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Showing papers by "Karolinska Institutet published in 1995"


Journal ArticleDOI
31 Aug 1995-Nature
TL;DR: Analysis of the nucleotide sequence of the open reading frame of the E5-1 gene led to the discovery of two missense substitutions at conserved amino-acid residues in affected members of pedigrees with a form of familial AD that has a later age of onset than the AD3 subtype (50–70 years versus 30–60 years for AD3).
Abstract: We report the cloning of a novel gene (E5-1) encoded on chromosome 1 which has substantial nucleotide and amino-acid sequence similarity to the S182 gene on chromosome 14q24.3. Mutations, including three new missense mutations in the S182 gene, are associated with the AD3 subtype of early-onset familial Alzheimer's disease (AD). Both the E5-1 and the S182 proteins are predicted to be integral membrane proteins with seven membrane-spanning domains, and a large exposed loop between the sixth and seventh transmembrane domains. Analysis of the nucleotide sequence of the open reading frame (ORF) of the E5-1 gene led to the discovery of two missense substitutions at conserved amino-acid residues in affected members of pedigrees with a form of familial AD that has a later age of onset than the AD3 subtype (50-70 years versus 30-60 years for AD3). These observations imply that the E5-1 gene on chromosome 1 and the S182 gene on chromosome 14q24.3 are members of a family of genes (presenilins) with related functions, and indicates that mutations in conserved residues of E5-1 could also play a role in the genesis of AD. Our results also indicate that still other AD susceptibility genes exist.

2,067 citations


Journal ArticleDOI
TL;DR: It is reported that exposure of cortical neurons to relatively short durations or low concentrations of NMDA, S-nitrosocysteine, or 3-morpholinosydnonimine, which generate low levels of peroxynitrite, induces a delayed form of neurotoxicity predominated by apoptotic features.
Abstract: N-Methyl-D-aspartate (NMDA) receptor-mediated neurotoxicity may depend, in part, on the generation of nitric oxide (NO.) and superoxide anion (O2.-), which react to form peroxynitrite (OONO-). This form of neurotoxicity is thought to contribute to a final common pathway of injury in a wide variety of acute and chronic neurologic disorders, including focal ischemia, trauma, epilepsy, Huntington disease, Alzheimer disease, amyotrophic lateral scelerosis, AIDS dementia, and other neurodegenerative diseases. Here, we report that exposure of cortical neurons to relatively short durations or low concentrations of NMDA, S-nitrosocysteine, or 3-morpholinosydnonimine, which generate low levels of peroxynitrite, induces a delayed form of neurotoxicity predominated by apoptotic features. Pretreatment with superoxide dismutase and catalase to scavenge O2.- partially prevents the apoptotic process triggered by S-nitrosocysteine or 3-morpholinosydnonimine. In contrast, intense exposure to high concentrations of NMDA or peroxynitrite induces necrotic cell damage characterized by acute swelling and lysis, which cannot be ameliorated by superoxide dismutase and catalase. Thus, depending on the intensity of the initial insult, NMDA or nitric oxide/superoxide can result in either apoptotic or necrotic neuronal cell damage.

2,016 citations


Journal ArticleDOI
01 Oct 1995-Neuron
TL;DR: It is shown that glutamate can induce either early necrosis or delayed apoptosis in cultures of cerebellar granule cells, suggesting that mitochondrial function is a critical factor that determines the mode of neuronal death in excitotoxicity.

1,893 citations


Journal ArticleDOI
TL;DR: A brief review of current knowledge concerning some biochemical, physiological and medical aspects of the function of ubiquinone (coenzyme Q) in mammalian organisms can be found in this article.

1,197 citations


Journal ArticleDOI
TL;DR: In the combined SDR superfamily, only one residue is strictly conserved and ascribed a crucial enzymatic function (Tyr 151 in the numbering system of human NAD(+)-linked prostaglandin dehydrogenase), and such a function is supported by chemical modifications, site-directed mutagenesis, and an active site position in those tertiary structures that have been characterized.
Abstract: Short-chain dehydrogenases/reductases (SDR) constitute a large protein family. Presently, at least 57 characterized, highly different enzymes belong to this family and typically exhibit residue identities only at the 15-30% level, indicating early duplicatory origins and extensive divergence. In addition, another family of 22 enzymes with extended protein chains exhibits part-chain SDR relationships and represents enzymes of no less than three EC classes. Furthermore, subforms and species variants are known of both families. In the combined SDR superfamily, only one residue is strictly conserved and ascribed a crucial enzymatic function (Tyr 151 in the numbering system of human NAD(+)-linked prostaglandin dehydrogenase). Such a function for this Tyr residue in SDR enzymes in general is supported also by chemical modifications, site-directed mutagenesis, and an active site position in those tertiary structures that have been characterized. A lysine residue four residues downstream is also largely conserved. A model for catalysis is available on the basis of these two residues. Binding of the coenzyme, NAD(H) or NADP(H), is in the N-terminal part of the molecules, where a common GlyXXXGlyXGly pattern occurs. Two SDR enzymes established by X-ray crystallography show a one-domain subunit with seven to eight beta-strands. Conformational patterns are highly similar, except for variations in the C-terminal parts. Additional structures occur in the family with extended chains. Some of the SDR molecules are known under more than one name, and one of the enzymes has been shown to be susceptible to native, chemical modification, producing reduced Schiff base adducts with pyruvate and other metabolic keto derivatives. Most SDR enzymes are dimers and tetramers. In those analyzed, the area of major subunit contacts involves two long alpha-helices (alpha E, alpha F) in similar and apparently strong subunit interactions. Future possibilities include verification of the proposed reaction mechanism and tracing of additional relationships, perhaps also with other protein families. Short-chain dehydrogenases illustrate the value of comparisons and diversified research in generating unexpected discoveries.

1,187 citations


Journal ArticleDOI
26 Jan 1995-Nature
TL;DR: It is concluded that intracerebral GDNF administration exerts both protective and reparative effects on the nigrostriatal dopamine system, which may have implications for the development of new treatment strategies for Parkinson's disease.
Abstract: GLIAL-CELL-LINE-DERIVED neurotrophic factor (GDNF), a recently cloned new member of the transforming growth factor-β superfamily, promotes survival of cultured fetal mesencephalic dopamine neurons1 and is expressed in the developing striatum2,3. There have, however, been no reports about effects of GDNF in situ. We have used the dopaminergic neurotoxin l–methyl–4–phenyl–1,2,3,6–tetrahydropyridine (MPTP), which produces parkinsonian symptoms in man, to determine whether GDNF might exert protective or regenerative effects in vivo in the adult nigrostriatal dopamine system in C57/B1 mice. GDNF injected over the substan-tia nigra or in striatum before MPTP potently protects the dopamine system, as shown by numbers of mesencephalic dopamine nerve cell bodies, dopamine nerve terminal densities and dopamine levels. When GDNF is given after MPTP, dopamine levels and fibre densities are significantly restored. In both cases, motor behaviour is increased above normal levels. We conclude that intracerebral GDNF administration exerts both protective and reparative effects on the nigrostriatal dopamine system, which may have implications for the development of new treatment strategies for Parkinson's disease.

1,117 citations


Book ChapterDOI
TL;DR: This chapter summarizes current methods to determine Trx and thioredoxin reductase (TR), which are likely to be involved in multiple signaling systems for redox control of cellular processes.
Abstract: Publisher Summary This chapter summarizes current methods to determine Trx and thioredoxin reductase (TR). Thioredoxin (Trx) is a small (Mr 12,000) multifunctional and ubiquitous protein characterized by having a redox-active disulfide/dithiol within the conserved active site sequence: -Trp-Cys-Gly-Pro-Cys-. Oxidized thioredoxin (Trx-S 2 ) has a disulfide, and reduced thioredoxin [Trx-(SH) 2 ] has a dithiol. Thioredoxin reductase specifically reduces Trx-S 2 to Trx-(SH) 2 using NADPH. The Trx-(SH) 2 form is a powerful protein disulfide reductase. Thus, Trx, TR, and NADPH, collectively called the thioredoxin system, operate as a powerful NADPH-dependent protein disulfide reductase system. Thioredoxin has been isolated and characterized from a wide variety of prokaryotic and eukaryotic species. Mammalian thioredoxins show about 25% sequence identity to the well-characterized E. coli protein with 108 residues. One classic function of the thioredoxin system is to act as a hydrogen donor for ribonucleotide reductase, which is essential for DNA synthesis. Redox control processes involve changes in the activity of an enzyme, a receptor, or a transcription factor via dithiol/disulfide interchange reactions. Mammalian thioredoxin reductase, with its broader substrate specificity, is likely to be involved in multiple signaling systems for redox control of cellular processes.

878 citations


Journal ArticleDOI
22 Jun 1995-Nature
TL;DR: The results highlight a previously unknown mechanism of viral escape from CTL surveillance, and support the view that the resistance of cells expressing EBNA1 to rejection mediated by CTL is a critical requirement for EBV persistence and pathogenesis.
Abstract: The Epstein-Barr virus (EBV)-encoded nuclear antigen (EBNA1) is expressed in latently EBV-infected B lymphocytes that persist for life in healthy virus carriers, and is the only viral protein regularly detected in all malignancies associated with EBV. Major histocompatibility complex (MHC) class I-restricted, EBNA1-specific cytotoxic T lymphocyte (CTL) responses have not been demonstrated. Using recombinant vaccinia viruses encoding chimaeric proteins containing an immunodominant human leukocyte antigen A11-restricted CTL epitope, amino acids 416-424 of the EBNA4 protein, inserted within the intact EBNA1, or within an EBNA1 deletion mutant devoid of the internal Gly-Ala repetitive sequence, we demonstrate that the Gly-Ala repeats generate a cis-acting inhibitory signal that interferes with antigen processing and MHC class I-restricted presentation. Insertion of the Gly-Ala repeats downstream of the 416-424 epitope inhibited CTL recognition of a chimaeric EBNA4 protein. The results highlight a previously unknown mechanism of viral escape from CTL surveillance, and support the view that the resistance of cells expressing EBNA1 to rejection mediated by CTL is a critical requirement for EBV persistence and pathogenesis.

851 citations


Journal ArticleDOI
TL;DR: Some of the patients received stereotactic radiation therapy concomitantly to more than one target, in others new metastases were also treated which appeared during the follow-up period, and a local rate of no progressive disease of 80% was observed.
Abstract: A stereotactic body frame with a fixation device has been developed for stereotactic radiation therapy of extracranial targets, a precision localization and positioning system in analogy with the stereotactic head frames used for intracranial targets. Results of the first 42 treated tumors in 31 patients are presented. Most of the patients had solitary tumors in liver, lung or retroperitoneal space. Clinical target volumes ranged from 2 to 622 cm3 (mean 78 cm3) and minimum doses to the planning target volumes (PTV) of 7.7-30 Gy/fraction (mean 14.2 Gy) were given on 1-4 occasions to a total minimum dose to the PTVs of 7.7-45 Gy (mean 30.2 Gy) to the periphery of the PTV and total mean doses to the PTVs of 8-66 Gy (mean 41 Gy). The central part of the tumor was usually given about 50% higher dose compared to that of the periphery of the PTV by a planned inhomogeneous dose distribution. Some of the patients received stereotactic radiation therapy concomitantly to more than one target, in others new metastases were also treated which appeared during the follow-up period. We observed a local rate of no progressive disease of 80% during a follow-up period of 1.5-38 months. Fifty percent of the tumors decreased in size or disappeared.

814 citations


Journal ArticleDOI
TL;DR: Investigation of human ob expression in subcutaneous and omental adipose tissue obtained from non–obese and massively obese subjects using in situ hybridization histochemistry and report on overexpression in obese people.
Abstract: Obesity is accompanied by complications such as hypertension, non–insulin–dependent diabetes mellitus and atherosclerosis, which in turn cause ischaemic heart disease, stroke and premature death1–3. The underlying mechanisms behind imbalance in energy intake and energy expenditure that lead to obesity are still controversial. In most populations, obesity is more common among women than men and is a multifactorial phenotype, which may result from a complex network of genetic and nongenetic factors. The relative importance of genetic factors for obesity is under debate4–7. Genome searches using polymorphic markers in inbred mice with phenotypes that result in extreme obesity8,9 and studies of human candidate genes are being performed in an attempt to identify genes that contribute to obesity. There is evidence that body weight is physiologically regulated4,10,11 and it has been postulated that the storage of fat may provide signals to the brain that the body is obese, which in turn may make the subject eat less and burn more fuel12,13. One of the molecules that may be involved in such signalling is the obese (ob) gene product. Mutations in ob result in profound obesity and type II diabetes in mice14,15. The mouse ob gene and its human homologue have been cloned and sequenced8. The gene is expressed in adipose tissue and the product has features of a secreted protein. We have investigated human ob expression in subcutaneous and omental adipose tissue obtained from non–obese and massively obese subjects using in situ hybridization histochemistry and report on overexpression in obese people.

756 citations


Journal ArticleDOI
TL;DR: Evidence for an independent, etiological role of PAI-1 in myocardial infarction is provided and the 4G allele of a recently described common 4/5-guanine-tract (4G/5G) polymorphism in the PAi-1 promoter is associated with higher plasma PAI -1 activity.
Abstract: Increased plasminogen-activator inhibitor 1 (PAI-1) activity is a common finding in patients with coronary heart disease. Here we provide evidence for an independent, etiological role of PAI-1 in myocardial infarction. The 4G allele of a recently described common 4/5-guanine-tract (4G/5G) polymorphism in the PAI-1 promoter is associated with higher plasma PAI-1 activity. The prevalence of the 4G allele is significantly higher in patients with myocardial infarction before the age of 45 than in population-based controls (allele frequencies of 0.63 vs. 0.53). Both alleles bind a transcriptional activator, whereas the 5G allele also binds a repressor protein to an overlapping binding site. In the absence of bound repressor, the basal level of PAI-1 transcription is increased.

Journal ArticleDOI
TL;DR: The findings indicate GDNF is a new neurotrophic factor for developing peripheral neurons and suggest possible non-neuronal roles for GDNF in the developing reproductive system.
Abstract: Glial cell line-derived neurotrophic factor (GDNF) is a neurotrophic polypeptide, distantly related to transforming growth factor-beta (TGF-beta), originally isolated by virtue of its ability to induce dopamine uptake and cell survival in cultures of embryonic ventral midbrain dopaminergic neurons, and more recently shown to be a potent neurotrophic factor for motorneurons. The biological activities and distribution of this molecule outside the central nervous system are presently unknown. We report here on the mRNA expression, biological activities and initial receptor binding characterization of GDNF and a shorter spliced variant termed GDNF beta in different organs and peripheral neurons of the developing rat. Both GDNF mRNA forms were found to be most highly expressed in developing skin, whisker pad, kidney, stomach and testis. Lower expression was also detected in developing skeletal muscle, ovary, lung, and adrenal gland. Developing spinal cord, superior cervical ganglion (SCG) and dorsal root ganglion (DRG) also expressed low levels of GDNF mRNA. Two days after nerve transection, GDNF mRNA levels increased dramatically in the sciatic nerve. Overall, GDNF mRNA expression was significantly higher in peripheral organs than in neuronal tissues. Expression of either GDNF mRNA isoform in insect cells resulted in the production of indistinguishable mature GDNF polypeptides. Purified recombinant GDNF promoted neurite outgrowth and survival of embryonic chick sympathetic neurons. GDNF produced robust bundle-like, fasciculated outgrowth from chick sympathetic ganglion explants. Although GDNF displayed only low activity on survival of newborn rat SCG neurons, this protein was found to increase the expression of vasoactive intestinal peptide and preprotachykinin-A mRNAs in cultured SCG neurons. GDNF also promoted survival of about half of the neurons in embryonic chick nodose ganglion and a small subpopulation of embryonic sensory neurons in chick dorsal root and rat trigeminal ganglia. Embryonic chick sympathetic neurons expressed receptors for GDNF with Kd 1-5 x 10(-9) M, as measured by saturation and displacement binding assays. Our findings indicate GDNF is a new neurotrophic factor for developing peripheral neurons and suggest possible non-neuronal roles for GDNF in the developing reproductive system.

Journal ArticleDOI
TL;DR: It is shown that the increased production of Aβ results from a cellular mechanism, which differs substantially from that responsible for the production of aβ from wild-type βAPP, and is explained by a competition between α and β-secretase.
Abstract: Several missense mutations causing early-onset Alzheimer's disease (AD) have been described in the gene coding for the beta-amyloid precursor protein (beta APP). A double mutation found in a Swedish family is located before the amyloid beta-peptide (A beta) region of beta APP and results in the increased production and secretion of A beta. Here we show that the increased production of A beta results from a cellular mechanism, which differs substantially from that responsible for the production of A beta from wild-type beta APP. In the latter case, A beta generation requires reinternalization and recycling of beta APP. In the case of the Swedish mutation the N-terminal beta-secretase cleavage of A beta occurs in Golgi-derived vesicles, most likely within secretory vesicles. Therefore, this cleavage occurs in the same compartment as the alpha-secretase cleavage, which normally prevents A beta production, explaining the increased A beta generation by a competition between alpha- and beta-secretase.

Journal ArticleDOI
01 Nov 1995-Pain
TL;DR: The data suggests that the brain employs different central mechanisms for chronic neuropathic pain and experimentally induced acute pain, respectively.
Abstract: This study was undertaken to explore whether the neural substrates demonstrated in brain imaging studies on experimentally induced pain are involved in the perception of chronic neuropathic pain. We investigated the cerebral representation of chronic lateralised ongoing pain in patients with painful mononeuropathy (PMN, i.e., pain in the distribution of a nerve, neuralgia) with positron emission tomography (PET), using regional cerebral blood flow (rCBF) as an index for neuronal activity. Eight patients (29-53 years) with PMN in the lower extremity (4 in the right, 4 in the left) were recruited. Paired comparisons of rCBF were made between the patient's habitual pain (HP) state and the pain alleviated (PA) state following a successful regional nerve block (RNB) with lidocaine. The ongoing neuropathic pain resulted in activation of bilateral anterior insula, posterior parietal, lateral inferior prefrontal, and posterior cingulate cortices as well as the posterior sector of the right anterior cingulate cortex (ACC), Brodmann area (BA) 24, regardless of the side of PMN. In addition, a reduction in rCBF was noted in the contralateral posterior thalamus. No significant change of rCBF was detected in the somatosensory areas, i.e., SI and SII. The cerebral activation pattern, while addressing the differences between the HP and PA states, emphasises the affective-motivational dimension in chronic ongoing neuropathic pain. The striking preferential activation of the right ACC (BA 24), regardless of the side of the PMN, not only confirms that the ACC participates in the sensorial/affectional aspect of the pain experience but also suggests a possible right hemispheric lateralisation of the ACC for affective processing in chronic ongoing neuropathic pain. Our data suggests that the brain employs different central mechanisms for chronic neuropathic pain and experimentally induced acute pain, respectively.

Journal ArticleDOI
TL;DR: It is reported that NO in humans is produced by epithelial cells in the paranasal sinuses and is present in sinus air in very high concentrations, close to the highest permissible atmospheric pollution levels.
Abstract: Nitric oxide (NO) is present in air derived from the nasal airways. However, the precise origin and physiological role of airway-derived NO are unknown. We report that NO in humans is produced by epithelial cells in the paranasal sinuses and is present in sinus air in very high concentrations, close to the highest permissible atmospheric pollution levels. In immunohistochemical and mRNA in situ hybridization studies we show that an NO synthase most closely resembling the inducible isoform is constitutively expressed apically in sinus epithelium. In contrast, only weak NO synthase activity was found in the epithelium of the nasal cavity. Our findings, together with the well-known bacteriostatic effects of NO, suggest a role for NO in the maintenance of sterility in the human paranasal sinuses.

Journal ArticleDOI
TL;DR: The physiologic metabolic role of the anabolic enzymes is discussed in relation to catabolic pathways, and alternative pathways for nucleoside analogue phosphorylation are surveyed, such as the phosphotransfer capacity of 5'-nucleotidase.

Journal ArticleDOI
18 May 1995-Nature
TL;DR: The stereochemistry suggests a mechanism by which heterodimers recognize the inter-half-site spacing between direct repeats of the consensus sequence 5′-AGGTCA-3′ separated by one to five base pairs.
Abstract: Nuclear receptor heterodimers recognize response elements composed of two direct repeats of the consensus sequence 5'-AGGTCA-3' separated by one to five base pairs The 19 A crystal structure of the complex formed by the DNA-binding domains of the 9-cis retinoic acid receptor and thyroid hormone receptor bound to a thyroid-response element shows that the subunits interact through a DNA-supported interface involving the carboxy-terminal extension of the DNA-binding domain of the thyroid hormone receptor The stereochemistry suggests a mechanism by which heterodimers recognize the inter-half-site spacing between direct repeats

Journal ArticleDOI
TL;DR: NGFI-B and NURR1 can increase the potential of RXR to modulate gene expression in a ligand-dependent manner by allowing a distinct class of direct repeats to serve as specific RXR response elements, and suggest a novel mechanism for convergence between vitamin A or retinoid and growth factor signaling pathways.
Abstract: In addition to its role as a 9-cis retinoic acid receptor, RXR has an important role in the regulation of multiple hormonal pathways through heterodimerization with nuclear receptors. Here, we show that two orphan receptors, NGFI-B and NURR1, which have been shown previously to interact with DNA as monomers, also can heterodimerize with RXR. These heterodimers bind selectively to a class of retinoic acid response elements composed of direct repeats spaced by 5 nucleotides. In this respect they are similar to heterodimers formed between RXR and the receptor for all-trans retinoic acid, RAR. However, whereas RXR is inhibited in the RXR-RAR heterodimer, NGFI-B/NURR1 promote efficient activation in response to RXR ligands and therefore shift RXR from a silent to an active heterodimerization partner. These data show that NGFI-B and NURR1 can increase the potential of RXR to modulate gene expression in a ligand-dependent manner by allowing a distinct class of direct repeats to serve as specific RXR response elements. Because expression of both NGFI-B and NURR1 is rapidly induced by various growth factors, these findings also suggest a novel mechanism for convergence between vitamin A or retinoid and growth factor signaling pathways.

Journal ArticleDOI
TL;DR: Caffeine has a number of central effects directly or indirectly related to adenosine receptors, some of which are potentially useful, and drug development based on the actions of caffeine should be interesting.
Abstract: Of the known biochemical actions of caffeine, only inhibition of adenosine receptors occurs at concentrations achieved during normal human consumption of the drug. Under normal physiological conditions, adenosine is present in sufficient concentrations to activate A1 and A2a receptors. Via actions on A, receptors, adenosine decreases neuronal firing and the release of neurotransmitters. The exact mechanisms are not known, but several possibilities are discussed. Via actions on A2a receptors, adenosine - and hence caffeine - can influence dopaminergic neurotransmission. Caffeine can induce rapid changes in gene expression and, somewhat later, marked adaptive changes. These include antiepileptic and neuroprotective changes. Thus, caffeine has a number of central effects directly or indirectly related to adenosine receptors. Some of these are potentially useful, and drug development based on the actions of caffeine should be interesting.

Journal ArticleDOI
TL;DR: The basic definitions and some distinctive characteristics of the two types of transmission are discussed and the evidence for different types of intercellular communication besides synaptic transmission in the central nervous system during phylogeny, and in vertebrates in physiological and pathological conditions is reviewed.

Journal ArticleDOI
TL;DR: In addition to its temporal and spatial regulation nestin expression appears to be regulated at the level of subcellular mRNA localization: in columnar neuroepithelial and radial glial cells nestin mRNA is predominantly localized to the pial endfeet.

Journal ArticleDOI
TL;DR: 27, 24, and 7 alpha-hydroxycholesterol were the most abundant cholesterol oxidation products in human plasma and the other oxysterols determined were present in concentrations lower than 30 ng/ml.

Book ChapterDOI
TL;DR: In the combined SDR superfamily, only one residue is strictly conserved and ascribed a crucial enzymatic function (Tyr 151 in the numbering system of human NAD(+)-linked prostaglandin dehydrogenase), and such a function is supported by chemical modifications, site-directed mutagenesis, and an active site position in those tertiary structures that have been characterized.
Abstract: Short-chain dehydrogenases/reductases (SDR) constitute a large protein family. Presently, at least 57 characterized, highly different enzymes belong to this family and typically exhibit residue identities only at the 15-30% level, indicating early duplicatory origins and extensive divergence. In addition, another family of 22 enzymes with extended protein chains exhibits part-chain SDR relationships and represents enzymes of no less than three EC classes. Furthermore, subforms and species variants are known of both families. In the combined SDR superfamily, only one residue is strictly conserved and ascribed a crucial enzymatic function (Tyr 151 in the numbering system of human NAD(+)-linked prostaglandin dehydrogenase). Such a function for this Tyr residue in SDR enzymes in general is supported also by chemical modifications, site-directed mutagenesis, and an active site position in those tertiary structures that have been characterized. A lysine residue four residues downstream is also largely conserved. A model for catalysis is available on the basis of these two residues. Binding of the coenzyme, NAD(H) or NADP(H), is in the N-terminal part of the molecules, where a common GlyXXXGlyXGly pattern occurs. Two SDR enzymes established by X-ray crystallography show a one-domain subunit with seven to eight beta-strands. Conformational patterns are highly similar, except for variations in the C-terminal parts. Additional structures occur in the family with extended chains. Some of the SDR molecules are known under more than one name, and one of the enzymes has been shown to be susceptible to native, chemical modification, producing reduced Schiff base adducts with pyruvate and other metabolic keto derivatives. Most SDR enzymes are dimers and tetramers. In those analyzed, the area of major subunit contacts involves two long alpha-helices (alpha E, alpha F) in similar and apparently strong subunit interactions. Future possibilities include verification of the proposed reaction mechanism and tracing of additional relationships, perhaps also with other protein families. Short-chain dehydrogenases illustrate the value of comparisons and diversified research in generating unexpected discoveries.

Journal ArticleDOI
TL;DR: Two divergently transcribed operons in Escherichia coli required for the expression of fibronectin‐ and Congo red‐binding curli polymers were identified and characterized by transposon mutagenesis, sequencing and transcriptional analyses, as well as for their ability to produce the curli subunit protein.
Abstract: Two divergently transcribed operons in Escherichia coli required for the expression of fibronectin- and Congo red-binding curli polymers were identified and characterized by transposon mutagenesis, sequencing and transcriptional analyses, as well as for their ability to produce the curli subunit protein. The csgBA operon encodes CsgA, the major subunit protein of the fibre, and CsgB, a protein with sequence homology to CsgA. A non-polar csgB mutant is unaffected in its production of CsgA, but the subunit protein is not assembled into insoluble fibre polymers. A third open reading frame, orfC, positioned downstream of csgA may affect some functional property of curli since an insertion in this putative gene abolishes the autoagglutinating ability typical of curliated cells without affecting the production of the fibre. The promoter for the oppositely transcribed csgDEFG operon was identified by primer extension and shown, like the csgBA promoter, to be dependent upon the alternate stationary phase-specific sigma factor sigma s in wild-type cells, but not in mutants lacking the nucleoid associated protein H-NS. Insertions in csgD abolish completely trancription from the csgBA promoter. Therefore, any regulatory effect on the csgBA promoter might be secondary to events controlling the csgDEFG promoter and/or activation of CsgD. Insertions in csgE, csgF and csgG abolish curli formation but allow CsgA expression suggesting that one or more of these gene products are involved in secretion/assembly of the CsgA subunit protein. No amino acid sequence homologies were found between the CsgE, CsgF and CsgG proteins and secretion/assembly proteins for other known bacterial fibres, suggesting that the formation of curli follows a novel pathway.

Journal ArticleDOI
08 Jun 1995-Nature
TL;DR: It is shown that clusters of vesicles at synaptic release sites are composed of two pools, a distal pool containing synapsin and a proximal pool devoid of synAPSin and located adjacent to the presynaptic membrane.
Abstract: Nerve terminals are unique among cellular secretory systems in that they can sustain vesicular release at a high rate. Although little is known about the mechanisms that account for the distinctive features of neurotransmitter release, it can be assumed that neuron-specific proteins are involved. One such protein family, the synapsins, are believed to regulate neurotransmitter release through phosphorylation-dependent interactions with synaptic vesicles and cytoskeletal elements. Here we show that clusters of vesicles at synaptic release sites are composed of two pools, a distal pool containing synapsin and a proximal pool devoid of synapsin and located adjacent to the presynaptic membrane. Presynaptic injection of synapsin antibodies resulted in the loss of the distal pool, without any apparent effect on the proximal pool. Depletion of this distal pool was associated with a marked depression of neurotransmitter release evoked by high-frequency (18-20 Hz) but not by low-frequency (0.2 Hz) stimulation. Thus the availability of the synapsin-associated pool of vesicles seems to be required to sustain release of neurotransmitter in response to high-frequency bursts of impulses.

Journal ArticleDOI
TL;DR: The composition of the glial scar after traumatic CNS injury in rat and mouse is more complex than previously assumed: expression of the intermediate filament nestin is induced in reactive astrocytes, suggesting that defined nestin regulatory regions mediate the injury response.
Abstract: Neuronal regeneration does generally not occur in the central nervous system (CNS) after injury, which has been attributed to the generation of glial scar tissue In this report we show that the composition of the glial scar after traumatic CNS injury in rat and mouse is more complex than previously assumed: expression of the intermediate filament nestin is induced in reactive astrocytes Nestin induction occurs within 48 hours in the spinal cord both at the site of lesion and in degenerating tracts and lasts for at least 13 months Nestin expression is induced with similar kinetics in the crushed optic nerve In addition to the expression in reactive astrocytes, we also observed nestin induction within 48 hours after injury in cells close to the central canal in the spinal cord, while nestin expressing cells at later timepoints were found progressively further out from the central canal This dynamic pattern of nestin induction after injury was mimicked by lacZ expressing cells in nestin promoter/lacZ transgenic mice, suggesting that defined nestin regulatory regions mediate the injury response We discuss the possibility that the spatiotemporal pattern of nestin expression reflects a population of nestin positive cells, which proliferates and migrates from a region close to the central canal to the site of lesion in response to injury

Journal ArticleDOI
TL;DR: The construction of the first complete genetic linkage map of the laboratory rat is reported, identifying 432 markers that show polymorphisms between the SHR and BN rat strains and mapped them in a single SHR × BN F2 intercross.
Abstract: We report the construction of the first complete genetic linkage map of the laboratory rat. By testing 1171 simple sequence length polymorphisms (SSLPs), we have identified 432 markers that show polymorphisms between the SHR and BN rat strains and mapped them in a single (SHR × BN) F2 intercross. The loci define 21 large linkage groups corresponding to the 21 rat chromosomes, together with a pair of nearby markers on chromosome 9 that are not linked to the rest of the map. Because 99.5% of the markers fall into one of the 21 large linkage groups, the maps appear to cover the vast majority of the rat genome. The availability of the map should facilitate whole genome scans for genes underlying qualitative and quantitative traits relevant to mammalian physiology and pathobiology.

Journal ArticleDOI
TL;DR: This work has localized the PS-1 gene to a 75 kb region and present the structure of this gene, evidence for alternative splicing and describe six novel mutations in early onset FAD pedigrees all of which alter residues conserved in the STM26 (Presenilin 2: PS-2) gene.
Abstract: Genetic linkage studies place a gene causing early onset familial Alzheimer's disease (FAD) on chromosome 14q24.3 (refs 1–4). Five mutations within the S182 (Presenilin 1: PS–1) gene, which maps to this region, have recently been reported in several early onset FAD kindreds5. We have localized the PS-1 gene to a 75 kb region and present the structure of this gene, evidence for alternative splicing and describe six novel mutations in early onset FAD pedigrees all of which alter residues conserved in the STM26 (Presenilin 2: PS-2) gene.

Journal ArticleDOI
TL;DR: Experimental and clinical evidence suggest that acupuncture may affect the sympathetic system via mechanisms at the hypothalamic and brainstem levels, and that the hypothalamus beta-endorphinergic system has inhibitory effects on the vasomotorcenter, VMC.

Journal ArticleDOI
TL;DR: It is demonstrated not only that patients with PSC and UC have a significantly higher risk of developing colorectal neoplasia compared with patients having UC only, but also that Patients with P SC and UC having coloreCTal neoperasia are more prone to develop cholangiocarcinoma.