PubMed Health⌕ Search

Biomedical subjects

D B Wilson

Publications and source records attributed to D B Wilson.

At least 19 recordsLinked to original sources
PubMed1992

Stereoselective hydrolysis catalyzed by related beta-1,4-glucanases and beta-1,4-xylanases.

Over 80 beta-1,4-glucanases and beta-1,4-xylanases can be classified into one of eight families on the basis of amino acid sequence similarities in their catalytic domains (Gilkes, N. R., Henrissat, B., Kilburn, D. G., Miller, R. C., Jr., and Warren, R. A. J. (1991) Microbiol. Rev. 55, 303-315). As a test of this classification, the stereochemical course of hydrolysis of 10 enzymes representative of five families has been determined using proton NMR. These data, together with published data for six additional enzymes, show that representatives of a given enzyme family have the same stereoselectivity: four families catalyze hydrolysis with retention of anomeric configuration, two with inversion. The results support the hypothesis that family members share a common general fold, active site topology, and catalytic mechanism.

Endo-1,4-beta Xylanases↗
PubMed1992

Human transcription factor GATA-2. Evidence for regulation of preproendothelin-1 gene expression in endothelial cells.

Previously, we showed that the promoter of the gene encoding preproendothelin-1 (PPET-1) contains a GATA motif that is essential for activity and interacts with a nuclear factor similar in size and binding specificity to the erythroid transcription factor GATA-1. To identify this endothelial GATA-binding protein, a human endothelial cell cDNA library was screened with oligonucleotide probes for a portion of the zinc finger domain of GATA-1. A 2.6-kilobase cDNA encoding a 470 amino acid protein was obtained. Sequence analysis revealed a predicted protein which is the human counterpart of a related chicken protein, designated GATA-2. Human GATA-2 is expressed by a variety of cells, including erythroid, HeLa, and endothelial cells. A complex of a GATA-containing probe and recombinant GATA-2 expressed in COS cells comigrates with that present in gel shift experiments with nuclear extract derived from endothelial cells. In addition, expressed human GATA-2 protein transactivates reporter gene constructs containing either minimal GATA promoter elements or the native PPET-1 promoter in a cotransfection assay. Retinoic acid treatment of endothelial cells results in down-regulation of GATA-2 expression as well as down-regulation of PPET-1 gene expression. Human homologs of other known GATA-binding transcription factors are either absent from endothelial cells (in the case of GATA-1) or made in small quantities and not significantly affected by retinoid acid in these cells (in the case of GATA-3), making it unlikely that they regulate the PPET-1 gene. We propose that GATA-2 is the GATA-binding protein required for PPET-1 gene expression in endothelial cells.

Amino Acid Sequence↗
PubMed1992

Aberrant convergence of the neural folds in the mouse mutant vl.

Progressive changes in the dorsolateral angles (DA) and ventral angle (VA) during elevation and convergence of the caudal neural folds were morphometrically analyzed in normal and dysraphic abnormal embryos of the mouse mutant vacuolated lens (vl), and correlations with the configuration of microfilaments in the apices of neuroepithelial cells were made by means of ultrastructural cytochemistry. In 22-28 somite stage abnormal (vl/vl) embryos, the DA and VA are larger than those in their normal counterparts at each comparable level of the caudal neural folds, suggesting that defective convergence involves both the DA and VA in this mutant. In 30-35 somite stage abnormal embryos, the VA is likewise larger than that in normal embryos in which the neural folds have converged and closed; however, the DAs are much smaller, indicating that a medial collapse of the dorsal ends of the neural folds may occur secondary to the closure failure. At the DA, the ultrastructural configuration of microfilaments is similar in abnormal and normal embryos in terms of their circumferential arrangement around the perimeters of the neuroepithelial cell apices. In abnormal embryos, however, the bundles of microfilaments are more delicate and less prominent than in normal embryos; thus it is possible that a quantitative and/or functional deficiency in these elements may be involved in the failure of the abnormal neuroepithelium to bend properly during convergence of the neural folds.

Actin Cytoskeleton↗
PubMed1992

Adrenocorticotropic cell distribution in adult and embryonic pituitaries of the little (lit) mutant mouse.

Immunofluorescence and colloidal gold immunocytochemistry were used to analyze the regional distribution of adrenocorticotropic (ACTH) cells and their ultrastructural relationship with growth hormone (GH) cells in adult and 17-day embryonic little (lit/lit) mice that exhibit GH cell defects. Adult lit/lit pituitaries lack the distinctive regionalization of ACTH cells that characterizes normal pituitaries, although typical ultrastructural relationships occur between some ACTH and GH cells. In 17-day embryos, normal and lit/lit pituitaries show similar distributions of ACTH cells. However, in lit/lit pituitaries the pars distalis cells are more loosely arranged, with poorly defined clusters and cords, than in normal glands. The results indicate that whereas the lack of ACTH cell regionalization in adult glands may be a secondary effect of the GH cell defect, the differences in overall integrity of the 17-day embryonic glands suggest the possibility of a developmental disturbance during early stages of gestation in this mutant.

Adrenocorticotropic Hormone↗
PubMed1992

Recombinant tissue-type plasminogen activator versus a novel dosing regimen of urokinase in acute pulmonary embolism: a randomized controlled multicenter trial.

Thrombolysis of acute pulmonary embolism can be accomplished more rapidly and safely with 100 mg of recombinant human tissue-type plasminogen activator (rt-PA) (Activase) than with a conventional dose of urokinase (Abbokinase) given as a 4,400-U/kg bolus dose, followed by 4,400 U/kg per h for 24 h. To determine the effects of a more concentrated urokinase dose administered over a shorter time course, this trial enrolled 90 patients with baseline perfusion lung scans and angiographically documented pulmonary embolism. They were randomized to receive either 100 mg/2 h of rt-PA or a novel dosing regimen of urokinase: 3 million U/2 h with the initial 1 million U given as a bolus injection over 10 min. Both drugs were delivered through a peripheral vein. To assess efficacy after initiation of therapy, repeat pulmonary angiograms at 2 h were performed in 87 patients and then graded in a blinded manner by a panel of six investigators. Of the 42 patients allocated to rt-PA therapy, 79% showed angiographic improvement at 2 h, compared with 67% of the 45 patients randomized to urokinase therapy (95% confidence interval for the difference in these proportions [rt-PA minus urokinase] is -6.6% to 30.4%; p = 0.11). The mean change in perfusion lung scans between baseline and 24 h was similar for both treatments. Three patients (two treated with rt-PA and one with urokinase) had an intracranial hemorrhage, which was fatal in one. The results indicate that a 2-h regimen of rt-PA and a new dosing regimen of urokinase exhibit similar efficacy and safety for treatment of acute pulmonary embolism.

Acute Disease↗
PubMed1992

Properties of a genetically reconstructed Prevotella ruminicola endoglucanase.

A pUC19-derived plasmid was constructed that coded for a hybrid cellulase with the Thermomonospora fusca E2 cellulose-binding domain at its C terminus joined to the Prevotella ruminicola 40.5-kDa carboxymethyl cellulase (CMCase). The hybrid enzyme was purified and characterized enzymatically. It bound tightly to cellulose, and its specific activities on carboxymethyl cellulose, amorphous cellulose, and ball-milled cellulose were 1.5, 10, and 8 times that of the 40.5-kDa CMCase, respectively. Furthermore, the modified enzyme gave synergism with an exocellulase in the degradation of filter paper, while the 40.5-kDa CMCase did not.

Actinomycetales↗
PubMed1992

Abnormal elevation of the neural folds in the loop-tail mutant mouse.

The processes of elevation and convergence of the spinal neural folds were analyzed in normal (+/+; Lp/+) and abnormal (Lp/Lp) embryos of the loop-tail mutant mouse in order to determine possible mechanisms underlying the dysraphic defect characterized by a failure of the neural fold to close in this mutant. The results indicate that the neural folds are already defective during very early phases of elevation, with greater distances between the apical points of the paired walls of the neural groove, larger ventral angles and higher ratios of luminal/basal linear distances occurring in the abnormal embryos relative to those in normal embryos. The cross-sectional area of the neuroepithelium is also greater in abnormals, suggesting that faulty elongation of the neuraxis may contribute to the dysraphic condition.

Animals↗
PubMed1992

Identification and subcellular localization of human rab5b, a new member of the ras-related superfamily of GTPases.

Members of the mammalian rab family of GTPases are associated with specific subcellular compartments, where these proteins are postulated to function in vesicular transport. By screening a human umbilical vein endothelial cell library with degenerate oligonucleotide probes, we have isolated a 1.6-kb cDNA clone encoding a 215-amino-acid protein belonging to the rab family of GTPases. This newly identified rab protein is 81% identical to human rab5, the canine counterpart of which has been localized to the plasma membrane and early endosomes. In light of this homology, we have named this new member of the GTPase superfamily "rab5b." Northern analysis using the rab5b cDNA as a probe revealed a 3.6-kb mRNA in a variety of cell types, including human umbilical vein endothelial cells, K562 erythroleukemia cells, U937 monoblastic cells, and HeLa cells. A fusion protein between glutathione-S-transferase (GST) and rab5b was expressed in bacteria and purified to homogeneity. The recombinant protein was shown to bind GTP and GDP. As is typical of other recombinant rab proteins, the rab5b-GST fusion protein displayed a low intrinsic rate of GTP hydrolysis (0.005/min). An antiserum to rab5b was prepared and used to determine the apparent molecular size and subcellular distribution of the protein. Western blotting with this antibody revealed a 25-kD protein in COS cells transfected with rab5b and in nontransfected HeLa cells. Indirect immunofluorescence and subcellular fractionation showed that rab5b localizes to the plasma membrane. We speculate that rab5b plays a role in vesicular trafficking at the plasma membrane in various cell types.

Amino Acid Sequence↗
PubMed1992

Biochemistry and genetics of actinomycete cellulases.

The order Actinomycetales includes a number of genera that contain species that actively degrade cellulose and these include both mesophilic and facultative thermophilic species. Cellulases produced by strains from two of the genera containing thermophilic organisms have been studied extensively: Microbispora bispora and Thermomonospora fusca. Fractionation of M. bispora cellulases has identified six different enzymes, all of which were purified to near homogeneity and partially characterized. Two of these enzymes appear to be exocellulases and gave synergism with each other and with the endocellulases. The structural genes of five M. bispora cellulases have been cloned and one was sequenced. Fractionation of T. fusca cellulases has identified five different enzymes, all of which were purified to near homogeneity and partially characterized. One of the T. fusca enzymes gives synergism in the hydrolysis of crystalline cellulose with several T. fusca endocellulases and with Trichoderma reesei CBHI but not with T. reesei CBHII. Each T. fusca cellulase contains distinct catalytic and cellulose binding domains. The structural genes of four of the T. fusca endoglucanases have been cloned and sequenced, while three cellulase genes have been cloned from "T. curvata". The T. fusca cellulase genes are expressed at a low level in Escherichia soli, but at a high level in Streptomyces lividans. Sequence comparisons have shown that there are no significant amino acid homologies between any of the catalytic domains of the four T. fusca cellulases, but each of them shows extensive homology to several other cellulases and fits in one of the five existing cellulase gene families. There have been extensive studies of the regulation of the synthesis of these cellulases and a number of regulatory mutants have been isolated. This work has shown that the different T. fusca cellulases are coordinately regulated over a 100-fold range by two independent controls; induction by cellobiose and repression by any good carbon source.

Actinomycetales↗
PubMed1992

Histopathology of the pituitary gland in neonatal little (lit) mutant mice.

The pituitary gland in the little (lit) mutant mouse was analyzed with respect to the cytoarchitecture of the pars distalis and the volumetric density of immunoreactive growth hormone (GH) cell granules in neonatal lit/lit and normal C57BL mice. At 8 days postnatally the volume of GH granules/total tissue was significantly less in the lit/lit pars distalis, and the cells were loosely arranged, as compared with the normal pars distalis. In newborn mice a statistically significant difference could not be detected between normal and lit/lit mice with respect to the volumetric density of GH granules; however, differences occurred in the cytoarchitectural organization of the pars distalis. These differences included prominent vascular channels and well-defined cords and clusters of cells in the normal newborn mice, in contrast to indistinct vascular elements and a more diffuse arrangement of cells in lit/lit.

Animals↗
PubMed1991

Analysis of T cell receptor beta chains in Lewis rats with experimental allergic encephalomyelitis: conserved complementarity determining region 3.

This study explores the usage of T cell antigen receptor (TCR) beta chain elements in Lewis rats with experimentally induced allergic encephalomyelitis (EAE). TCRs from 15 different T cell clones and hybridomas derived from animals immunized with myelin basic protein (MBP), and all having specificity for the 21-mer encephalitogenic fragment MBP 68-88, utilized V beta 8.2. In addition, there was a marked conservation of the first two amino acid residues of the junctional complementarity determining region 3 (CDR3) associated with the V beta 8.2 receptors. 12 of 15 contained an aspartic acid followed by serine regardless of the associated J beta element. At the nucleotide level, this conservation of AspSer residues was accomplished with few or no nongermline-encoded nucleotide (N) additions. A similar pattern of AspSer usage and N region nucleotide additions was observed in a number of V beta 8.2 isolates derived from MBP-immunized lymph nodes. In contrast, V beta 8.2 polymerase chain reaction amplified isolates from Lewis T cells activated with concanavalin A or from lymph nodes of complete Freund's adjuvant-immunized animals showed no AspSer utilization (0/31) in the CDR3, and four to nine N region nucleotide additions. We conclude from this finding that AspSer residues in the CDR3, limited N region nucleotide additions, along with V beta 8.2 sequences, contribute to TCR specificity for MBP 68-88. This raises the possibility that encephalitogenic, disease-causing T cells either represent a population that derives from late fetal life or alternatively, that they are rare cells with this particular TCR phenotype contributed to the T cell pool throughout adulthood and are selected by antigen. In either case, the CDR3 AspSer sequences as well as V beta 8.2 sequences are candidates for the receptor target structures recognized by regulator T cells in recovery from and resistance to active EAE. In this respect, a preliminary analysis of TCR utilization in three T cell clones specific for MBP 68-88 isolated from animals recovered from active EAE indicates that while all three use V beta 8.2, only one contains AspSer in the CDR3.

Amino Acid Sequence↗
PubMed1991

Break for SCIDs.

Explore the source record for details and available documents.

Animals↗
PubMed1991

Generation of T cells with lytic specificity for atypical antigens. I. A mitochondrial antigen in the rat.

F1 rats primed with normal parental strain lymphocyte populations and restimulated in culture with parental lymphoblasts generate potent cytotoxic T cell responses to unusual antigen systems. Here we describe in the Lewis (L)/DA anti-DA combination an antigen system most likely of mitochondrial origin with the following properties: it is transmitted maternally from DA strain females, inherited in an extra-chromosomal manner, restricted by class I RT1Aa major histocompatibility complex gene products, extinguished on target cells treated with chloramphenicol, and its pattern of expression in different rat strains correlates with restriction fragment-length polymorphisms of mitochondrial DNA. Sequence analysis of the rat ND1 gene indicates that the maternally transferred factor in the rat is not a homologue of the maternally transmitted factor responsible for the mitochondrial antigen in mice. In keeping with its inheritance from DA females, this antigen is present on target cells from (DA female x L male)F1 donors and all other F1 combinations derived from DA female parents, but absent from target cells from some F1 combinations (L/DA and Wistar-Furth [WF]/DA) derived from DA strain males. The presence of this antigen in other F1 combinations (Brown Norway [BN]/DA, August 2880 [AUG]/DA, and PVG/DA) indicates that this mitochondrial antigen system is shared by the DA, BN, and PVG strains, but not by the L and WF strains.

Amino Acid Sequence↗
PubMed1991

Generation of T cells with lytic specificity for atypical antigens. II. A novel antigen system in the rat dependent on homozygous expression of major histocompatibility complex genes of the class I-like RT1C region.

Lymphocytes from parental strain DA rats can induce potent killer cell responses to atypical antigen systems in F1 Lewis (L)/DA and DA/L recipients. Here, we describe an antigen system, H, present on homozygous parental target cells, but not on F1 cells. This antigen system is unusual in several respects: it does not involve class I RT1A gene products usually used by killer cell responses in the rat, it maps to the major histocompatibility complex (MHC) class I-like RT1C region, and it requires homozygous expression of RT1Cav1 alleles. This may be another example, this time involving the RT1C region, of an MHC gene product antigenically altered by an MHC-linked trans-activating modifier gene.

Animals↗
PubMed1991

Generation of T cells with lytic specificity for atypical antigens. III. Priming F1 animals with antigen-bearing cells also having reactivity for host alloantigens allows for potent lytic T cell responses.

Here, we explore the conditions required for generating two different highly potent F1 antiparental killer cell populations to unusual antigens in rats. The first, L/DA anti-DA, has lytic specificity for two antigen systems: MTA, a mitochondrial antigen expressed on DA and DA Lewis (L) target cells restricted by RT1A class I molecules; and H, an antigen that maps to the class I-like RT1C region and is present only on parental target cells from donors homozygous at the major histocompatibility complex. The second killer population is generated in the reciprocal DA/L anti-DA combination and has lytic specificity only for the H antigen system. We show that the killer cells are T cells, and that generation of these F1 cytotoxic T lymphocytes (CTL) requires an in vivo priming step in which it is essential that the inoculated parental cells bear the relevant target antigens and possess alloreactivity for F1 host antigens. The requirement for alloreactivity and antigen on the same priming cell population suggests that these potent lytic responses depend on a situation akin to a hapten-carrier effect that bypasses otherwise ineffective helper responses by the host to these unusual antigens. Restimulation of F1 lymphocytes in culture is also necessary, requiring the presence of antigen on irradiated lymphoblast stimulator cells, but alloreactivity to responder cell antigens is not necessary; normal, nonactivated lymph node cells are completely ineffective as stimulators. For effective lysis, the target cells need not possess the potential for alloreactivity to responder F1 CTL. We also demonstrate in a preliminary way additional antigen systems defined by killer populations raised with other F1 antiparental strain combinations.

Animals↗