bluuebunny/crossref_metadata_2025 huggingface dataset:
Influence of Polymorphism in the Genes for the Interleukin (IL)-1 Receptor Antagonist and IL-1β on Tuberculosis
Authors: Wilkinson Robert J. , Patel Punita , Llewelyn Martin , Hirsch Christina S. , Pasvol Geoffrey , Snounou Georges , Davidson Robert N. , Toossi Zahra
Published: July 2002

Several lines of evidence suggest that host genetic factors controlling the immune response influence infection by Mycobacterium tuberculosis. The proinflammatory cytokine interleukin (IL)-1β and its antagonist, IL-1Ra (IL-1 receptor agonist), are strongly induced by M. tuberculosis and are encoded by polymorphic genes. The induction of both IL-1Ra mRNA and secreted protein by M. tuberculosis in IL-1Ra allele A2–positive (IL-1Ra A2+) healthy subjects was 1.9-fold higher than in IL-1Ra A2− subjects. The M. tuberculosis–induced expression of mRNA for IL-1β was higher in subjects of the IL-1β (+3953) A1+ haplotype (P = 0.04). The molar ratio of IL-1Ra/IL-1β induced by M. tuberculosis was markedly higher in IL-1Ra A2+ individuals (P < 0.05), with minor overlap between the groups, reflecting linkage between the IL-1Ra A2 and IL-1β (+3953) A2 alleles. In M. tuberculosis–stimulated peripheral blood mononuclear cells, the addition of IL-4 increased IL-1Ra secretion, whereas interferon γ increased and IL-10 decreased IL-1β production, indicative of a differential influence on the IL-1Ra/IL-1β ratio by cytokines. In a study of 114 healthy purified protein derivative–reactive subjects and 89 patients with tuberculosis, the frequency of allelic variants at two positions (−511 and +3953) in the IL-1β and IL-1Ra genes did not differ between the groups. However, the proinflammatory IL-1Ra A2−/IL-1β (+3953) A1+ haplotype was unevenly distributed, being more common in patients with tuberculous pleurisy (92%) in comparison with healthy M. tuberculosis–sensitized control subjects or patients with other disease forms (57%, P = 0.028 and 56%, P = 0.024, respectively). Furthermore, the IL-1Ra A2+ haplotype was associated with a reduced Mantoux response to purified protein derivative of M. tuberculosis: 60% of tuberculin-nonreactive patients were of this type. Thus, the polymorphism at the IL-1 locus influences the cytokine response and may be a determinant of  delayed-type hypersensitivity and disease expression in human tuberculosis.
Glycosylphosphatidylinositol Anchors ofPlasmodium falciparum
Authors: Naik Ramachandra S. , Branch OraLee H. , Woods Amina S. , Vijaykumar Matam , Perkins Douglas J. , Nahlen Bernard L. , Lal Altaf A. , Cotter Robert J. , Costello Catherine E. , Ockenhouse Christian F. , Davidson Eugene A. , Gowda D. Channe
Published: July 2002

Induction of proinflammatory cytokine responses by glycosylphosphatidylinositols (GPIs) of intraerythrocytic Plasmodium falciparum is believed to contribute to malaria pathogenesis. In this study, we purified the GPIs of P. falciparum to homogeneity and determined their structures by biochemical degradations and mass spectrometry. The parasite GPIs differ from those of the host in that they contain palmitic (major) and myristic (minor) acids at C-2 of inositol, predominantly C18:0 and C18:1 at sn-1 and sn-2, respectively, and do not contain additional phosphoethanolamine substitution in their core glycan structures. The purified parasite GPIs can induce tumor necrosis factor α release from macrophages. We also report a new finding that adults who have resistance to clinical malaria contain high levels of persistent anti-GPI antibodies, whereas susceptible children lack or have low levels of short-lived antibody response. Individuals who were not exposed to the malaria parasite completely lack anti-GPI antibodies. Absence of a persistent anti-GPI antibody response correlated with malaria-specific anemia and fever, suggesting that anti-GPI antibodies provide protection against clinical malaria. The antibodies are mainly directed against the acylated phosphoinositol portion of GPIs. These results are likely to be valuable in studies aimed at the evaluation of chemically defined structures for toxicity versus immunogenicity with implications for the development of GPI-based therapies or vaccines.
Function of Cd3ε-Mediated Signals in T Cell Development
Authors: Sommers Connie L. , Dejarnette Jan B. , Huang Kun , Lee Jan , El-Khoury Dalal , Shores Elizabeth W. , Love Paul E.
Published: July 2002

The T cell antigen receptor (TCR) and pre-TCR complexes are composed of multiple signal-transducing subunits (CD3γ, CD3δ, CD3ε, and ζ) that each contain one or more copies of a semiconserved functional motif, the immunoreceptor tyrosine-based activation motif (ITAM). Although biochemical studies indicate that individual TCR-ITAMs may bind selectively or with different affinity to various effector molecules, data from other experiments suggest that at least some ITAMs are functionally equivalent. In this study, we examined the role of CD3ε ITAM-mediated signals in T cell development by genetically reconstituting CD3ε-deficient mice with transgenes encoding either wild-type or ITAM-mutant (signaling defective) forms of the protein. The results demonstrate that signals transduced by CD3ε are not specifically required for T cell maturation but instead contribute quantitatively to TCR signaling in a manner similar to that previously observed for ζ chain. Unexpectedly, analysis of TCR-transgenic/CD3ε-mutant mice reveals a potential role for CD3ε signals in T cell survival.
Analysis of Thymocyte Development Reveals That the Gtpase Rhoa Is a Positive Regulator of T Cell Receptor Responses in Vivo
Authors: Corre Isabelle , Gomez Manuel , Vielkind Susina , Cantrell Doreen A.
Published: July 2002

Loss of function of the guanine nucleotide binding protein RhoA blocks pre-T cell differentiation and survival indicating that this GTPase is a critical signaling molecule during early thymocyte development. Previous work has shown that the Rho family GTPase Rac-1 can initiate changes in actin dynamics necessary and sufficient for pre-T cell development. The present data now show that Rac-1 actions in pre-T cells require Rho function but that RhoA cannot substitute for Rac-1 and induce the actin cytoskeletal changes necessary for pre-T cell development. Activation of Rho is thus not sufficient to induce pre-T cell differentiation or survival in the absence of the pre-T cell receptor (TCR). The failure of RhoA activation to impact on pre-TCR–mediated signaling was in marked contrast to its actions on T cell responses mediated by the mature TCR α/β complex. Cells expressing active RhoA were thus hyperresponsive in the context of TCR-induced proliferation in vitro and in vivo showed augmented positive selection of thymocytes expressing defined TCR complexes. This reveals that RhoA function is not only important for pre-T cells but also plays a role in determining the fate of mature T cells.
Block of N-type Calcium Channels in Chick Sensory Neurons by External Sodium
Authors: Polo-Parada Luis , Korn Stephen J.
Published: July 2002

L-type Ca2+ channels select for Ca2+ over sodium Na+ by an affinity-based mechanism. The prevailing model of Ca2+ channel permeation describes a multi-ion pore that requires pore occupancy by at least two Ca2+ ions to generate a Ca2+ current. At [Ca2+] < 1 μM, Ca2+ channels conduct Na+. Due to the high affinity of the intrapore binding sites for Ca2+ relative to Na+, addition of μM concentrations of Ca2+ block Na+ conductance through the channel. There is little information, however, about the potential for interaction between Na+ and Ca2+ for the second binding site in a Ca2+ channel already occupied by one Ca2+. The two simplest possibilities, (a) that Na+ and Ca2+ compete for the second binding site or (b) that full time occupancy by one Ca2+ excludes Na+ from the pore altogether, would imply considerably different mechanisms of channel permeation. We are studying permeation mechanisms in N-type Ca2+ channels. Similar to L-type Ca2+ channels, N-type channels conduct Na+ well in the absence of external Ca2+. Addition of 10 μM Ca2+ inhibited Na+ conductance by 95%, and addition of 1 mM Mg2+ inhibited Na+ conductance by 80%. At divalent ion concentrations of 2 mM, 120 mM Na+ blocked both Ca2+ and Ba2+ currents. With 2 mM Ba2+, the IC50 for block of Ba2+ currents by Na+ was 119 mM. External Li+ also blocked Ba2+ currents in a concentration-dependent manner, with an IC50 of 97 mM. Na+ block of Ba2+ currents was dependent on [Ba2+]; increasing [Ba2+] progressively reduced block with an IC50 of 2 mM. External Na+ had no effect on voltage-dependent activation or inactivation of the channel. These data suggest that at physiological concentrations, Na+ and Ca2+ compete for occupancy in a pore already occupied by a single Ca2+. Occupancy of the pore by Na+ reduced Ca2+ channel conductance, such that in physiological solutions, Ca2+ channel currents are between 50 and 70% of maximal.
Calcium Modulation of Ligand Affinity in the Cyclic GMP–gated Ion Channels of Cone Photoreceptors
Authors: Hackos David H. , Korenbrot Juan I.
Published: July 2002

To investigate modulation of the activation of cGMP-gated ion channels in cone photoreceptors, we measured currents in membrane patches detached from the outer segments of single cones isolated from striped bass retina. The sensitivity of these channels to activation by cGMP depends on the history of exposure to divalent cations of the membrane's cytoplasmic surface. In patches maintained in 20 μM Ca++ and 100 μM Mg++ after excision, the current amplitude dependence on cGMP is well described by a Hill equation with average values of K1/2, the concentration necessary to activate half the maximal current, of 86 μM and a cooperativity index, n, of 2.57. Exposing the patch to a solution free of divalent cations irreversibly increases the cGMP sensitivity; the average value of K1/2 shifts to 58.8 μM and n shifts to 1.8. Changes in cGMP sensitivity do not affect other functional parameters of the ion channels, such as the interaction and permeation of mono- and divalent cations. Modulation of cGMP activation depends on the action of an endogenous factor that progressively dissociates from the channel as Ca++ concentration is lowered below 1 μM. The activity of the endogenous modulator is not well mimicked by exogenously added calmodulin, although this protein competes with the endogenous modulator for a common binding site. Thus, the modulation of cGMP affinity in cones depends on the activity of an unidentified molecule that may not be calmodulin.
Mechanism of Ion Permeation in Skeletal Muscle Chloride Channels
Authors: Fahlke Christoph , Dürr Christine , George Alfred L.
Published: July 2002

Voltage-gated Cl− channels belonging to the ClC family exhibit unique properties of ion permeation and gating. We functionally probed the conduction pathway of a recombinant human skeletal muscle Cl− channel (hClC-1) expressed both in Xenopus oocytes and in a mammalian cell line by investigating block by extracellular or intracellular I− and related anions. Extracellular and intracellular I− exert blocking actions on hClC-1 currents that are both concentration and voltage dependent. Similar actions were observed for a variety of other halide (Br−) and polyatomic (SCN−, NO3−, CH3SO3−) anions. In addition, I− block is accompanied by gating alterations that differ depending on which side of the membrane the blocker is applied. External I− causes a shift in the voltage-dependent probability that channels exist in three definable kinetic states (fast deactivating, slow deactivating, nondeactivating), while internal I− slows deactivation. These different effects on gating properties can be used to distinguish two functional ion binding sites within the hClC-1 pore. We determined KD values for I− block in three distinct kinetic states and found that binding of I− to hClC-1 is modulated by the gating state of the channel. Furthermore, estimates of electrical distance for I− binding suggest that conformational changes affecting the two ion binding sites occur during gating transitions. These results have implications for understanding mechanisms of ion selectivity in hClC-1, and for defining the intimate relationship between gating and permeation in ClC channels.
The Voltage Dependence of a Cloned Mammalian Renal Type II Na+/Pi Cotransporter (NaPi-2)
Authors: Forster Ian , Hernando Nati , Biber Jürg , Murer Heini
Published: July 2002

The voltage dependence of the rat renal type II Na+/Pi cotransporter (NaPi-2) was investigated by expressing NaPi-2 in Xenopus laevis oocytes and applying the two-electrode voltage clamp. In the steady state, superfusion with inorganic phosphate (Pi) induced inward currents (Ip) in the presence of 96 mM Na+ over the potential range −140 ≤ V ≤ +40 mV. With Pi as the variable substrate, the apparent affinity constant (KmPi) was strongly dependent on Na+, increasing sixfold for a twofold reduction in external Na+. KmPi increased with depolarizing voltage and was more sensitive to voltage at reduced Na+. The Hill coefficient was close to unity and the predicted maximum Ip (Ipmax) was 40% smaller at 50 mM Na+. With Na+ as the variable substrate, KmNa was weakly dependent on both Pi and voltage, the Hill coefficient was close to 3 and Ipmax was independent of Pi at −50 mV. The competitive inhibitor phosphonoformic acid suppressed the steady state holding current in a Na+-dependent manner, indicating the existence of uncoupled Na+ slippage. Voltage steps induced pre–steady state relaxations typical for Na+-coupled cotransporters. NaPi-2-dependent relaxations were quantitated by a single, voltage-dependent exponential. At 96 mM Na+, a Boltzmann function was fit to the steady state charge distribution (Q-V) to give a midpoint voltage (V0.5) in the range −20 to −50 mV and an apparent valency of ∼0.5 e−. V0.5 became more negative as Na+ was reduced. Pi suppressed relaxations in a dose-dependent manner, but had little effect on their voltage dependence. Reducing external pH shifted V0.5 to depolarizing potentials and suppressed relaxations in the absence of Na+, suggesting that protons interact with the unloaded carrier. These findings were incorporated into an ordered kinetic model whereby Na+ is the first and last substrate to bind, and the observed voltage dependence arises from the unloaded carrier and first Na+ binding step.
Recombinant pICln Forms Highly Cation-selective Channels when Reconstituted into Artificial and Biological Membranes
Authors: Li Canhui , Breton Sylvie , Morrison Rebecca , Cannon Carolyn L. , Emma Francesco , Sanchez-Olea Roberto , Bear Christine , Strange Kevin
Published: July 2002

pICln has been proposed to be the swelling-activated anion channel responsible for ICl, swell, or a channel regulator. We tested the anion channel hypothesis by reconstituting recombinant pICln into artificial and biological membranes. Single channels were observed when pICln was reconstituted into planar lipid bilayers. In the presence of symmetrical 300 mM KCl, the channels had a high open probability and a slope conductance of 48 pS, and were outwardly rectifying. Reduction of trans KCl to 50 mM shifted the reversal potential by −31.2 ± 0.06 mV, demonstrating that the channel is at least seven times more selective for cations than for anions. Consistent with this finding, channel conductance was unaffected by substitution of Cl− with glutamate, but was undetectable when K+ was replaced by N-methyl-d-glucamine. Reconstitution of pICln into liposomes increased 86Rb+ uptake by three- to fourfold, but had no effect on 36Cl− uptake. Phosphorylation of pICln with casein kinase II or mutation of G54, G56, and G58 to alanine decreased channel open probability and 86Rb+ uptake. When added to the external medium bathing Sf9 cells, pICln inserted into the plasma membrane and increased cell cation permeability. Taken together, these observations demonstrate that channel activity is due to pICln and not minor contaminant proteins. However, these findings do not support the hypothesis that pICln is the anion-selective ICl, swell channel. The observed cation channel activity may reflect an as yet to be defined physiological function of pICln, or may be a consequence of in vitro reconstitution of purified, recombinant protein.
Location of the Permeation Pathway in the Recombinant Type 1 Inositol 1,4,5-Trisphosphate Receptor
Authors: Ramos-Franco Josefina , Galvan Daniel , Mignery Gregory A. , Fill Michael
Published: July 2002

The inositol 1,4,5-trisphosphate receptor (InsP3R) forms ligand-regulated intracellular Ca2+ release channels in the endoplasmic reticulum of all mammalian cells. The InsP3R has been suggested to have six transmembrane regions (TMRs) near its carboxyl terminus. A TMR-deletion mutation strategy was applied to define the location of the InsP3R pore. Mutant InsP3Rs were expressed in COS-1 cells and single channel function was defined in planar lipid bilayers. Mutants having the fifth and sixth TMR (and the interceding lumenal loop), but missing all other TMRs, formed channels with permeation properties similar to wild-type channels (gCs = 284; gCa = 60 pS; PCa/PCs = 6.3). These mutant channels bound InsP3, but ligand occupancy did not regulate the constitutively open pore (Po > 0.80). We propose that a region of 191 amino acids (including the fifth and sixth TMR, residues 2398–2589) near the COOH terminus of the protein forms the InsP3R pore. Further, we have produced a constitutively open InsP3R pore mutant that is ideal for future site-directed mutagenesis studies of the structure–function relationships that define Ca2+ permeation through the InsP3R channel.
Gat1 (Gaba:Na+:Cl−) Cotransport Function
Authors: Lu Chin-Chih , Hilgemann Donald W.
Published: July 2002

To explain cotransport function, the “alternating access” model requires that conformational changes of the empty transporter allow substrates to bind alternatively on opposite membrane sides. To test this principle for the GAT1 (GABA:Na+:Cl−) cotransporter, we have analyzed how its charge-moving partial reactions depend on substrates on both membrane sides in giant Xenopus oocyte membrane patches. (a) “Slow” charge movements, which require extracellular Na+ and probably reflect occlusion of Na+ by GAT1, were defined in three ways with similar results: by application of the high-affinity GAT1 blocker (NO-711), by application of a high concentration (120 mM) of cytoplasmic Cl−, and by removal of extracellular Na+ via pipette perfusion. (b) Three results indicate that cytoplasmic Cl− and extracellular Na+ bind to the transporter in a mutually exclusive fashion: first, cytoplasmic Cl− (5–140 mM) shifts the voltage dependence of the slow charge movement to more negative potentials, specifically by slowing its “forward” rate (i.e., extracellular Na+ occlusion); second, rapid application of cytoplasmic Cl− induces an outward current transient that requires extracellular Na+, consistent with extracellular Na+ being forced out of its binding site; third, fast charge-moving reactions, which can be monitored as a capacitance, are “immobilized” both by cytoplasmic Cl− binding and by extracellular Na+ occlusion (i.e., by the slow charge movement). (c) In the absence of extracellular Na+, three fast (submillisecond) charge movements have been identified, but no slow components. The addition of cytoplasmic Cl− suppresses two components (τ < 1 ms and 13 μs) and enables a faster component (τ < 1 μs). (d) We failed to identify charge movements of fully loaded GAT1 transporters (i.e., with all substrates on both sides). (e) Under zero-trans conditions, inward (forward) GAT1 current shows pronounced pre–steady state transients, while outward (reverse) GAT1 current does not. (f) Turnover rates for reverse GAT1 transport (33°C), calculated from the ratio of steady state current magnitude to total charge movement magnitude, can exceed 60 s−1 at positive potentials.
Role of Aquaporin-4 in Airspace-to-Capillary Water Permeability in Intact Mouse Lung Measured by a Novel Gravimetric Method
Authors: Song Yuanlin , Ma Tonghui , Matthay Michael A. , Verkman A.S.
Published: July 2002

The mammalian peripheral lung contains at least three aquaporin (AQP) water channels: AQP1 in microvascular endothelia, AQP4 in airway epithelia, and AQP5 in alveolar epithelia. In this study, we determined the role of AQP4 in airspace-to-capillary water transport by comparing water permeability in wild-type mice and transgenic null mice lacking AQP1, AQP4, or AQP1/AQP4 together. An apparatus was constructed to measure lung weight continuously during pulmonary artery perfusion of isolated mouse lungs. Osmotically induced water flux (Jv) between the airspace and capillary compartments was measured from the kinetics of lung weight change in saline-filled lungs in response to changes in perfusate osmolality. Jv in wild-type mice varied linearly with osmotic gradient size (4.4 × 10−5 cm3 s−1 mOsm−1) and was symmetric, independent of perfusate osmolyte size, weakly temperature dependent, and decreased 11-fold by AQP1 deletion. Transcapillary osmotic water permeability was greatly reduced by AQP1 deletion, as measured by the same method except that the airspace saline was replaced by an inert perfluorocarbon. Hydrostatically induced lung edema was characterized by lung weight changes in response to changes in pulmonary arterial inflow or pulmonary venous outflow pressure. At 5 cm H2O outflow pressure, the filtration coefficient was 4.7 cm3 s−1 mOsm−1 and reduced 1.4-fold by AQP1 deletion. To study the role of AQP4 in lung water transport, AQP1/AQP4 double knockout mice were generated by crossbreeding of AQP1 and AQP4 null mice. Jv were (cm3 s−1 mOsm−1 × 10−5, SEM, n = 7–12 mice): 3.8 ± 0.4 (wild type), 0.35 ± 0.02 (AQP1 null), 3.7 ± 0.4 (AQP4 null), and 0.25 ± 0.01 (AQP1/AQP4 null). The significant reduction in Pf in AQP1 vs. AQP1/AQP4 null mice was confirmed by an independent pleural surface fluorescence method showing a 1.6 ± 0.2-fold (SEM, five mice) reduced Pf in the AQP1/AQP4 double knockout mice vs. AQP1 null mice. These results establish a simple gravimetric method to quantify osmosis and filtration in intact mouse lung and provide direct evidence for a contribution of the distal airways to airspace-to-capillary water transport.
Dynamics of Signaling between Ca2+ Sparks and Ca2+- Activated K+ Channels Studied with a Novel Image-Based Method for Direct Intracellular Measurement of Ryanodine Receptor Ca2+ Current
Authors: ZhuGe Ronghua , Fogarty Kevin E. , Tuft Richard A. , Lifshitz Lawrence M. , Sayar Kemal , Walsh John V.
Published: July 2002

Ca2+ sparks are highly localized cytosolic Ca2+ transients caused by a release of Ca2+ from the sarcoplasmic reticulum via ryanodine receptors (RyRs); they are the elementary events underlying global changes in Ca2+ in skeletal and cardiac muscle. In smooth muscle and some neurons, Ca2+ sparks activate large conductance Ca2+-activated K+ channels (BK channels) in the spark microdomain, causing spontaneous transient outward currents (STOCs) that regulate membrane potential and, hence, voltage-gated channels. Using the fluorescent Ca2+ indicator fluo-3 and a high speed widefield digital imaging system, it was possible to capture the total increase in fluorescence (i.e., the signal mass) during a spark in smooth muscle cells, which is the first time such a direct approach has been used in any system. The signal mass is proportional to the total quantity of Ca2+ released into the cytosol, and its rate of rise is proportional to the Ca2+ current flowing through the RyRs during a spark (ICa(spark)). Thus, Ca2+ currents through RyRs can be monitored inside the cell under physiological conditions. Since the magnitude of ICa(spark) in different sparks varies more than fivefold, Ca2+ sparks appear to be caused by the concerted opening of a number of RyRs. Sparks with the same underlying Ca2+ current cause STOCs, whose amplitudes vary more than threefold, a finding that is best explained by variability in coupling ratio (i.e., the ratio of RyRs to BK channels in the spark microdomain). The time course of STOC decay is approximated by a single exponential that is independent of the magnitude of signal mass and has a time constant close to the value of the mean open time of the BK channels, suggesting that STOC decay reflects BK channel kinetics, rather than the time course of [Ca2+] decline at the membrane. Computer simulations were carried out to determine the spatiotemporal distribution of the Ca2+ concentration resulting from the measured range of ICa(spark). At the onset of a spark, the Ca2+ concentration within 200 nm of the release site reaches a plateau or exceeds the [Ca2+]EC50 for the BK channels rapidly in comparison to the rate of rise of STOCs. These findings suggest a model in which the BK channels lie close to the release site and are exposed to a saturating [Ca2+] with the rise and fall of the STOCs determined by BK channel kinetics. The mechanism of signaling between RyRs and BK channels may provide a model for Ca2+ action on a variety of molecular targets within cellular microdomains.
Substrate and Product Dependence of Force and Shortening in Fast and Slow Smooth Muscle
Authors: Löfgren Mia , Malmqvist Ulf , Arner Anders
Published: July 2002

To explore the molecular mechanisms responsible for the variation in smooth muscle contractile kinetics, the influence of MgATP, MgADP, and inorganic phosphate (Pi) on force and shortening velocity in thiophosphorylated “fast” (taenia coli: maximal shortening velocity Vmax = 0.11 ML/s) and “slow” (aorta: Vmax = 0.015 ML/s) smooth muscle from the guinea pig were compared. Pi inhibited active force with minor effects on the Vmax. In the taenia coli, 20 mM Pi inhibited force by 25%. In the aorta, the effect was markedly less (<10%), suggesting differences between fast and slow smooth muscles in the binding of Pi or in the relative population of Pi binding states during cycling. Lowering of MgATP reduced force and Vmax. The aorta was less sensitive to reduction in MgATP (Km for Vmax: 80 μM) than the taenia coli (Km for Vmax: 350 μM). Thus, velocity is controlled by steps preceding the ATP binding and cross-bridge dissociation, and a weaker binding of ATP is not responsible for the lower Vmax in the slow muscle. MgADP inhibited force and Vmax. Saturating concentrations of ADP did not completely inhibit maximal shortening velocity. The effect of ADP on Vmax was observed at lower concentrations in the aorta compared with the taenia coli, suggesting that the ADP binding to phosphorylated and cycling cross-bridges is stronger in slow compared with fast smooth muscle.
Molecular Basis of Ca2+ Activation of the Mouse Cardiac Ca2+ Release Channel (Ryanodine Receptor)
Authors: Li Pin , Chen S.R. Wayne
Published: July 2002

Activation of the cardiac ryanodine receptor (RyR2) by Ca2+ is an essential step in excitation-contraction coupling in heart muscle. However, little is known about the molecular basis of activation of RyR2 by Ca2+. In this study, we investigated the role in Ca2+ sensing of the conserved glutamate 3987 located in the predicted transmembrane segment M2 of the mouse RyR2. Single point mutation of this conserved glutamate to alanine (E3987A) reduced markedly the sensitivity of the channel to activation by Ca2+, as measured by using single-channel recordings in planar lipid bilayers and by [3H]ryanodine binding assay. However, this mutation did not alter the affinity of [3H]ryanodine binding and the single-channel conductance. In addition, the E3987A mutant channel was activated by caffeine and ATP, was inhibited by Mg2+, and was modified by ryanodine in a fashion similar to that of the wild-type channel. Coexpression of the wild-type and mutant E3987A RyR2 proteins in HEK293 cells produced individual single channels with intermediate sensitivities to activating Ca2+. These results are consistent with the view that glutamate 3987 is a major determinant of Ca2+ sensitivity to activation of the mouse RyR2 channel, and that Ca2+ sensing by RyR2 involves the cooperative action between ryanodine receptor monomers. The results of this study also provide initial insights into the structural and functional properties of the mouse RyR2, which should be useful for studying RyR2 function and regulation in genetically modified mouse models.
Regulation of Cloned Atp-Sensitive K Channels by Adenine Nucleotides and Sulfonylureas
Authors: John Scott A. , Weiss James N. , Ribalet Bernard
Published: July 2002

KATP channels, comprised of the pore-forming protein Kir6.x and the sulfonylurea receptor SURx, are regulated in an interdependent manner by adenine nucleotides, PIP2, and sulfonylureas. To gain insight into these interactions, we investigated the effects of mutating positively charged residues in Kir6.2, previously implicated in the response to PIP2, on channel regulation by adenine nucleotides and the sulfonylurea glyburide. Our data show that the Kir6.2 “PIP2-insensitive” mutants R176C and R177C are not reactivated by MgADP after ATP-induced inhibition and are also insensitive to glyburide. These results suggest that R176 and R177 are required for functional coupling to SUR1, which confers MgADP and sulfonylurea sensitivity to the KATP channel. In contrast, the R301C and R314C mutants, which are also “PIP2-insensitive,” remained sensitive to stimulation by MgADP in the absence of ATP and were inhibited by glyburide. Based on these findings, as well as previous data, we propose a model of the KATP channel whereby in the presence of ATP, the R176 and R177 residues on Kir6.2 form a specific site that interacts with NBF1 bound to ATP on SUR1, promoting channel opening by counteracting the inhibition by ATP. This interaction is facilitated by binding of MgADP to NBF2 and blocked by binding of sulfonylureas to SUR1. In the absence of ATP, since KATP channels are not blocked by ATP, they do not require the counteracting effect of NBF1 interacting with R176 and R177 to open. Nevertheless, channels in this state remain activated by MgADP. This effect may be explained by a direct stimulatory interaction of NBF2/MgADP moiety with another region of Kir6.2 (perhaps the NH2 terminus), or by NBF2/MgADP still promoting a weak interaction between NBF1 and Kir6.2 in the absence of ATP. The region delimited by R301 and R314 is not involved in the interaction with NBF1 or NBF2, but confers additional PIP2 sensitivity.
In Vivo Airway Surface Liquid Cl− Analysis with Solid-State Electrodes
Authors: Caldwell Ray A. , Grubb Barbara R. , Tarran Robert , Boucher Richard C. , Knowles Michael R. , Barker Pierre M.
Published: July 2002

The pathogenesis of cystic fibrosis (CF) airways disease remains controversial. Hypotheses that link mutations in CFTR and defects in ion transport to CF lung disease predict that alterations in airway surface liquid (ASL) isotonic volume, or ion composition, are critically important. ASL [Cl−] is pivotal in discriminating between these hypotheses, but there is no consensus on this value given the difficulty in measuring [Cl−] in the “thin” ASL (∼30 μm) in vivo. Consequently, a miniaturized solid-state electrode with a shallow depth of immersion was constructed to measure ASL [Cl−] in vivo. In initial experiments, the electrode measured [Cl−] in physiologic salt solutions, small volume (7.6 μl) test solutions, and in in vitro cell culture models, with ≥93% accuracy. Based on discrepancies in reported values and/or absence of data, ASL Cl− measurements were made in the following airway regions and species. First, ASL [Cl−] was measured in normal human nasal cavity and averaged 117.3 ± 11.2 mM (n = 6). Second, ASL [Cl−] measured in large airway (tracheobronchial) regions were as follows: rabbit trachea and bronchus = 114.3 ± 1.8 mM; (n = 6) and 126.9 ± 1.7 mM; (n = 3), respectively; mouse trachea = 112.8 ± 4.2 mM (n = 13); and monkey bronchus = 112.3 ± 10.9 mM (n = 3). Third, Cl− measurements were made in small (1–2 mm) diameter airways of the rabbit (108.3 ± 7.1 mM, n = 5) and monkey (128.5 ± 6.8 mM, n = 3). The measured [Cl−], in excess of 100 mM throughout all airway regions tested in multiple species, is consistent with the isotonic volume hypothesis to describe ASL physiology.
Index of B-Fredholm operators and generalization of a Weyl theorem
Authors: Berkani M.
Published: July 2002

The aim of this paper is to show that if

S
S

and

T
T

are commuting B-Fredholm operators acting on a Banach space

X
X

, then

S
T

ST

is a B-Fredholm operator and

i
n
d
(
S
T
)
=
i
n
d
(
S
)
+
i
n
d
(
T
)

ind(ST)=ind(S)+ind(T)

, where

i
n
d

ind

means the index. Moreover if

T
T

is a B-Fredholm operator and

F
F

is a finite rank operator, then

T
+
F

T+F

is a B-Fredholm operator and

i
n
d
(
T
+
F
)
=
i
n
d
(
T
)
.

ind(T+F)= ind(T).

We also show that if

0
0

is isolated in the spectrum of

T
T

, then

T
T

is a B-Fredholm operator of index

0
0

if and only if

T
T

is Drazin invertible. In the case of a normal bounded linear operator

T
T

acting on a Hilbert space

H
H

, we obtain a generalization of a classical Weyl theorem.
Operators with singular continuous spectrum, V. Sparse potentials
Authors: Simon B. , Stolz G.
Published: July 2002

By presenting simple theorems for the absence of positive eigenvalues for certain one-dimensional Schrödinger operators, we are able to construct explicit potentials which yield purely singular continuous spectrum.
Finitistic dimension and Ziegler spectrum
Authors: Krause Henning
Published: July 2002

Given a two-sided artinian ring

Λ
\Lambda

, it is shown that the Ziegler spectrum of

Λ
\Lambda

forms a test class for certain homological properties of

Λ
\Lambda

. We discuss the finitistic dimension of

Λ
\Lambda

, Nunke’s condition, and also the relation between the big and the little finitistic dimension.
Rotation invariant ambiguity functions
Authors: Jiang Qingtang
Published: July 2002

Let

W
(
ψ
;
x
,
y
)

W(\psi ; x, y)

be the wideband ambiguity function. It is obtained in this note that

y

−

α
+
2

2

W
(
ψ
;
x
,
y
)
(
α
>
−
1
)

y^{-\frac {\alpha +2}2}W(\psi ; x, y) (\alpha >-1)

is

S
O
(
2
)

SO(2)

-invariant if and only if the Fourier transform of

ψ
\psi

is a Laguerre function.
Topological horseshoes
Authors: Kennedy Judy , Yorke James
Published: July 2002

When does a continuous map have chaotic dynamics in a set

Q
Q

? More specifically, when does it factor over a shift on

M
M

symbols? This paper is an attempt to clarify some of the issues when there is no hyperbolicity assumed. We find that the key is to define a “crossing number” for that set

Q
Q

. If that number is

M
M

and

M
>
1

M>1

, then

Q
Q

contains a compact invariant set which factors over a shift on

M
M

symbols.
Nonlinear Cauchy-Riemann operators in ℝⁿ
Authors: Iwaniec Tadeusz
Published: July 2002

This paper has arisen from an effort to provide a comprehensive and unifying development of theLpL^{p}-theory of quasiconformal mappings inRn\mathbb {R}^{n}. The governing equations for these mappings form nonlinear differential systems of the first order, analogous in many respects to the Cauchy-Riemann equations in the complex plane. This approach demands that one must work out certain variational integrals involving the Jacobian determinant. Guided by such integrals, we introduce two nonlinear differential operators, denoted byD−\mathcal {D}^{-}andD+\mathcal {D}^{+}, which act on weakly differentiable deformationsf:Ω→Rnf:\Omega \to \mathbb {R}^{n}of a domainΩ⊂Rn\Omega \subset \mathbb {R}^{n}.

Solutions to the so-called Cauchy-Riemann equationsD−f=0\mathcal {D}^{-}f=0andD+f=0\mathcal {D}^{+}f=0are simply conformal deformations preserving and reversing orientation, respectively. These operators, though genuinely nonlinear, possess the important feature of being rank-one convex. Among the many desirable properties, we give the fundamentalLpL^{p}-estimate<mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" alttext="double-vertical-bar script upper D Superscript plus Baseline f double-vertical-bar Subscript p Baseline less-than-or-equal-to upper A Subscript p Baseline left-parenthesis n right-parenthesis double-vertical-bar script upper D Superscript minus Baseline f double-vertical-bar Subscript p Baseline period">‖D+f‖p≤Ap(n)‖D−f‖p.\begin{equation*}\|\mathcal {D}^{+}f\|_{p} \le A_{p}(n)\|\mathcal {D}^{-}f\|_{p}. \end{equation*}

In quest of the best constantAp(n)A_{p}(n), we are faced with fascinating problems regarding quasiconvexity of some related variational functionals. Applications to quasiconformal mappings are indicated.
The quantum analog of a symmetric pair: a construction in type (𝐶_{𝑛},𝐴₁×𝐶_{𝑛-1})
Authors: Baldoni Welleda , Frajria Pierluigi
Published: July 2002

Let

I

\mathcal {I}

be the ideal in the enveloping algebra of

s
p

(
n
,

C

)

\mathfrak {sp}(n,\mathbb C)

generated by the maximal compact subalgebra of

s
p

(
n
−
1
,
1
)

\mathfrak {sp}(n-1,1)

. In this paper we construct an analog of

I

\mathcal I

in the quantized enveloping algebra

U

\mathcal {U}

corresponding to a type

C

n

C_{n}

diagram at generic

q
q

. We find generators for

I

\mathcal {I}

and explicit bases for

U

/

I

\mathcal {U}/\mathcal {I}

.
Poisson transforms on vector bundles
Authors: Yang An
Published: July 2002

Let

G
G

be a connected real semisimple Lie group with finite center, and

K
K

a maximal compact subgroup of

G
G

. Let

(
τ
,
V
)

(\tau ,V)

be an irreducible unitary representation of

K
K

, and

G

×
K

V

G\times _K\,V

the associated vector bundle. In the algebra of invariant differential operators on

G

×
K

V

G\times _K\,V

the center of the universal enveloping algebra of

Lie
⁡
(
G
)

\operatorname {Lie}(G)

induces a certain commutative subalgebra

Z
τ

Z_\tau

. We are able to determine the characters of

Z
τ

Z_\tau

. Given such a character we define a Poisson transform from certain principal series representations to the corresponding space of joint eigensections. We prove that for most of the characters this map is a bijection, generalizing a famous conjecture by Helgason which corresponds to

τ
\tau

the trivial representation.
Isoperimetric Estimates on Sierpinski Gasket Type Fractals
Authors: Strichartz Robert
Published: July 2002

For a compact Hausdorff space

F
F

that is pathwise connected, we can define the connectivity dimension

β
\beta

to be the infimum of all

b
b

such that all points in

F
F

can be connected by a path of Hausdorff dimension at most

b
b

. We show how to compute the connectivity dimension for a class of self–similar sets in

R

n

\mathbb {R}^{n}

that we call point connected , meaning roughly that

F
F

is generated by an iterated function system acting on a polytope

P
P

such that the images of

P
P

intersect at single vertices. This class includes the polygaskets, which are obtained from a regular

n
n

–gon in the plane by contracting equally to all

n
n

vertices, provided

n
n

is not divisible by 4. (The Sierpinski gasket corresponds to

n
=
3

n = 3

.) We also provide a separate computation for the octogasket (

n
=
8

n = 8

), which is not point connected. We also show, in these examples, that

inf

H

β

(

γ

x
,
y

)

1

/

β

\inf \mathcal {H}_{\beta }(\gamma _{x,y})^{1/\beta }

, where the infimum is taken over all paths

γ

x
,
y

\gamma _{x,y}

connecting

x
x

and

y
y

, and

H

β

\mathcal {H}_{\beta }

denotes Hausdorff measure, is equivalent to the original metric on 

F
F

. Given a compact subset

F
F

of the plane of Hausdorff dimension

α
\alpha

and connectivity dimension

β
\beta

, we can define the isoperimetric profile function

h
(
L
)

h(L)

to be the supremum of

H

α

(
F
∩
D
)

\mathcal {H}_{\alpha }(F \cap D)

, where

D
D

is a region in the plane bounded by a Jordan curve (or union of Jordan curves)

γ
\gamma

entirely contained in

F
F

, with

H

β

(
γ
)
≤
L

\mathcal {H}_{\beta }(\gamma ) \le L

. The analog of the standard isperimetric estimate is

h
(
L
)
≤
c

L

α

/

β

h(L) \le cL^{\alpha /\beta }

. We are particularly interested in finding the best constant

c
c

and identifying the extremal domains where we have equality. We solve this problem for polygaskets with

n
=
3
,
5
,
6
,
8

n = 3,5,6,8

. In addition, for

n
=
5
,
6
,
8

n = 5,6,8

we find an entirely different estimate for

h
(
L
)

h(L)

as

L
→
∞

L \rightarrow \infty

, since the boundary of

F
F

has infinite

H

β

\mathcal {H}_{\beta }

measure. We find that the isoperimetric profile function is discontinuous, and that the extremal domains have relatively simple polygonal boundaries. We discuss briefly the properties of minimal paths for the Sierpinski gasket, and the isodiametric problem in the intrinsic metric.
Linear meromorphic differential equations: A modern point of view
Authors: Varadarajan V.
Published: July 2002

A large part of the modern theory of differential equations in the complex domain is concerned with regular singularities and holonomic systems. However the theory of differential equations with irregular singularities has a long history and has become very active in recent years. Substantial links of this theory to the theory of algebraic groups, commutative algebra, resurgent functions, and Galois differential methods have been discovered. This survey attempts a general introduction to some of these aspects, with emphasis on reduction theory, asymptotic analysis, Stokes phenomena, and certain moduli problems.
Quivers, Floer cohomology, and braid group actions
Authors: Khovanov Mikhail , Seidel Paul
Published: July 2002

We consider the derived categories of modules over a certain familyAmA_m(m≥1m \geq 1) of graded rings, and Floer cohomology of Lagrangian intersections in the symplectic manifolds which are the Milnor fibres of simple singularities of typeAm.A_m.We show that each of these two rather different objects encodes the topology of curves on an(m+1)(m+1)-punctured disc. We prove that the braid groupBm+1B_{m+1}acts faithfully on the derived category ofAmA_m-modules, and that it injects into the symplectic mapping class group of the Milnor fibers. The philosophy behind our results is as follows. Using Floer cohomology, one should be able to associate to the Milnor fibre a triangulated category (its construction has not been carried out in detail yet). This triangulated category should contain a full subcategory which is equivalent, up to a slight difference in the grading, to the derived category ofAmA_m-modules. The full embedding would connect the two occurrences of the braid group, thus explaining the similarity between them.
The Dolbeault complex in infinite dimensions II
Authors: Lempert László
Published: July 2002

We study the equation

∂
¯

u
=
f

\overline {\partial }u=f

on a ball

B
(
R
)
⊂

l

1

B(R)\subset l^{1}

, and prove that it is solvable if

f
f

is a Lipschitz continuous, closed

(
0
,
1
)

(0,1)

-form.
Harish-Chandra modules for quantum symmetric pairs
Authors: Letzter Gail
Published: July 2002

Let

U
U

denote the quantized enveloping algebra associated to a semisimple Lie algebra. This paper studies Harish-Chandra modules for the recently constructed quantum symmetric pairs

U
U

,

B
B

in the maximally split case. Finite-dimensional

U
U

-modules are shown to be Harish-Chandra as well as the

B
B

-unitary socle of an arbitrary module. A classification of finite-dimensional spherical modules analogous to the classical case is obtained. A one-to-one correspondence between a large class of natural finite-dimensional simple

B
B

-modules and their classical counterparts is established up to the action of almost

B
B

-invariant elements.
Assessing protein coding region integrity in cDNA sequencing projects.
Authors: Salamov A A , Nishikawa T , Swindells M B
Published: July 2002

MOTIVATION: In cDNA sequencing projects, it is vital to know whether the protein coding region of a sequence is complete, or whether errors have occurred during library construction. Here we present a linear discriminant approach that predicts this completeness by estimating the probability of each ATG being the initiation codon. RESULTS: Because of the current shortage of full-length cDNA data on which to base this work, tests were performed on a non-redundant set of 660 initiation codon-containing DNA sequences that had been conceptually spliced into mRNA/cDNA. We also used an edited set of the same sequences that only contained the region following the initiation codon as a negative control. Using the criterion that only a single prediction is allowed for each sequence, a cut-off was selected at which discrimination of both positive and negative sets was equal. At this cut-off, 67% of each set could be correctly distinguished, with the correct ATG codon also being identified in the positive set. Reliability could be increased further by raising the cut-off or including homologues, the relative merits of which are discussed. AVAILABILITY: The prediction program, called ATGpr, and other data are available at http://www.hri.co.jp/atgpr CONTACT: swintech@hri.co.jp
A bioinformatics tool to select sequences for microarray
studies of mouse models of oncogenesis
Authors: Edgerton Mary E. , Taylor Ronald , Powell John I. , Hunter Lawrence , Simon Richard , Liu Edison T.
Published: July 2002

One of the challenges to the effective utilization of cDNA microarray analysis in mouse models of oncogenesis is the choice of a critical set of probes that are informative for human disease. Given the thousands of genes with a potential role in human oncogenesis and the hundreds of thousands of mouse sequences available for use as probes, selection of an informative set of mouse probes can be an overwhelming task. We have developed a web based sequence mining tool using DataBase Independent (DBI) Perl to annotate publicly available sequences. The Mouse Oncochip Design Tool uses the Mouse Genome Database (MGD) developed and maintained by the Jackson Laboratories for mouse DNA sequences. There are over 380 000 sequences in their database. The output list has been ordered to present the genes more likely to be informative in a mouse model of human cancer using a candidate set of oncogenes to order the list. Mouse sequences that represent genes that are homologous with a member of a human oncogene set are listed first. In addition it provides a set of links for information on clone source gene function.

Contact: http://nciarray.nci.nih.gov/cgi-bin/me/mouse_design.cgi

* To whom correspondence should be addressed.

5 Current address: Department of Pathology and Biomedical Informatics, Vanderbilt University Medical Center, MCN C-3321, Nashville TN 37232, USA.

6 Current address: Department of Pharmacology, School of Medicine, University of Colorado Health Sciences Center, 4200 E Ninth Avenue, Denver CO 80262, USA.

7 Current address: Genome Institute of Singapore, 1 Research Link, IMA Building #04-01, National University of Singapore, Singapore 117604.
Length recognition at the N‐terminal tail for the initiation of FtsH‐mediated proteolysis
Authors: Chiba Shinobu , Akiyama Yoshinori , Mori Hiroyuki , Matsuo Ei‐ichi , Ito Koreaki
Published: July 2002

FtsH‐mediated proteolysis against membrane proteins is processive, and presumably involves dislocation of the substrate into the cytosol where the enzymatic domains of FtsH reside. To study how such a mode of proteolysis is initiated, we manipulated N‐terminal cytosolic tails of three membrane proteins. YccA, a natural substrate of FtsH was found to require the N‐terminal tail of 20 amino acid residues or longer to be degraded by FtsH in vivo. Three unrelated sequences of this segment conferred the FtsH sensitivity to YccA. An artificially constructed TM9‐PhoA protein, derived from SecY, as well as the SecE protein, were sensitized to FtsH by addition of extra amino acid sequences to their N‐terminal cytosolic tails. Thus, FtsH recognizes a cytosolic region of sufficient length (∼20 amino acids) to initiate the processive proteolysis against membrane proteins. Such a region is typically at the N‐terminus and can be diverse in amino acid sequences.