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Biomedical subjects

S Lev

Publications and source records attributed to S Lev.

At least 37 records · Page 2Linked to original sources
PubMed1995

Protein tyrosine kinase PYK2 involved in Ca(2+)-induced regulation of ion channel and MAP kinase functions.

The protein tyrosine kinase PYK2, which is highly expressed in the central nervous system, is rapidly phosphorylated on tyrosine residues in response to various stimuli that elevate the intracellular calcium concentration, as well as by protein kinase C activation. Activation of PYK2 leads to modulation of ion channel function and activation of the MAP kinase signalling pathway. PYK2 activation may provide a mechanism for a variety of short- and long-term calcium-dependent signalling events in the nervous system.

3T3 Cells↗
PubMed1995

The carbonic anhydrase domain of receptor tyrosine phosphatase beta is a functional ligand for the axonal cell recognition molecule contactin.

Receptor-type protein tyrosine phosphatase beta (RPTP beta) is expressed in the developing nervous system and contains a carbonic anhydrase (CAH) domain as well as a fibronectin type III repeat in its extracellular domain. Fusion proteins containing these domains were used to search for ligands of RPTP beta. The CAH domain bound specifically to a 140 kDa protein expressed on the surface of neuronal cells. Expression cloning in COS7 cells revealed that this protein is contactin, a GPI membrane-anchored neuronal cell recognition molecule. The CAH domain of RPTP beta induced cell adhesion and neurite growth of primary tectal neurons, and differentiation of neuroblastoma cells. These responses were blocked by antibodies against contactin, demonstrating that contactin is a neuronal receptor for RPTP beta. These experiments show that an individual domain of RPTP beta acts as a functional ligand for the neuronal receptor contactin. The interaction between contactin and RPTP beta may generate unidirectional or bidirectional signals during neural development.

Amino Acid Sequence↗
PubMed1995

The fourth immunoglobulin domain of the stem cell factor receptor couples ligand binding to signal transduction.

Receptor dimerization is ubiquitous to the action of all receptor tyrosine kinases, and in the case of dimeric ligands, such as the stem cell factor (SCF), it was attributed to ligand bivalency. However, by using a dimerization-inhibitory monoclonal antibody to the SCF receptor, we confined a putative dimerization site to the nonstandard fourth immunoglobulin-like domain of the receptor. Deletion of this domain not only abolished ligand-induced dimerization and completely inhibited signal transduction, but also provided insights into the mechanism of the coupling of ligand binding to dimer formation. These results identify an intrinsic receptor dimerization site and suggest that similar sites may exist in other receptors.

Animals↗
PubMed1995

Cerebrospinal fluid flow waveforms: analysis in patients with Chiari I malformation by means of gated phase-contrast MR imaging velocity measurements.

PURPOSE: To determine the features of the cardiac cycle-related cerebrospinal fluid (CSF) flow pulsations in with Chiari I malformation before and after decompressive surgery. MATERIALS AND METHODS: Fifteen patients and 23 healthy subjects underwent cine phase-contrast magnetic resonance imaging. Ten patients were also examined after decompressive surgery. Four regions of interest were selected in the upper cervical subarachnoid space and premedullary cistern. Velocity and fraction of the cardiac cycle waveforms were plotted, and amplitude, temporal, and CSF displacement parameters were assessed. RESULTS: In the Chiari I patients, impaired systolic and unaltered diastolic CSF flow pulsations immediately below the foramen magnum were identified. After surgery, the systolic flow pulsations immediately below the foramen magnum improved. Good correlation with post-surgical improvement was observed. CONCLUSION: CSF flow waveform analysis helps demonstrate abnormalities in CSF flow at the foramen magnum and the benefits of decompressive surgery in patients with the Chiari I malformation.

Adolescent↗
PubMed1995

Osteosarcoma: subtle, rare, and misleading plain film features.

Osteosarcoma is a malignant, mesenchymal, osteoid, and bone-forming tumor [1-3]. In most cases, typical radiographic features clearly illustrate the aggressive bone-forming nature of the lesion. These features include long-bone metaphyseal location, mixed areas of lysis and sclerosis, cortical destruction, periosteal new bone, and soft-tissue mass. However, numerous factors contribute to misleading radiographic patterns of osteosarcoma. They include histologic low-grade, lytic, or minimally sclerotic lesions, early detection, confinement to the intramedullary canal, benign-appearing periosteal reaction, rare intraosseous locations (subchondral, diaphyseal, intracortical), and rare skeletal sites (e.g., soft tissues, skull, ribs, tarsal bones). Secondary osteosarcomas, such as those arising from infarcts or fibrous dysplasia, may also produce a confusing radiologic picture. The purpose of this pictorial essay is to illustrate cases of osteosarcoma that proved to be difficult diagnostic dilemmas because of their subtle, rare, or misleading plain film features.

Adolescent↗
PubMed1994

Steel factor and c-kit protooncogene: genetic lessons in signal transduction.

Despite extensive research on the molecular mechanisms of signal transduction by growth factors and their oncogenic receptor tyrosine kinases, the physiological relevance of these pathways, especially in mammals, remains largely unknown. A unique exception is the Steel factor (SLF) and its c-kit-encoded receptor, because many natural germ line mutations of both the ligand and the receptor exist in mice. The protooncogene c-kit encodes a cell surface receptor that belongs to the immunoglobulin gene family and carries an intrinsic tyrosine kinase activity in its cytoplasmic portion. The precursor of the Kit ligand, SLF, is also a transmembrane protein that exists as a soluble factor as well as a cell surface protein. The interaction of Kit with SLF leads to receptor dimerization, kinase activation, and tyrosine phosphorylation of cytoplasmic proteins that contain Src homology 2 motifs. Various mutations in Kit and SLF result in a defective signaling pathway and underly the complex phenotypes of W and Sl mice, respectively. The early development of at least four cell lineages is affected. These are erythrocytes, melanocytes, germ cells, and mast cells. Correlation between the behavior of these lineages and specific mutations uncovered interesting physiological aspects of the mechanism of signal transduction by a polypeptide growth factor. These include the different degrees of severity of affected lineages, indications for distinct functions during early embryonic development and at late phases, the significance of synergy between a growth factor and lymphokines, the interaction between mutant and wild-type proteins in heterozygous animals, and the possibility that a surface-anchored ligand may act differently than a soluble factor. Predictably, the lessons learned with Kit and Sl mice will be widely relevant to other pairs of ligands and receptors that control the function of different cell lineages and physiological processes.

Animals↗
PubMed1993

Soluble c-kit proteins and antireceptor monoclonal antibodies confine the binding site of the stem cell factor.

The binding of the stem cell factor (SCF) to the c-kit-encoded receptor tyrosine kinase stimulates a variety of biochemical responses that culminate in cellular proliferation, migration, or survival. The extracellular domain of p145kit consists of five immunoglobulin-like domains. To confine the ligand binding site to this portion of the receptor we generated a panel of murine monoclonal antibodies (mAbs) to the Kit protein and identified two mAbs that efficiently displaced receptor-bound SCF and also inhibited proliferation of SCF-dependent human megakaryocytes. To map the epitopes of these mAbs we constructed and expressed soluble portions of the extracellular domain of Kit, which included either the two amino-terminal Ig-like domains (denoted Kit 1-2), three Ig-like domains (Kit 1-2-3), or the entire extracellular portion (Kit-X). All three recombinant proteins were recognized by the ligand inhibitory mAbs, suggesting that the SCF binding site resides in the amino-terminal half of the ecto-domain. Consistent with this conclusion, all of the soluble proteins inhibited SCF binding to Kit-expressing cells, and they also underwent specific covalent cross-linking to the radiolabeled ligand. However, whereas Kit 1-2-3 and Kit-X displayed comparable ligand affinities, deletion of the third Ig-like domain, in Kit 1-2, involved significant reduction in SCF binding. Hence, the binding site of SCF probably includes Ig-like domains 1 and 2, but structural determinants distal to this portion may also participate in ligand recognition.

Animals↗
PubMed1993

Structure-function analyses of the kit receptor for the steel factor.

Binding of the Steel factor (SLF) to the product of the c-kit proto-oncogene stimulates the receptor's intrinsic tyrosine kinase that phosphorylates a set of cytoplasmic signaling molecules. Germ-line mutations in the genes that encode the receptor or the ligand result in remarkably similar phenotypes that affect melanogenesis, erythropoiesis and gametogenesis in mice. We concentrated on the initial events of the signal transduction pathway that underlies these processes. The extracellular portion of Kit is comprised of five immunoglobulin-(Ig)-like domains. Ligand binding to this domain induces rapid and extensive dimerization of the receptor molecules in a mechanism that involves monovalent binding of the dimeric ligand, followed by an increase in receptors' affinity and gradual stabilization of the dimers. It thus appears that Kit has at least two functions: ligand binding and ligand-induced receptor dimerization, in addition to the kinase activity. Both functions are independent of the transmembrane and cytoplasmic domains, as a recombinant soluble ectodomain retained high affinity to SLF and ligand-dependent dimerization. In order to correlate these functions with specific structures, we employed ligand-competitive monoclonal antibodies, soluble deletion mutants of the ectodomain and chimeric human-mouse Kit proteins. These approaches indicated that the N-terminal three Ig-like domains constitute the binding site, whose core is the second domain. Further experiments suggested that a putative dimerization site is distinct from the binding cleft and may be located on the fourth Ig-like domain.

Animals↗
PubMed1993

Interspecies molecular chimeras of kit help define the binding site of the stem cell factor.

The extracellular portion of the kit-encoded receptor for the stem cell factor (SCF) comprises five immunoglobulin (Ig)-like domains. To localize the ligand recognition site, we exploited the lack of binding of human SCF to the murine receptor by using human-mouse hybrids of Kit and species-specific monoclonal antibodies (MAbs) that inhibit ligand binding. Replacement of the three N-terminal Ig-like domains of the murine Kit with the corresponding portion of the human receptor conferred upon the chimeric receptor high-affinity binding of the human ligand as well as of human-specific ligand-inhibitory MAbs. By constructing five chimeric murine Kit proteins which individually contain each of these three human Ig-like units or pairs of them, we found that the second human domain confers upon the mouse Kit high-affinity binding of the human ligand and also binding of species-specific SCF-competitive MAbs. Nevertheless, the flanking Ig-like domains also affect high-affinity recognition of SCF. Moreover, it appears that the determinants that define ligand specificity of the murine and the human receptors do not structurally coincide. This observation allowed us to identify a chimeric receptor that displayed a dual specificity; namely, it bound with high affinity either the human or the murine SCF molecules and reacted with mouse- as well as human-specific ligand-inhibitory MAbs. Conversely, another chimera, which included all of the five Ig-like domains, bound neither ligand. In conclusion, interdomain packing involving the second Ig-like domain of human Kit and noncontiguous structural motifs of the receptor are involved in SCF recognition.

Animals↗
PubMed1992

Dimerization and activation of the kit receptor by monovalent and bivalent binding of the stem cell factor.

The protooncogene c-kit encodes a tyrosine kinase receptor for the stem cell factor (SCF). Mutants of c-kit were shown to confer a pleiotropic defective phenotype and often display negative dominance in heterozygous mice. To explore the involvement of receptor dimerization in this genetic phenomenon, we employed both a human ligand, which does not recognize the murine receptor, and a rodent SCF, which binds to the human receptor with 100-fold reduced affinity as compared with human SCF. SCF binding to living cells was found to induce rapid and complete receptor dimerization that involved activation of the catalytic tyrosine kinase function. Although receptor dimerization can be attributed to the dimeric nature of the ligand, no dissociation of Kit dimers occurred at high excess of SCF, suggesting that receptor-receptor interactions are also involved in dimer stabilization. This was supported by in vitro formation of heterodimers between the human and murine Kit proteins through monovalent binding of species-specific human SCF. By coexpression of human and mouse Kit in murine fibroblasts, we found that receptor heterodimerization in living cells involved an increase in the affinity of human Kit for rat SCF and also an accelerated rate of receptor down-regulation. When a human Kit mutant lacking the kinase insert domain was coexpressed with the murine wild-type receptor, we observed a significant decrease in both the activation of the intact tyrosine kinase and its coupling to an effector protein, namely phosphatidylinositol 3'-kinase. Our results favor a receptor activation model that assumes an initial step of monovalent ligand binding, followed by an intermediate receptor dimer bound by one arm of the ligand molecule. This model predicts the existence of an intrinsic receptor dimerization site and provides a structural basis for genetic dominance of mutant SCF receptors.

Animals↗
PubMed1992

A recombinant ectodomain of the receptor for the stem cell factor (SCF) retains ligand-induced receptor dimerization and antagonizes SCF-stimulated cellular responses.

The stem cell factor (SCF) is a polypeptide ligand that is essential for the development of germ cells, hematopoietic progenitor cells, and melanocyte precursors. It binds to a tyrosine kinase membrane receptor that is encoded by the c-kit proto-oncogene. We have constructed an expression vector that directs the synthesis of the entire extracellular ligand-binding domain of the Kit/SCF receptor. When expressed and amplified in Chinese hamster ovary cells, a secreted 90-kDa glycoprotein could be harvested from the growth medium of the cells in a soluble form. This extracellular portion of the Kit/SCF receptor, denoted Kit-X, was recognized by antibodies specific to the SCF receptor; and when injected into animals, it raised antibodies that were reactive with the complete membrane form of the receptor. Direct binding and covalent cross-linking of radiolabeled SCF showed that Kit-X fully retained high affinity ligand binding and also underwent efficient dimerization in the presence of the ligand. The capacity of Kit-X to act as an antagonist of SCF was assayed on cultured cells that overexpress the receptor. Simultaneous addition of SCF and Kit-X to these cells resulted in a stoichiometric inhibition of SCF binding and a consequent decrease in autophosphorylation of the SCF receptor on tyrosine residues. The inhibition extended to later SCF-mediated responses, including the association of the receptor with phosphatidylinositol 3'-kinase and coupling to the Raf1 protein kinase. These results indicate that the recombinant ectodomain of the Kit-SCF receptor can be used as a specific antagonist of SCF actions and may enable detailed molecular analysis of ligand-receptor interactions.

Amino Acid Sequence↗
PubMed1992

Interkinase domain of kit contains the binding site for phosphatidylinositol 3' kinase.

Our previous analysis of the signal transduction pathway used by the c-kit-encoded receptor for the stem cell factor (SCF) indicated efficient coupling to the type I phosphatidylinositol 3' kinase (PI3K). In an attempt to localize the receptor's site of interaction with PI3K, we separately deleted either the noncatalytic 68-amino-acid-long interkinase domain or the carboxyl-terminal portion distal to the catalytic sequences. Loss of ligand-induced association of PI3K with the former deletion mutant and retention of the PI3K association by the carboxyl-terminally deleted receptor implied interactions of PI3K with the kinase insert. This was further supported by partial inhibition of the association by an anti-peptide antibody directed against the kinase insert and lack of effect of an antibody directed to the carboxyl tail of the SCF receptor. A bacterially expressed kinase insert domain was used as a fusion protein to directly test its presumed function as a PI3K association site. This protein bound PI3K from cell lysate as demonstrated by PI3K activity and by an associated phosphoprotein of 85 kDa. The association was dependent on phosphorylation of the tyrosine residues on the expressed kinase insert. On the basis of these observations, we conclude that the kinase insert domain of the SCF receptor selectively interacts with the p85 regulatory subunit of PI3K and that this association requires phosphorylation of tyrosine residues in the kinase insert region, with apparently no involvement of the bulk cytoplasmic structure or tyrosine kinase function of the receptor.

1-Phosphatidylinositol 4-Kinase↗
PubMed1991

A specific combination of substrates is involved in signal transduction by the kit-encoded receptor.

The kit protooncogene encodes a transmembrane tyrosine kinase related to the receptors for the platelet derived growth factor (PDGF-R) and the macrophage growth factor (CSF1-R), and was very recently shown to bind a stem cell factor. To compare signal transduction by the kit kinase with signaling by homologous receptors we constructed a chimeric protein composed of the extracellular domain of the epidermal growth factor receptor (EGF-R) and the transmembrane and cytoplasmic domains of kit. We have previously shown that the chimeric receptor transmits potent mitogenic and transforming signals in response to the heterologous ligand. Here we demonstrate that upon ligand binding, the ligand-receptor complex undergoes endocytosis and degradation and induces short- and long-term cellular effects. Examination of the signal transduction pathway revealed that the activated kit kinase strongly associates with phosphatidylinositol 3'-kinase activity and a phosphoprotein of 85 kd. In addition, the ligand-stimulated kit kinase is coupled to modifications of phospholipase C gamma and the Raf1 protein kinase. However, it does not lead to a significant change in the production of inositol phosphate. Comparison of our results with the known signaling pathways of PDGF-R and CSF1-R suggests that each receptor is coupled to a specific combination of signal transducers.

Animals↗
PubMed1990

Receptor functions and ligand-dependent transforming potential of a chimeric kit proto-oncogene.

The c-kit proto-oncogene, the cellular homolog of the transforming gene of a feline retrovirus, encodes a transmembrane tyrosine kinase homologous to receptors for growth factors. To study the cellular function of c-kit, we constructed a chimeric molecule composed of the extracellular portion of the receptor for epidermal growth factor (EGF) and the transmembrane and cytoplasmic domains of p145kit. The hybrid molecule was properly expressed in murine fibroblasts and displayed specific binding of EGF (Kd, 3 x 10(-8) M). Activation of the chimeric receptor by EGF stimulated the tyrosine kinase activity of kit and led to the generation of a potent mitogenic signal. Moreover, cells expressing the chimeric receptor acquired a transformed phenotype once they were stimulated with the heterologous ligand.

Amino Acid Sequence↗
PubMed1988

Effect of ambulatory blood pressure monitoring on the diagnosis and cost of treatment for mild hypertension.

Detection of mild hypertension by a small number of casual blood pressures may be inaccurate for the determination of average blood pressure. Nonetheless, casual pressures remain the basis for the diagnosis and treatment of hypertensive patients. We compared casual and noninvasive ambulatory blood pressure monitoring in a consecutive series of 60 subjects evaluated for possible mild hypertension on the basis of casual pressures. Ambulatory blood pressure monitoring was performed on days of usual activity. Correlations between casual systolic and average ambulatory systolic pressures or casual diastolic and average ambulatory diastolic pressures were not significant. Nearly half of the subjects had average ambulatory systolic pressures less than 130 mm Hg. Sixty percent had average ambulatory diastolic pressures less than 85 mm Hg. Nearly 40% had both systolic and diastolic pressures less than those limits. A preliminary analysis of the effects of these results on the short-term cost of antihypertensive treatment was made, assuming that treatment could be withheld from those with average ambulatory pressures less than 130/85 mm Hg. This approach suggests that ambulatory blood pressure monitoring need not increase overall cost, if the results of this evaluation are used in the decision to treat.

Adult↗
PubMed1985

On the vertical caloric nystagmus.

Eye movements in response to caloric stimuli are recorded traditionally in the horizontal plane only. Twin channel electronystagmographic traces of 124 patients undergoing evaluation of vertigo were reviewed. Thirty traces (24%) were found to include a "true" vertical component to the evoked nystagmus. There was no correlation between the existence and the nature of any vestibular pathology and the presence of the vertical nystagmus. Thus, a vertical component to the caloric nystagmus (the so-called "perverted" caloric nystagmus) is not necessarily a pathological sign, and can be a normal variant, caused by excitation of the vertical semicircular canal - most probably the posterior one. The mechanisms of heat transfer in the temporal bone and the source of this vertical caloric nystagmus are discussed in the context of the published literature.

Caloric Tests↗
PubMed1984

Square wave jerks--incidence, characteristic, and significance.

Square wave jerks (SWJ) are saccadic eye movements which, when recorded with open eyes are considered to be a pathological sign, caused by fixation instability, and pointing to a central neurological lesion. In reviewing 429 electronystagmographic traces, 85 (20%) were found to include the pattern of SWJ. Among them, in 38 traces, SWJ have been recorded with open eyes during fixation. Many of these cases were found to suffer from definite peripheral vestibular pathology, and some of them did not reveal any sign of neurological pathology in the clinical evaluation and by electronystagmography. Thus, SWJ are non-specific rapid eye movements, which do not necessarily point to neurological pathology. The literature is reviewed and the possible neural mechanism for the production of SWJ is discussed.

Adult↗