRNAs encoding these forms also occur in other tissues with the proportion of the mini Oct-2 mRNA being much higher in the spleen than in the brain. However, the levels of the mini Oct-2 mRNA increase in neuronal cell lines in response to differentiation-inducing stimuli and decrease upon exposure to growth factors. Hence the splicing of the Oct-2 transcript can be regulated in both a tissue specific manner and in neuronal cells in response to specific stimuli. The significance of this effect is discussed in terms of the differing ability of different forms of Oct-2 to activate or inhibit gene expression.

Animals↗
PubMed1995

The inhibitory domain in the Oct-2 transcription factor represses gene activity in a cell type-specific and promoter-independent manner.

The Oct-2 transcription factor contains an N-terminal inhibitory domain which can act to inhibit promoter activity when linked to either its corresponding DNA-binding POU domain or the heterologous DNA binding domain of the yeast transcription factor GAL4. This inhibitory effect is independent of the number of DNA binding sites or their context in the target promoter. In contrast the effect is cell type-specific and can be relieved by over-expression of the isolated inhibitory domain in the absence of a DNA binding domain. These results suggest that the inhibitory domain acts by decreasing the activity of the basal transcriptional complex but that it operates indirectly by recruiting a second cell type-specific factor to the promoter which then interacts with the basal complex decreasing its activity.

Animals↗
PubMed1995

Direct evidence that the POU family transcription factor Oct-2 represses the cellular tyrosine hydroxylase gene in neuronal cells.

The POU family transcription factor Oct-2 was originally identified in B lymphocytes but has been shown to be expressed in neuronal cells, although it is absent in most other cell types. Cotransfection of Oct-2 expression vectors into nonneuronal cells with a tyrosine hydroxylase promoter/reporter plasmid suggests that Oct-2 can repress this promoter in this artificial situation. Here we report that reduction of endogenous Oct-2 levels in a neuronal cell line by an antisense approach results in an increase in endogenous tyrosine hydroxylase levels. In contrast, the level of the neuronal marker protein PGP9.5 remains unchanged in the antisense lines whereas that of the neuronal nitric oxide synthase decreases. Hence, the tyrosine hydroxylase gene is a natural target for repression by Oct-2 in neuronal cells. The significance of this effect is discussed in terms of the processes that regulate tyrosine hydroxylase gene expression and the role of Oct-2 in neuronal cells.

Animals↗
PubMed1995

Immediate upstream promoter regions required for neurospecific expression of SNAP-25.

The promoter structure and regulation of Snap, a gene encoding the presynaptic t-SNARE SNAP-25 implicated in synaptic vesicle docking and fusion, was studied. Transcription start-site analysis revealed two major start sites located 42 nucleotides apart. Nucleotide sequence of a promoter region 2073 nucleotides upstream of the first transcription site contains three AP-1, one CRE sequence, and three Sp1-like sites close to the TATA box. Further upstream of these sites two TG repeats were found. The ability of regions within the 5' upstream sequence to promote basal neural-specific expression in tissue culture cells was evaluated using a series of constructs containing both Snap gene start sites with progressively restricted 5' sequence linked to the chloramphenicol acetyl transferase (CAT) reporter gene. CAT expression was maximal in neuron-like undifferentiated ND7 and PC12 cells transfected with constructs containing Snap sequences up to 127 bp from the start site. In contrast, nonneuronal fibroblast cell lines did not express significant amounts of CAT, suggesting that this short 127-bp sequence is sufficient to drive neural specific expression of SNAP-25. Band shift analysis of oligonucleotides spanning from -127 to -41 bp of the Snap promoter revealed three distinct DNA-protein complexes generated by brain nuclear extracts and one by liver nuclear extracts, indicating that transcription factors that bind to this 86-bp sequence located just upstream of the TATA box are involved in regulation of basal neurospecific expression of this gene.

Animals↗
PubMed1995

Down regulation of the octamer binding protein Oct-1 during growth arrest and differentiation of a neuronal cell line.

The octamer binding transcription/DNA replication factor Oct-1 is present in virtually all cell types including proliferating cell lines of neuronal origin but is not detectable in mature non-dividing neurons. Cell cycle arrest in G0/G1 and morphological differentiation of a neuronal cell line is accompanied by a decline in the level of Oct-1 DNA binding, although the level of DNA binding by another octamer binding protein, Oct-2 is unaltered. This effect is paralled by a decline in the level of the Oct-1 mRNA in the non-dividing cells. The decrease in Oct-1 levels occurs only with the production of a mature, non-dividing neuronal phenotype and not when the cells are arrested in late G1 and do not undergo morphological differentiation. The potential role of Oct-1 and other octamer binding proteins in gene regulation in neuronal cells and in their differentiation is discussed.

Animals↗
PubMed1995

Differential cytoprotection against heat stress or hypoxia following expression of specific stress protein genes in myogenic cells.

Cells respond to sub-lethal hear stress by preferential synthesis and accumulation of several members of functionally and compartmentally distinct families of heat shock (or stress) proteins (such as hsp70, hsp90, hsp60 and hsp27). Some of these have been implicated in the development of thermotolerance and resistance to other environmental stresses. The aim of this investigation was to determine the ability of hsp70, hsp90 or hsp60 to individually protect embryonal rat heart-derived H9c2 myocytes against both (i) heat stress and (ii) substrate-free hypoxia. When H9c2 cells were subjected to a sub-lethal stress (43 degrees C for 30 min) they were shown to have elevated levels of hsp70, hsp90 and hsp60 which was maximal between 14-24 h and was associated with increased survival at 24 h against a subsequent lethal heat stress (47 degrees C for 2 h) (20.4 +/- 2.6% v 7.3 +/- 1.8%; P < 0.002). H9c2 myocytes were transfected with a plasmid containing human hsp70i, hsp90 beta or hsp60 expressed under the control of the constitutively active human beta-actin promoter or with control vector alone (containing no hsp gene). Stable colonies of primary transfectants selected for neomycin resistance showed different degrees of over-expression of hsp70i, hsp90 beta or hsp60 expression as determined by Western blotting using specific monoclonal antisera. Cells constitutively expressing high levels of hsp70i showed significantly higher survival against lethal heat stress compared to cells transfected with vector alone (39.2 +/- 6.5% v 4.5 +/- 1.0%; P < 0.0001) and also against 20 h of substrate-free hypoxia (86.4 +/- 3.0% v 3.5 +/- 0.7%; P < 0.0001). Cells constitutively expressing high levels of hsp90 beta also showed significantly higher survival against heat stress (21.5 +/- 3.0% v 9.8 +/- 3.7%; P < 0.05) but were not resistant to 20 hours of substrate-free hypoxia (0.5 +/- 1.0% v 2.0 +/- 1.7%). Cells transfected with the hsp60 plasmid showed no increased survival against either heat stress or substrate-free hypoxia. These results demonstrate that transfection of H9c2 cells with either hsp70i, hsp90 beta or hsp60 genes confers different patterns of protection against heat stress and substrate-free hypoxia, indicating functional differences between stress proteins in their ability to protect against divergent stresses.

Actins↗
PubMed1995

Differential stress protein mRNA expression during early ischaemic preconditioning in the rabbit heart and its relationship to adenosine receptor function.

The aim of this study was: (1) to elucidate in more detail the relationship between stress protein expression and brief periods of ischaemia and reperfusion, such as occur during early (classical) ischaemic preconditioning (PC) in the rabbit myocardium; (2) to determine whether stress protein expression is affected by adenosine receptor modulation, since adenosine is a mediator of the preconditioning response. We have studied the expression of the 60 kDa (hsp60); 70 kDa (hsp70-inducible and constitutive isoforms) and 27 kDa (hsp27) stress proteins and the mitochondrial ATP-synthase beta-subunit using Northern blotting. Hsp60, hsp70 and hsp27 expression were also determined at the protein level by Western blotting. Total RNA and proteins were prepared from frozen samples of ischaemic left ventricle and non-ischaemic right ventricle rabbit myocardium after the following treatments (1) sham-operated; (2) 15 min stabilization + 5 min coronary occlusion + 10 min reperfusion (PC); (3) PC + 30 min coronary occlusion (I); (4) PC + 30 min coronary occlusion + 2 h reperfusion (I/R) (5) the adenosine receptor antagonist 8-(p-sulpho-phenyl) theophyline (SPT) given 5 min prior to PC; (6) the adenosine receptor agonist 2-chlorocyclopentyl-N6-adenosine (CCPA) given in place of PC. A transient, approximately two-fold elevation in hsp60 mRNA occurred following 5 min coronary occlusion + 10 min reperfusion (PC) which was stable during a subsequent 30 min ischaemia (I), but returned to baseline during the second (2 h) reperfusion (I/R). An inducible hsp70 mRNA species appeared within 10 min of the second (30 min) coronary occlusion (I) which continued to increase to high levels during the second (2 h) reperfusion (I/R). Hsp27 mRNA expression was not altered following PC or subsequent ischaemia and reperfusion (I/R). ATP synthase beta-subunit mRNA did not change during PC or I but decreased during the subsequent 2 h reperfusion (I/R). Western blot analysis showed no change in left ventricle ischaemic zone hsp60, hsp70i/hsc70 or hsp27 protein during PC compared to an approximately two-fold elevation of hsp70i 24 h following whole body heat stress or 24 h following 4 x 5 min coronary occlusion (as reported by Marber et al., 1993). However, hsp70i, hsp60 and hsp27 showed significant decreases in immunodetectable protein following subsequent ischaemia and reperfusion (I/R). SPT inhibited the increase in hsp60 mRNA following PC (P < or = 0.05), but had no effect on hsp70, hsp27 or ATP-synthase mRNA levels. Therefore, differential expression of mRNAs for hsp60 and hsp70 occurred following ischaemia and reperfusion, with hsp70 mRNA expression involving a significant reperfusion-dependent component. CCPA had no effect on expression of mRNAs for hsp60, hsp70, hsp27 or ATP-synthase. We conclude that the early phase of adenosine receptor-dependent preconditioning in the rabbit heart is not mediated via stress protein expression. However, brief ischaemia and reperfusion resulted in differential changes in individual stress protein gene expression which may be due to different physiological and/or biochemical components of ischaemia and reperfusion in the heart. In addition, partial dependence of hsp60 expression on adenosine receptor modulation was observed.

Adaptation, Physiological↗
PubMed1995

The occurrence, nature and distribution of flares in a cohort of patients with systemic lupus erythematosus: a rheumatological view.

This study examines the occurrence, nature and distribution of disease flares in systemic lupus erythematosus (SLE) according to organ involvement. One-hundred-and-fourteen patients were seen in the Lupus Clinic of the Bloomsbury Rheumatology Unit over a 3-yr period. At each visit a data sheet was completed to assess their disease activity in eight separate organs/systems according to the British Isles Lupus Assessment Group (BILAG) activity index. This information was entered into a computer and a score for each organ system was obtained. The record of each individual patient was examined to identify flares in the individual organs; 458 flares occurred in 107 patients. The majority of patients (69%) experienced more than one flare during the study period. Fifty-four per cent of patients had flares in more than one system simultaneously, but the majority (70%) of flares involved only one system. The most severe organ involvement (A score) was most commonly observed in the musculoskeletal system, whereas severe renal disease occurred only three times. These results indicate that although SLE is a multisystem disease, the flares that occur tend to be confined to one system at a time. Further, these results demonstrate that in our rheumatology practice the most common 'A' flare observed was severe polyarthritis (A score in musculoskeletal).

Adult↗
PubMed1995

Bcl-2 expression is unaltered in unfractionated peripheral blood mononuclear cells in patients with systemic lupus erythematosus.

It has been postulated that the high levels of autoantibodies observed in systemic lupus erythematosus (SLE) patients could result from abnormal longevity of polyclonally activated B-cells. This increased survival could be due to dysfunction of apoptosis, the normal regulatory process governing their life span. The protein product of the bcl-2 gene can enhance lymphoid cell survival by interfering with apoptosis. Moreover, transgenic mice overexpressing bcl-2 in their B-cells developed an autoimmune syndrome resembling SLE. To determine whether overexpression of bcl-2 occurs in SLE patients, bcl-2 protein was measured in peripheral blood mononuclear cells from 73 SLE patients, 20 healthy individuals and 47 patients with other autoimmune diseases. Only three lupus patients had raised levels of bcl-2 and there were no statistically significant differences in the mean bcl-2 levels measured in SLE patients compared to controls. Bcl-2 levels did not correlate with overall disease activity in SLE patients.

Adult↗