A case of syncope on olfactory stimulation.
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Adult↗
Biomedical subjects
M Forte
Publications and source records attributed to M Forte.
The acquisition of anti-pneumococcal capsular polysaccharide Haemophilus influenzae type b and tetanus toxoid antibodies, with age, in the UK.
Antibody levels specific for capsular polysaccharides of Streptococcus pneumoniae and Haemophilus influenzae type b (Hib) and to tetanus toxoid (TT), were measured in serum samples of 750 age-stratified subjects from the UK. The study subjects comprised healthy adult volunteers and hospitalized children undergoing elective surgery, excluding those with a history of infection or under investigation for immunological or haematological disorders. These antibody levels were calibrated by comparison with serum pool obtained from healthy adult volunteers, who were immunized with Hib polyribose-phosphate vaccine (Merieux). The data are intended to provide reference ranges to assist in the interpretation of specific antibody measurements in the clinical setting. Maternal IgG pneumococcal capsular polysaccharide (PCP) specific antibody levels, geometric-mean titre (GMT) 1/22, were lost by 6 months of age (GMT of 1/9). They remained low until 3-5 years (GMT of 1/20), and consisted principally of IgG1. Thereafter, IgG anti-PCP antibody titres increased steadily to adult levels (GMT of 1/275), of which 80% was IgG2. Anti-PCP antibody titres of the IgM isotype rose steadily from a GMT 1/21 (0-6 months) to 1/420 (3-5 years), a level which was maintained until adulthood. Anti-Hib antibody concentrations, determined by RABA, again demonstrated the decline in maternal antibody, from 0.18 micrograms/ml in the 0-6 month age cohort, to 0.09 microgram/ml between 6 and 12 months. Geometric-mean antibody concentrations remained below 0.2 micrograms/ml until 3-5 years, then increased with age, attaining the mean adult level of 1.02 micrograms/ml. Anti-TT antibody concentrations were measured in the same sera, by ELISA. Two peaks in anti-TT antibody levels were seen in children of 0.059 IU/ml and 0.166 IU/ml corresponding to the schedule of routine childhood immunization in the first year and at 5 years of age.
Adolescent↗
Cloning and expression of a rat somatostatin receptor enriched in brain.
The tetradecapeptide somatostatin (SRIF) is a hormone release-inhibiting substance that mediates diverse effects in brain and peripheral organs via specific receptors. A cDNA encoding a rat SRIF receptor was identified by use of degenerate oligonucleotide primers and polymerase chain reaction amplification of cDNA prepared from transcripts expressed in rat brain. The complete cDNA encodes a protein of 391 amino acids with seven potential transmembrane domains. Expression of the cDNA product in transfected COS-7 cell lines provides the same high affinity of binding to [125I-Tyr11]SRIF-14 as that of rat cerebral cortex tissues. However, the binding of [125I-Tyr11]SRIF-14 to cloned rat SRIF receptor is not displaced by MK678, a SRIF analog that partially displaces [125I-Tyr11]SRIF-14 binding sites in membranes of rat cerebral cortex. Northern analysis and in situ hybridization indicate that mRNA (4.0 kilobases) for cloned rat SRIF receptor is preferentially expressed in rat brain regions such as cerebral cortex and hippocampus with no detectable expression in most peripheral organs. This pattern contrasts with the exclusive peripheral expression of a recently cloned human SRIF receptor. The cDNA probe of rat receptor detects mRNA from mouse brain but not from human cerebral cortex and cerebellum.
Amino Acid Sequence↗
Cloning, functional expression, and developmental regulation of a neuropeptide Y receptor from Drosophila melanogaster.
Neuropeptide Y, peptide YY, and pancreatic polypeptide are homologous 36-amino acid peptides that differ from most other peptide transmitters by having a relatively rigid conformation in aqueous solutions, defined as the pancreatic polypeptide fold, and a critical C-terminal tyrosine amide. These peptides serve as gastrointestinal hormones and neurotransmitters. A cDNA encoding a novel G protein-coupled receptor activated by neuropeptide Y was cloned from Drosophila by use of degenerate oligonucleotide primers and polymerase chain reaction amplification of cDNA prepared from transcripts expressed early in embryogenesis. The cDNA encodes a protein of 449 amino acids with the characteristics of a G protein-coupled receptor and shares significant amino acid identity with mammalian tachykinin receptors. When expressed in Xenopus oocytes, the PR4 protein is activated by mammalian neuropeptides in the order: peptide YY greater than neuropeptide Y much greater than pancreatic polypeptide. Northern analysis showed that PR4 receptor is expressed at equivalent levels in adult Drosophila head and body and that the expression of the PR4 receptor is regulated during development. The molecular characterization of this receptor should lead to a better understanding of the functional role of this important family of hormone receptors in adult organisms and during development.
Amino Acid Sequence↗
Toward the molecular structure of the mitochondrial channel, VDAC.
A summary is presented of the most recent information about the structure and mechanism of closure of the mitochondrial channel, VDAC. Considerable information has come from studies involving electron microscopy of two-dimensional crystals and from electrophysiological studies of wild-type channels and site-directed mutants. Available evidence points to a beta-barrel as the basic structural model for VDAC. Two models for voltage- or effector- induced closure have been proposed, the first involving removal of strands from the wall of the pore, the second invoking movement of protein domains into the lumen. Experimental strategies to resolve the actual mechanism are presented.
Amino Acid Sequence↗
Determination of the number of polypeptide subunits in a functional VDAC channel from Saccharomyces cerevisiae.
Genes encoding VDAC proteins containing specific site-directed amino acid alterations were introduced into wild-type Saccharomyces cerevisiae. The mutant VDAC proteins form channels with ion selectivities very different from that of the wild-type channel. Therefore, the resulting yeast strains express two different genes capable of coding for functional, yet distinct, VDAC channels. If VDAC were an oligomeric channel, analysis of VDAC from these strains should have revealed not only the presence of channels with wild-type or mutant selectivity but also channels with intermediate selectivities. While channels with wild-type and mutant selectivities were observed with approximately equal frequency, no channels with intermediate selectivity were observed. Sufficient observations were performed with two different mutant genes K61E.K65E and K19E.K61E) that the likelihood of having missed hybrid channels was less than 1 in 10(7). These findings favor the hypothesis that each functional VDAC channel is composed of a single 30-kDa polypeptide chain.
Fungal Proteins↗
Probing molecular structure and structural changes of voltage-gated channel by expressing mutant channels in yeast and reconstituting them into planar membranes.
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Amino Acid Sequence↗
Large scale rearrangement of protein domains is associated with voltage gating of the VDAC channel.
The VDAC channel of the mitochondrial outer membrane is voltage-gated like the larger, more complex voltage-gated channels of the plasma membrane. However, VDAC is a low molecular weight (30 kDa), abundant protein, which is readily purified and reconstituted, making it an ideal system for analyzing the molecular basis for ion selectivity and voltage-gating. We have probed the VDAC channel by subjecting the cloned yeast (S. cerevisiae) VDAC gene to site-directed mutagenesis and introducing the resulting mutant channels into planar bilayers to detect the effects of specific sequence changes on channel properties. This approach has allowed us to formulate and test a model of the open state structure of the VDAC channel. Now we have applied the same approach to analyzing the structure of the channel's low-conducting "closed state" (essentially closed to important metabolites). We have identified protein domains forming the wall of the closed conformation and domains that seem to be removed from the wall of the pore during channel closure. The latter can explain the reduction in pore diameter and volume and the dramatically altered channel selectivity resulting from the channel closure. This process would make a natural coupling between motion of the sensor and channel gating.
Amino Acid Sequence↗
Cytolytic T-cell activity against mycobacterial antigens in HIV.
OBJECTIVES: The declining incidence of tuberculosis (TB) in developed countries has recently been reversed with the advent of HIV disease. This study proposes to document in vitro T-cell responses to mycobacterial antigens in HIV-infected individuals. DESIGN: T-cell-mediated immunity is recognized as one of the mechanisms of defence against TB. The cellular immunodeficiency and the importance of TB in the context of HIV disease has prompted use of in vitro assays of lymphocyte proliferation and cytolytic activity. METHODS: Peripheral blood mononuclear cells isolated from 29 HIV-infected patients (four with recent TB) and 11 healthy volunteers were stimulated with purified protein derivative (PPD). The responding blasts were presented to autologous antigen-primed macrophages to measure specific cytolytic T-lymphocyte (CTL) activity in vitro. RESULTS: T-cell proliferative responses were significantly lower in late stages of HIV disease. The degree of specific CTL activity was higher in healthy individuals than in Centers for Disease Control (CDC) stage II-III (P = 0.037), and CDC stage IV patients (P = 0.029). CONCLUSIONS: The clinical presentation of TB tends to be typical in early stages of HIV disease and atypical in late stages. The manifestations reflect the degree of immunodepression. This study documents the declining proliferative and cytolytic T-cell-mediated responses in HIV patients with progression of immunodeficiency.
Adolescent↗
T-lymphocyte responses to Pneumocystis carinii in healthy and HIV-positive individuals.
Pneumocystis carinii pneumonia (PCP) is a well-recognized cause of morbidity in patients with impaired T-cell function. In this study of cellular immunity to P. carinii, peripheral blood mononuclear cells from 25 HIV antibody-positive (HIV+) patients and 11 healthy individuals were stimulated in vitro with P. carinii antigen. The responding T-cell blasts were cocultured with autologous P. carinii antigen-pulsed macrophages to measure P. carinii-specific cytolytic T-lymphocyte activity (CTL). T-cell blasts from two healthy donors were used to generate P. carinii-specific clones by limiting dilution. T cells from HIV+ patients proliferated less to P. carinii antigen than T cells from healthy volunteers. In contrast, the level of specific cytotoxicity was identical in all groups when equal numbers of CTLs were used. Within the group of symptomatic patients, CTL activity was higher in those with a history of PCP (p = 0.033). Pneumocystis carinii antigen-specific T-cell clones proved to be CD4+ and MHC class II restricted; six of eight clones tested showed P. carinii-specific cytolytic activity. Cell-mediated immune response to P. carinii in healthy individuals include CD4+, class II MHC-restricted T cells with P. carinii-specific cytotoxicity. There is an increasing loss of P. carinii-specific proliferative responses in HIV+ patients as disease progresses, but a cytotoxic response is still detected in the absence of proliferation.
Adolescent↗
Human cytochrome c oxidase subunit VIb: characterization and mapping of a multigene family.
We report the isolation and sequence of a human heart cDNA coding for cytochrome c oxidase (COX) subunit VIb (COX VIb). This cDNA extends 50 bp upstream from the region coding for the mature peptide. By Northern analysis, a single transcript of approx. 550 nucleotides (nt) has been identified in six human tissues. Southern analysis of human genomic DNA demonstrates the presence of multiple loci that show high homology to the cDNA. These loci cosegregate with either five or six different human chromosomes in human-rodent somatic cell hybrids. Using the COX6b cDNA, genomic sequences representing two of these loci have been isolated and characterized. The nt sequence analysis suggests that both loci represent COX6b pseudogenes.
Amino Acid Sequence↗
Drosophila stimulatory G protein alpha subunit activates mammalian adenylyl cyclase but interacts poorly with mammalian receptors: implications for receptor-G protein interaction.
Heterotrimeric guanine nucleotide binding proteins (G proteins) transduce signals from cell-surface receptors to intracellular effector proteins. Two forms of stimulatory G protein (Gs) alpha-like subunit have been described in Drosophila melanogaster. To examine the function of these subunits we have used vaccinia virus vectors to express both proteins in cyc- cells, a murine S49 cell line deficient for Gs alpha activity. Receptor-independent activation of each Drosophila Gs alpha has demonstrated that both forms are capable of activating mammalian adenylyl cyclase and thus have the activity expected of stimulatory G proteins. However, the Drosophila Gs alpha subunits interact poorly with mammalian Gs-coupled receptors. These observations have helped to identify a region of high variability in Gs alpha proteins that may be important for receptor interactions.
Adenylyl Cyclases↗
Cloning, heterologous expression and developmental regulation of a Drosophila receptor for tachykinin-like peptides.
We identified clones encoding a Drosophila receptor for tachykinin-like peptides by low stringency screening of an embryonic cDNA library with probes from the bovine substance K receptor. The cDNAs encode a seven transmembrane domain protein (DTKR) of 519 amino acids with 40-48% amino acid identity to mammalian tachykinin receptors within transmembrane regions. Xenopus oocytes injected with DTKR cRNAs showed selective responses to vertebrate substance P, its agonists and not to other vertebrate tachykinin peptides. These responses were eliminated by treatment of oocytes with pertussis toxin. In the adult fly, Northern and PCR analysis demonstrated preferential expression of DTKR in the head; in situ hybridization indicated that DTKR is accumulated in the cell bodies of neurons in the adult CNS. The levels of DTKR transcript are regulated during development. Northern and PCR amplification analysis showed that while DTKR transcripts are present at all stages, high levels of expression occur in later stages of embryogenesis (starting at 10-14 h), coinciding with the beginning of major periods of neural development. Whole mount embryo in situ hybridization demonstrated that DTKR is expressed at these later stages of embryogenesis (11-15 h) in the brain and in a specific subset of neurons in each neuromere of the developing ventral ganglion. The gene encoding DTKR was mapped by in situ hybridization to a single location at 99D on the right arm of chromosome 3. These observations demonstrate that the tachykinin family of peptide transmitters and their receptors represent an evolutionarily ancient form of cellular communication within the nervous system.(ABSTRACT TRUNCATED AT 250 WORDS)
Amino Acid Sequence↗
Restricted spatial and temporal expression of G-protein alpha subunits during Drosophila embryogenesis.
Of the known signal transduction mechanisms, the most evolutionarily ancient is mediated by a family of heterotrimeric guanine nucleotide binding proteins or G proteins. In simple organisms, this form of sensory transduction is used exclusively to convey signals of developmental consequence. In metazoan organisms, however, the developmental role of G-protein-coupled sensory transduction has been more difficult to elucidate because of the wide variety of signals (peptides, small molecules, odorants, hormones, etc.) that use this form of sensory transduction. We have begun to examine the role of G-protein-coupled signaling during development by investigating the expression during Drosophila embryogenesis of a limited set of G proteins. Since these proteins are a common component of all G-protein-coupled signaling systems, their developmental pattern of expression should indicate when and where programmed changes in gene activity are initiated by, or involve the participation of, G-protein-coupled signaling events. We have focused on the spatial and temporal expression pattern of three different Drosophila G-protein alpha subunits by northern blot analysis, in situ hybridization and immunocytochemistry using antibodies directed to peptides specifically found in each alpha subunit. From the spatial and temporal restriction of the expression of each protein, our results suggest that different forms of G-protein-coupled sensory transduction may mediate developmental interactions during both early and late stages of embryogenesis and may participate in a variety of specific developmental processes such as the establishment of embryonic position, the ontogeny of the nervous system and organogenesis.
Animals↗
In-vitro interaction of human macrophages with Pneumocystis carinii.
Pneumocystis carinii is an important opportunistic pathogen in patients with compromised cell-mediated immunity. T-cell and macrophage function are believed to be of prime importance in defence against this organism. The present ultrastructural study is aimed at the analysis of the interaction between human macrophages and P. carinii in vitro. Adherent peripheral blood mononuclear cells from healthy volunteers were exposed in vitro to Pneumocystis derived from lungs of steroid-treated rats. The macrophages were harvested at different intervals and studied by transmission and scanning electron microscopy. The material used for inoculation of macrophages was of identical morphology to previously described P. carinii. When mixed with Pneumocystis in vitro, the macrophages appeared to move towards the organism, extended pseudopods and ingested trophozoites and cysts. Within 24 h, intracellular Pneumocystis underwent progressive degeneration inside macrophage vacuoles. This study highlights the possible role of macrophages in host defence against P. carinii.
Animals↗
Selectivity changes in site-directed mutants of the VDAC ion channel: structural implications.
The gene encoding the yeast mitochondrial outer membrane channel VDAC was subjected to site-directed mutagenesis to change amino acids at 29 positions to residues differing in charge from the wild-type sequence. The mutant genes were then expressed in yeast, and the physiological consequences of single and multiple amino acid changes were assessed after isolation and insertion of mutant channels into phospholipid bilayers. Selectivity changes were observed at 14 sites distributed throughout the length of the molecule. These sites are likely to define the position of the protein walls lining the aqueous pore and hence, the transmembrane segments. These results have been used to develop a model of the open state of the channel in which each polypeptide contributes 12 beta strands and one alpha helix to form the aqueous transmembrane pathway.
Amino Acid Sequence↗
Immunolocalization of G protein alpha-subunits in the Drosophila CNS.
In order to uncover the role of G proteins in the integrative functioning and development of the nervous system, we have begun a multidisciplinary study of the G proteins present in the fruit fly, Drosophila melanogaster. In this report, we describe the distribution of 3 different G protein alpha-subunits in the adult Drosophila CNS as determined by immunocytochemical localization using affinity-purified antibodies generated to synthetic oligopeptide sequences unique to each alpha-subunit. Western blot analysis of membranes prepared from Drosophila heads indicates that antibodies specific for the Drosophila Go alpha and Gs alpha homologs recognize the appropriate protein species predicted by molecular cloning (Quan et al., 1989; Thambi et al., 1989). The Gi alpha homolog could not be detected in head membranes by Western blotting, consistent with the negligible levels of expression observed for Gi alpha on Northern blots of head mRNA (Provost et al., 1988). However, a Drosophila Gi alpha fusion protein could be detected by these antibodies following expression in E. coli. Immunolocalization studies revealed that the Go alpha and Gs alpha homologs are expressed at highest levels in neuropils and at intermediate levels in the cortex of all brain and thoracic ganglion areas. Only the lamina contained low levels of these alpha-subunits in the CNS. Additionally, Gs alpha appears to be associated with the cell membranes of neuronal cell bodies, while Go alpha has a more diffuse distribution, suggesting its presence in the cytoplasm as well as cell membranes. In contrast to the wide distribution of Go alpha and Gs alpha, Gi alpha has a surprisingly restricted distribution in the CNS. It is present at high levels only in photoreceptor cell terminations, glomerulae of the antennal lobes, and the ocellar retina. Little or no Gi alpha was detected in other brain regions or in the thoracic ganglion. Gi alpha, then, appears to be uniquely associated with some primary sensory afferents and their terminations, suggesting the presence of specific receptor and/or effector systems which mediate the transmission of primary sensory information in Drosophila.
Animals↗
[Tinea capitis in adults. Description of 3 cases].
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Age Factors↗