bluuebunny/crossref_metadata_2025 huggingface dataset:
Myocyte proliferation in end-stage cardiac failure in humans
Authors: Kajstura Jan , Leri Annarosa , Finato Nicoletta , Di Loreto Carla , Beltrami Carlo A. , Anversa Piero
Published: July 2002

Introduced several decades ago, the dogma persists that cardiac myocytes are terminally differentiated cells and that division of muscle cells is impossible in the adult heart. More recently, nuclear mitotic divisions in myocytes occasionally were seen, but those observations were challenged on the assumption that the rate of cell proliferation was inconsequential for actual tissue regeneration. Moreover, mitoses were never detected in normal myocardium. However, the analysis of routine histologic preparations constituted the basis for the belief that myocytes were unable to reenter the cell cycle and divide, ignoring the limitations of these techniques. We report here by confocal microscopy that 14 myocytes per million were in mitosis in control human hearts. A nearly 10-fold increase in this parameter was measured in end-stage ischemic heart disease (152 myocytes per million) and in idiopathic dilated cardiomyopathy (131 myocytes per million). Because the left ventricle contains 5.8 × 10
9
myocytes, these mitotic indices imply that 81.2 × 10
3
, 882 × 10
3
, and 760 × 10
3
myocytes were in mitosis in the entire ventricular myocardium of control hearts and hearts affected by ischemic and idiopathic dilated cardiomyopathy, respectively. Additionally, mitosis lasts less than 1 hr, suggesting that large numbers of myocytes can be formed in the nonpathologic and pathologic heart with time. Evidence of cytokinesis in myocytes was obtained, providing unequivocal proof of myocyte proliferation.
The early phase of engraftment after murine blood cell transplantation is mediated by hematopoietic stem cells
Authors: Zijlmans J. Mark J. M. , Visser Jan W. M. , Laterveer Laurens , Kleiverda Karin , Heemskerk Diana P. M. , Kluin Philip M. , Willemze Roel , Fibbe Willem E.
Published: July 2002

Blood cell transplantation is largely replacing bone marrow transplantation because engraftment is more rapid. This accelerated engraftment is thought to be mediated by relatively mature committed hematopoietic progenitor cells. Herein, we have used a modified rhodamine (Rho) staining procedure to identify and purify Rho+/++(dull/bright) and Rho−(negative) subpopulations of hematopoietic progenitor cells in murine cytokine-mobilized blood. The Rho+/++cell population contained >99% of committed progenitor cells within vitrocolony-forming ability. The Rho−cell population contained the majority of hematopoietic stem cells within vivomarrow repopulating ability. The rate of hematopoietic reconstitution was identical in recipients of grafts containing only purified Rho−stem cells or purified Rho−stem cells in combination with large numbers of Rho+/++committed progenitor cells. In contrast, transplantation of 3-fold more hematopoietic stem cells resulted in accelerated reconstitution, indicating that the reconstitution rate was determined by the absolute numbers of Rho−stem cells in the graft. In addition, we observed a 5- to 8-fold reduced frequency of the subset of hematopoietic stem cells with long-term repopulating ability in cytokine-mobilized blood in comparison to steady-state bone marrow. Our results indicate that hematopoietic stem cells and not committed progenitor cells mediate early hematopoietic reconstitution after blood cell transplantation and that relative to bone marrow, the frequency of stem cells with long-term repopulating ability is reduced in mobilized blood.
Characterization of hematopoietic progenitor cells that express the transcription factor SCL, using a
lacZ
“knock-in” strategy
Authors: Elefanty Andrew G. , Begley C. Glenn , Metcalf Donald , Barnett Louise , Köntgen Frank , Robb Lorraine
Published: July 2002

Gene targeting experiments have demonstrated that the transcription factor SCL is essential for primitive and definitive hematopoiesis in the mouse. To study the functional properties of hematopoietic cells expressing
SCL
, we have generated mutant mice (
SCL
lacZ/w
) in which the
Escherichia coli lacZ
reporter gene has been “knocked in” to the
SCL
locus, thereby linking β-galactosidase expression to transcription from the
SCL
promoter. Bone marrow cells from heterozygous
SCL
lacZ/w
mice were sorted into fractions expressing high, intermediate and low levels of β-galactosidase (designated lacZ
high
, lacZ
int
, and lacZ
neg
). Cells that were lacZ
high
or lacZ
int
were enriched for day 12 spleen colony-forming units and myeloid and erythroid colony-forming cells (CFCs). These fractions included >99% of the erythroid and >90% of the myeloid CFCs. Culture of sorted bone marrow populations on stromal cells secreting interleukin-7 or in fetal thymic organ cultures showed that B and T lymphoid progenitors were also present in the lacZ
high
and lacZ
int
fractions. These data provide a functional correlation between
SCL
expression and colony-forming ability in immature hematopoietic cells. Our data also suggested that expression of
SCL
was transient and confined to hematopoietic stem and/or progenitor cells, because the differentiated progeny of most lineages (except the erythroid) were β-galactosidase-negative.
Transgenic DNA integrated into the oat genome is frequently interspersed by host DNA
Authors: Pawlowski Wojciech P. , Somers David A.
Published: July 2002

Integration of transgenic DNA into the plant genome was investigated in 13 transgenic oat (
Avena sativa
L.) lines produced using microprojectile bombardment with one or two cotransformed plasmids. In all transformation events, the transgenic DNA integrated into the plant genome consisted of intact transgene copies that were accompanied by multiple, rearranged, and/or truncated transgene fragments. All fragments of transgenic DNA cosegregated, indicating that they were integrated at single gene loci. Analysis of the structure of the transgenic loci indicated that the transgenic DNA was interspersed by the host genomic DNA. The number of insertions of transgenic DNA within the transgene loci varied from 2 to 12 among the 13 lines. Restriction endonucleases that do not cleave the introduced plasmids produced restriction fragments ranging from 3.6 to about 60 kb in length hybridizing to a probe comprising the introduced plasmids. Although the size of the interspersing host DNA within the transgene locus is unknown, the sizes of the transgene-hybridizing restriction fragments indicated that the entire transgene locus must be at least from 35–280 kb. The observation that all transgenic lines analyzed exhibited genomic interspersion of multiple clustered transgenes suggests a predominating integration mechanism. We propose that transgene integration at multiple clustered DNA replication forks could account for the observed interspersion of transgenic DNA with host genomic DNA within transgenic loci.
Isolation and characterization of mammalian homologs of the
Drosophila
gene
glial cells missing
Authors: Kim Jaesang , Jones Bradley W. , Zock Christiane , Chen Zhoufeng , Wang Hai , Goodman Corey S. , Anderson David J.
Published: July 2002

The
glial cells missing
(
gcm
) gene in
Drosophila
encodes a transcription factor that determines the choice between glial and neuronal fates. We report here the isolation of two mammalian
gcm
homologs,
Gcm1
and
Gcm2
, and the characterization of their expression patterns during embryonic development. Although
Gcm2
is expressed in neural tissues at a low level, the major sites of expression for both of the mammalian genes are nonneural, suggesting that the functions of the mammalian homologs have diverged and diversified. However, when expressed ectopically,
Gcm1
can substitute functionally for
Drosophila gcm
by transforming presumptive neurons into glia. Thus, certain biochemical properties, although not the specificity of the tissue in which the gene is expressed, have been conserved through the evolution of the
Gcm
gene family.
An inhibitor of HIV-1 protease modulates proteasome activity, antigen presentation, and T cell responses
Authors: André Patrice , Groettrup Marcus , Klenerman Paul , de Giuli Rita , Booth Bruce L. , Cerundolo Vincenzo , Bonneville Marc , Jotereau Francine , Zinkernagel Rolf M. , Lotteau Vincent
Published: July 2002

Inhibitors of the protease of HIV-1 have been used successfully for the treatment of HIV-1-infected patients and AIDS disease. We tested whether these protease inhibitory drugs exerted effects in addition to their antiviral activity. Here, we show in mice infected with lymphocytic choriomeningitis virus and treated with the HIV-1 protease inhibitor ritonavir a marked inhibition of antiviral cytotoxic T lymphocyte (CTL) activity and impaired major histocompatibility complex class I-restricted epitope presentation in the absence of direct effects on lymphocytic choriomeningitis virus replication. A potential molecular target was found: ritonavir selectively inhibited the chymotrypsin-like activity of the 20S proteasome. In view of the possible role of T cell-mediated immunopathology in AIDS pathogenesis, the two mechanisms of action (i.e., reduction of HIV replication and impairment of CTL responses) may complement each other beneficially. Thus, the surprising ability of ritonavir to block the presentation of antigen to CTLs may possibly contribute to therapy of HIV infections but potentially also to the therapy of virally induced immunopathology, autoimmune diseases, and transplantation reactions.
Targeted disruption of mouse centromere protein C gene leads to mitotic disarray and early embryo death
Authors: Kalitsis Paul , Fowler Kerry J. , Earle Elizabeth , Hill Joanne , Choo K. H. Andy
Published: July 2002

Centromere protein C (CENPC) is a key protein that has been localized to the inner kinetochore plate of active mammalian centromeres. Using gene targeting techniques, we have disrupted the mouseCenpcgene and shown that the gene is essential for normal mouse embryonic development. Heterozygous mice carrying one functional copy of the gene are healthy and fertile, whereas homozygous embryos fail to thrive. In these embryos, mitotic arrest and gross morphological degeneration become apparent as early as the morula stage of development. The degenerating embryos demonstrate highly irregular cell and nuclear morphologies, including the presence of a large number of micronuclei. Mitotic chromosomes of these embryos display a scattered and often highly condensed configuration and do not segregate in an ordered fashion. These results describing the phenotype of the mutant mouse embryos indicate that CENPC has a direct role in the mitotic progression from metaphase to anaphase.
Chondrocyte-derived apoptotic bodies and calcification of articular cartilage
Authors: Hashimoto Sanshiro , Ochs Robert L. , Rosen Fred , Quach Jacqueline , McCabe Greg , Solan Joell , Seegmiller J. Edwin , Terkeltaub Robert , Lotz Martin
Published: July 2002

Chondrocytes exposed to nitric oxide (NO) or antibody to Fas undergo cell death by apoptosis. This study examines structural and functional properties of chondrocyte-derived apoptotic bodies. In NO treated cartilage, the dense pericellular matrix that normally surrounds the cells is degraded and apoptotic bodies accumulate within and in the vicinity of the chondrocyte lacunae. Functional analysis shows that apoptotic bodies isolated from NO-treated chondrocytes or cartilage produce pyrophosphate. The levels of pyrophosphate produced by apoptotic bodies are increased by pretreatment of the chondrocytes with transforming growth factor β and decreased by interleukin 1. Apoptotic bodies contain alkaline phosphatase and NTP pyrophosphohydrolase activities and can precipitate calcium. These results suggest that chondrocyte-derived apoptotic bodies express functional properties that may contribute to the pathologic cartilage calcification observed in aging and osteoarthritis.
The voltage sensitive Lc-type Ca
2+
channel is functionally coupled to the exocytotic machinery
Authors: Wiser Ofer , Trus Michael , Hernández Ana , Renström Erik , Barg Sebastian , Rorsman Patrik , Atlas Daphne
Published: July 2002

Although N- and P-type Ca
2+
channels predominant in fast-secreting systems, Lc-type Ca
2+
channels (C-class) can play a similar role in certain secretory cells and synapses. For example, in retinal bipolar cells, Ca
2+
entry through the Lc channels triggers ultrafast exocytosis, and in pancreatic β-cells, evoked secretion is highly sensitive to Ca
2+
. These findings suggest that a rapidly release pool of vesicles colocalizes with the Ca
2+
channels to allow high Ca
2+
concentration and a tight coupling of the Lc channels at the release site. In binding studies, we show that the Lc channel is physically associated with synaptotagmin (p65) and the soluble N-ethylmaleimide-sensitive attachment proteins receptors: syntaxin and synaptosomal-associated protein of 25 kDa. Soluble N-ethylmaleimide-sensitive attachent proteins receptors coexpressed in
Xenopus
oocytes along with the Lc channel modify the kinetic properties of the channel. The modulatory action of syntaxin can be overcome by coexpressing p65, where at a certain ratio of p65/syntaxin, the channel regains its unaltered kinetic parameters. The cytosolic region of the channel, Lc
753–893
, separating repeats II–III of its α1C subunit, interacts with p65 and “pulls” down native p65 from rat brain membranes. Lc
753–893
injected into single insulin-secreting β-cell, inhibits secretion in response to channel opening, but not in response to photolysis of caged Ca
2+
, nor does it affect Ca
2+
current. These results suggest that Lc
753–893
competes with the endogenous channel for the synaptic proteins and disrupts the spatial coupling with the secretory apparatus. The molecular organization of the Lc channel and the secretory machinery into a multiprotein complex (named excitosome) appears to be essential for an effective depolarization evoked exocytosis.
Multistep regulation of DNA replication by Cdk phosphorylation of HsCdc6
Authors: Jiang Wei , Wells Nicholas J. , Hunter Tony
Published: July 2002

We have characterized HsCdc6, a human protein homologous to the budding yeast Cdc6p that is essential for DNA replication. We show that, unlike Cdc6p, the levels of HsCdc6 protein remain constant throughout the cell cycle in human cells. However, phosphorylation of HsCdc6 is regulated during the cell cycle. HsCdc6 is an excellent substrate for Cdk2
in vitro
and is phosphorylated
in vivo
at three sites (Ser-54, Ser-74, and Ser-106) that are phosphorylated by Cdk2
in vitro
, strongly suggesting that HsCdc6 is an
in vivo
Cdk substrate. HsCdc6 is nuclear in G
1
, but translocates to the cytoplasm at the start of S phase via Crm1-dependent export. An HsCdc6A1A2A3 mutant, which mimics unphosphorylated HsCdc6, is exclusively nuclear, and its expression inhibits initiation of DNA replication. An HsCdc6E1E2E3 mutant, which mimics phosphorylated HsCdc6, is exclusively cytoplasmic and is not associated with the chromatin/nuclear matrix fraction. Based on these results, we propose that phosphorylation of HsCdc6 by Cdks regulates DNA replication of at least two steps: first, by promoting initiation of DNA replication and, second, through nuclear exclusion preventing DNA rereplication.
Transglutaminase aggregates huntingtin into nonamyloidogenic polymers, and its enzymatic activity increases in Huntington’s disease brain nuclei
Authors: Karpuj Marcela V. , Garren Hideki , Slunt Hilda , Price Donald L. , Gusella James , Becher Mark W. , Steinman Lawrence
Published: July 2002

The protein huntingtin (htt), aggregated in neuronal nuclear inclusions, is pathognomonic of Huntington’s disease (HD). Constructs, translated
in vitro
from the N terminus of htt, containing either polyQ23 from a normal individual, or polyQ41 or polyQ67 from an HD patient, were all soluble. Transglutaminase (TGase) crosslinked these proteins, and the aggregations did not have the staining properties of amyloid. More TGase-catalyzed aggregates formed when the polyglutamine domain of htt exceeded the pathologic threshold of polyQ36. Furthermore, shorter htt constructs, containing 135 aa or fewer, formed more aggregates than did larger htt constructs. TGase activity in the HD brain was increased compared with the control, with notable increases in cell nuclei. The increased TGase activity was brain specific. In lymphoblastoid cells from HD patients, TGase activity was decreased. TGase-mediated crosslinking of htt may be involved in the formation of the nonamyloidogenic nuclear inclusions found in the HD brain. The staining properties of nuclear inclusions in the HD brain revealed that they were not amyloid.
Src, a molecular switch governing gain control of synaptic transmission mediated by
N-
methyl-
d
-aspartate receptors
Authors: Yu Xian-Min , Salter Michael W.
Published: July 2002

The
N-
methyl-
d
-aspartate (NMDA) receptor
is a principal subtype of glutamate receptor mediating fast excitatory
transmission at synapses in the dorsal horn of the spinal cord and
other regions of the central nervous system. NMDA receptors are crucial
for the lasting enhancement of synaptic transmission that occurs both
physiologically and in pathological conditions such as chronic pain.
Over the past several years, evidence has accumulated indicating that
the activity of NMDA receptors is regulated by the protein tyrosine
kinase, Src. Recently it has been discovered that, by means of
up-regulating NMDA receptor function, activation of Src mediates the
induction of the lasting enhancement of excitatory transmission known
as long-term potentiation in the CA1 region of the hippocampus. Also,
Src has been found to amplify the up-regulation of NMDA receptor
function that is produced by raising the intracellular concentration of
sodium. Sodium concentration increases in neuronal dendrites during
high levels of firing activity, which is precisely when Src becomes
activated. Therefore, we propose that the boost in NMDA receptor
function produced by the coincidence of activating Src and raising
intracellular sodium may be important in physiological and
pathophysiological enhancement of excitatory transmission in the dorsal
horn of the spinal cord and elsewhere in the central nervous system.
HIV-1 Nef increases T cell activation in a stimulus-dependent manner
Authors: Schrager Jeffrey A. , Marsh Jon W.
Published: July 2002

Lentiviral Nef increases viral replicationin vivo, plays a direct role in pathogenesis, and increases viral particle infectivity. We now find that HIV Nef also increases the activation of T cells, a cellular state required for optimal viral replication. This enhancement is stimulant-dependent. As defined by IL-2 generation, activation of T cells stimulated with classical mitogens [phorbol 12-myristate 13-acetate (PMA) + anti-CD3, PMA + phytohemagglutinin, and PMA + ionomycin] is unaffected by the expression of Nef. However, Nef increases IL-2 secretion when cells are stimulated through the T cell receptor and the costimulus receptor (CD28). This increase in activation, which depends on Nef myristylation, is caused by an increase in the number of cells reaching full activation and not by an increase in the amount of IL-2 secreted per cell. These findings demonstrate that Nef lowers the threshold of the dual-receptor T cell activation pathway. The capacity of Nef to increase T cell activity may be very importantin vivowhen Nef is the predominant or the only viral gene product expressed.
A DNA pentaplex incorporating nucleobase quintets
Authors: Chaput John C. , Switzer Christopher
Published: July 2002

Supramolecular self-assembly is an integral step in the formation of many biological structures. Here we report a DNA pentaplex that derives from a metal-assisted, hydrogen bond-mediated self-assembly process. In particular, cesium ions are found to induce pentameric assembly of DNA bearing the nonstandard nucleobase iso-guanine. The pentaplex was designed by using a simple algorithm to predict nucleobase structural requirements within a quintet motif. The design principles are general and should extend to complexes beyond pentaplex. Structures exhibiting molecularities of five or more were previously accessible to peptides, but not nucleic acids.
Crystal structure of a multifunctional 2-Cys peroxiredoxin heme-binding protein 23 kDa/proliferation-associated gene product
Authors: Hirotsu Shoko , Abe Yasuko , Okada Kengo , Nagahara Noriyuki , Hori Hiroyuki , Nishino Takeshi , Hakoshima Toshio
Published: July 2002

Heme-binding protein 23 kDa (HBP23), a rat isoform of human proliferation-associated gene product (PAG), is a member of the peroxiredoxin family of peroxidases, having two conserved cysteine residues. Recent biochemical studies have shown that HBP23/PAG is an oxidative stress-induced and proliferation-coupled multifunctional protein that exhibits specific bindings to c-Abl protein tyrosine kinase and heme, as well as a peroxidase activity. A 2.6-Å resolution crystal structure of rat HBP23 in oxidized form revealed an unusual dimer structure in which the active residue Cys-52 forms a disulfide bond with conserved Cys-173 from another subunit by C-terminal tail swapping. The active site is largely hydrophobic with partially exposed Cys-173, suggesting a reduction mechanism of oxidized HBP23 by thioredoxin. Thus, the unusual cysteine disulfide bond is involved in peroxidation catalysis by using thioredoxin as the source of reducing equivalents. The structure also provides a clue to possible interaction surfaces for c-Abl and heme. Several significant structural differences have been found from a 1-Cys peroxiredoxin, ORF6, which lacks the C-terminal conserved cysteine corresponding to Cys-173 of HBP23.
Plasmodium falciparum
domain mediating adhesion to chondroitin sulfate A: A receptor for human placental infection
Authors: Buffet Pierre A. , Gamain Benoit , Scheidig Christine , Baruch Dror , Smith Joseph D. , Hernandez-Rivas Rosaura , Pouvelle Bruno , Oishi Shinya , Fujii Nobutaka , Fusai Thierry , Parzy Daniel , Miller Louis H. , Gysin Jürg , Scherf Artur
Published: July 2002

Malaria during the first pregnancy causes a high rate of fetal and neonatal death. The decreasing susceptibility during subsequent pregnancies correlates with acquisition of antibodies that block binding of infected red cells to chondroitin sulfate A (CSA), a receptor for parasites in the placenta. Here we identify a domain within a particular
Plasmodium falciparum
erythrocyte membrane protein 1 that binds CSA. We cloned a
var
gene expressed in CSA-binding parasitized red blood cells (PRBCs). The gene had eight receptor-like domains, each of which was expressed on the surface of Chinese hamster ovary cells and was tested for CSA binding. CSA linked to biotin used as a probe demonstrated that two Duffy-binding-like (DBL) domains (DBL3 and DBL7) bound CSA. DBL7, but not DBL3, also bound chondroitin sulfate C (CSC) linked to biotin, a negatively charged sugar that does not support PRBC adhesion. Furthermore, CSA, but not CSC, blocked the interaction with DBL3; both CSA and CSC blocked binding to DBL7. Thus, only the DBL3 domain displays the same binding specificity as PRBCs. Because protective antibodies present after pregnancy block binding to CSA of parasites from different parts of the world, DBL-3, although variant, may induce cross-reactive immunity that will protect pregnant women and their fetuses.
Two-dimensional freezing in the liquid–vapor interface of a dilute Pb:Ga alloy
Authors: Yang Bin , Gidalevitz David , Li Dongxu , Huang Zhengqing , Rice Stuart A.
Published: July 2002

We report the results of x-ray reflectivity and grazing incidence x-ray diffraction studies of the liquid–vapor interface of a dilute alloy of Pb in Ga over the temperature range of 23–76°C. Our data show that the liquid–vapor interface of this alloy is stratified for several atomic diameters into the bulk liquid and that a monolayer of Pb forms the outermost stratum of the interface. Over the temperature range of 23–56°C, the monolayer of Pb is in an ordered hexagonal phase. At about 58°C, this monolayer undergoes a first-order transition to a hexatic phase, which remains stable to 76°C. An analogy between the observed transition and the first-order melting transition in a one-component classical plasma is suggested.
Inhibition of luciferase expression in transgenic
Aedes aegypti
mosquitoes by Sindbis virus expression of antisense luciferase RNA
Authors: Johnson Barbara W. , Olson Ken E. , Allen-Miura Tanya , Rayms-Keller Alfredo , Carlson Jonathan O. , Coates Craig J. , Jasinskiene Nijole , James Anthony A. , Beaty Barry J. , Higgs Stephen
Published: July 2002

A rapid and reproducible method of inhibiting the expression of specific genes in mosquitoes should further our understanding of gene function and may lead to the identification of mosquito genes that determine vector competence or are involved in pathogen transmission. We hypothesized that the virus expression system based on the mosquito-borne
Alphavirus
, Sindbis (Togaviridae), may efficiently transcribe effector RNAs that inhibit expression of a targeted mosquito gene. To test this hypothesis, germ-line-transformed
Aedes aegypti
that express luciferase (LUC) from the mosquito
Apyrase
promoter were intrathoracically inoculated with a double subgenomic Sindbis (dsSIN) virus TE/3′2J/anti-luc (Anti-luc) that transcribes RNA complementary to the 5′ end of the LUC mRNA. LUC activity was monitored in mosquitoes infected with either Anti-luc or control dsSIN viruses expressing unrelated antisense RNAs. Mosquitoes infected with Anti-luc virus exhibited 90% reduction in LUC compared with uninfected and control dsSIN-infected mosquitoes at 5 and 9 days postinoculation. We demonstrate that a gene expressed from the mosquito genome can be inhibited by using an antisense strategy. The dsSIN antisense RNA expression system is an important tool for studying gene function
in vivo
.
Molecular and functional remodeling of electrogenic membrane of hypothalamic neurons in response to changes in their input
Authors: Tanaka M. , Cummins T. R. , Ishikawa K. , Black J. A. , Ibata Y. , Waxman S. G.
Published: July 2002

Neurons respond to stimuli by integrating generator and synaptic potentials and generating action potentials. However, whether the underlying electrogenic machinery within neurons itself changes, in response to alterations in input, is not known. To determine whether there are changes in Na
+
channel expression and function within neurons in response to altered input, we exposed magnocellular neurosecretory cells (MNCs) in the rat supraoptic nucleus to different osmotic milieus by salt-loading and studied Na
+
channel mRNA and protein, and Na
+
currents, in these cells.
In situ
hybridization demonstrated significantly increased mRNA levels for α-II, Na6, β1 and β2 Na
+
channel subunits, and immunohistochemistry/immunoblotting showed increased Na
+
channel protein after salt-loading. Using patch-clamp recordings to examine the deployment of functional Na
+
channels in the membranes of MNCs, we observed an increase in the amplitude of the transient Na
+
current after salt-loading and an even greater increase in amplitude and density of the persistent Na
+
current evoked at subthreshold potentials by slow ramp depolarizations. These results demonstrate that MNCs respond to salt-loading by selectively synthesizing additional, functional Na
+
channel subtypes whose deployment in the membrane changes its electrogenic properties. Thus, neurons may respond to changes in their input not only by producing different patterns of electrical activity, but also by remodeling the electrogenic machinery that underlies this activity.
Modulation of glycine receptors in retinal ganglion cells by zinc
Authors: Han Yi , Wu Samuel M.
Published: July 2002

Effects of zinc, an endogenous neuromodulator in the central nervous system, on glycine receptors (GlyRs) in retinal ganglion cells were investigated by using the whole-cell voltage-clamp technique. Zn
2+
at low concentration (<2 μM) potentiated the glycine-induced chloride current and at higher concentration (>10 μM) suppressed it. This biphasic regulatory action of zinc acted selectively on the fast component of the glycine-induced current mediated by the strychnine-sensitive GlyRs, but not on the slow component mediated by the 5,7-dichlorokynurenic acid-sensitive GlyRs. Dose-response studies showed that 1 μM Zn
2+
increased the maximum glycine response (
I
∝
) and shifted the EC
50
to the left, suggesting that Zn
2+
at low concentrations acts as an allosteric activator of the strychnine-sensitive GlyRs. Zn
2+
at a concentration of 100 μM did not alter
I
∝
and shifted the EC
50
to the right, indicating that Zn
2+
at high concentrations acts as a competitive inhibitor of the GlyRs. Physiological functions of zinc modulation of GlyRs in retinal ganglion cells are discussed.
Specific interaction of the yeast
cis
-Golgi syntaxin Sed5p and the coat protein complex II component Sec24p of endoplasmic reticulum-derived transport vesicles
Authors: Peng Renwang , Grabowski Reiner , De Antoni Anna , Gallwitz Dieter
Published: July 2002

The generation of transport vesicles at the endoplasmic reticulum (ER) depends on cytosolic proteins, which, in the form of subcomplexes (Sec23p/Sec24p; Sec13p/Sec31p) are recruited to the ER membrane by GTP-bound Sar1p and form the coat protein complex II (COPII). Using affinity chromatography and two-hybrid analyses, we found that the essential COPII component Sec24p, but not Sec23p, binds to the
cis
-Golgi syntaxin Sed5p. Sec24p/Sed5p interaction
in vitro
was not dependent on the presence of [Sar1p⋅GTP]. The binding of Sec24p to Sed5p is specific; none of the other seven yeast syntaxins bound to this COPII component. Whereas the interaction site of Sec23p is within the N-terminal half of the 926-aa-long Sec24p (amino acid residues 56–549), Sed5p binds to the N- and C-terminal halves of the protein. Destruction by mutagenesis of a potential zinc finger within the N-terminal half of Sec24p led to a nonfunctional protein that was still able to bind Sec23p and Sed5p. Sec24p/Sed5p binding might be relevant for cargo selection during transport-vesicle formation and/or for vesicle targeting to the
cis
-Golgi.
Demonstration of the parity-violating energy difference between enantiomers
Authors: Szabó-Nagy Andrea , Keszthelyi Lajos
Published: July 2002

Racemic mixtures of (+) and (−) sodium potassium tartrate, tris(1,2-ethanediamine)cobalt(III), and tris(1,2-ethanediamine)iridium(III) molecules were crystallized, and the optical activities of the resulting crystalline materials, dissolved in water, were carefully measured to study the influence of the parity-violating energy difference in the crystallization process. Although no effect was found in the case of tartrate, enantiomeric excess appeared in the crystallization of the cobalt and iridium complexes. These investigations, performed in our laboratory, demonstrated the contribution of the parity-violating neutral weak current to the forces acting in molecules.
A molecular trigger of lipid binding-induced opening of a helix bundle exchangeable apolipoprotein
Authors: Narayanaswami Vasanthy , Wang Jianjun , Schieve Dean , Kay Cyril M. , Ryan Robert O.
Published: July 2002

Apolipophorin III (apoLp-III) from the sphinx moth,
Manduca sexta,
is a helix bundle protein that interacts reversibly with lipoproteins. Its five elongated amphipathic α-helices are organized in an antiparallel fashion, with helices 3 and 4 connected by a short 6-residue (PDVEKE) linker helix, termed helix 3′. Upon interaction with lipoproteins, apoLp-III opens to expose a continuous hydrophobic interior. It was postulated that helix bundle opening is preceded by an initiation step wherein helix 3′ serves to recognize available lipoprotein surface binding sites. To test this hypothesis, helix 3′ was replaced by residues that have a propensity to form a type I β-turn, NPNG. This mutant apoLp-III was defective in lipoprotein binding assays. To define a more precise mode of interaction, the relevance of the presence of the hydrophobic Val-97 flanked by Asp-96 and Glu-98 was investigated by site-directed mutagenesis. V97N and D96N/V97N/E98Q apoLp-III were unable to compete with wild-type apoLp-III to initiate an interaction with lipoproteins, whereas D96N/E98Q apoLp-III was as competent as wild-type apoLp-III. The results suggest that Val-97 is critical, whereas Asp-96 and Glu-98 are irrelevant for initiating binding to lipoproteins. A model of binding is presented wherein apoLp-III is oriented with the helix 3′ end of the molecule juxtaposed to the lipoprotein surface. Recognition of lipoprotein surface hydrophobic defects by Val-97 triggers opening of the helix bundle and facilitates formation of a stable binding interaction.
High recombination rate in natural populations of
Plasmodium falciparum
Authors: Conway David J. , Roper Cally , Oduola Ayoade M. J. , Arnot David E. , Kremsner Peter G. , Grobusch Martin P. , Curtis Chris F. , Greenwood Brian M.
Published: July 2002

Malaria parasites are sexually reproducing protozoa, although the extent of effective meiotic recombination in natural populations has been debated. If meiotic recombination occurs frequently, compared with point mutation and mitotic rearrangement, linkage disequilibrium between polymorphic sites is expected to decline with increasing distance along a chromosome. The rate of this decline should be proportional to the effective meiotic recombination rate in the population. Multiple polymorphic sites covering a 5-kb region of chromosome 9 (the
msp1
gene) have been typed in 547 isolates from six populations in Africa to test for such a decline and estimate its rate in populations of
Plasmodium falciparum
. The magnitude of two-site linkage disequilibrium declines markedly with increasing molecular map distance between the sites, reaching nonsignificant levels within a map range of 0.3–1.0 kb in five of the populations and over a larger map distance in the population with lowest malaria endemicity. The rate of decline in linkage disequilibrium over molecular map distance is at least as rapid as that observed in most chromosomal regions of other sexually reproducing eukaryotes, such as humans and
Drosophila
. These results are consistent with the effective recombination rate expected in natural populations of
P. falciparum
, predicted on the basis of the underlying molecular rate of meiotic crossover and the coefficient of inbreeding caused by self-fertilization events. This is conclusive evidence to reject any hypothesis of clonality or low rate of meiotic recombination in
P. falciparum
populations. Moreover, the data have major implications for the design and interpretation of population genetic studies of selection on
P. falciparum
genes.
Tracking the intracellular path of poly(ethylenimine)/DNA complexes for gene delivery
Authors: Godbey W. T. , Wu Kenneth K. , Mikos Antonios G.
Published: July 2002

Poly(ethylenimine) (PEI) is one of a number of polycations that has been used successfully to transfer genes into living cells. Although PEI shows promise in the field of gene therapy, to date no rigorous proof of mechanism has been published regarding the fate of PEI/DNA administered for transfection. Here we show, by using fluorescent labeling and confocal microscopy, the paths of PEI/DNA complexes from endocytosis to gene expression. We found that complexes attach to cell surfaces and migrate into clumps that are endocytosed. The endocytotic vesicles grow in number and size and are occasionally seen to lyse. Most interesting is the fact that endocytosed PEI, whether administered with or without DNA, undergoes nuclear localization in the form of ordered structures.
Model selection for quantitative trait locus analysis in polyploids
Authors: Doerge R. W. , Craig Bruce A.
Published: July 2002

Over the years, substantial gains have been made in locating regions of agricultural genomes associated with characteristics, diseases, and agroeconomic traits. These gains have relied heavily on the ability to statistically estimate the association between DNA markers and regions of a genome (quantitative trait loci or QTL) related to a particular trait. The majority of these advances have focused on diploid species, even though many important agricultural crops are, in fact, polyploid. The purpose of our work is to initiate an algorithmic approach for model selection and QTL detection in polyploid species. This approach involves the construction of all possible chromosomal configurations (models) that may result in a gamete, model reduction based on estimation of marker dosage from progeny data, and lastly model selection. While simplified for initial explanation, our approach has demonstrated itself to be extendible to many breeding schemes and less restricted settings.
Role of endocytosis in the activation of the extracellular signal-regulated kinase cascade by sequestering and nonsequestering G protein-coupled receptors
Authors: Pierce Kristen L. , Maudsley Stuart , Daaka Yehia , Luttrell Louis M. , Lefkowitz Robert J.
Published: July 2002

Acting through a number of distinct pathways, many G protein-coupled receptors (GPCRs) activate the extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) cascade. Recently, it has been shown that in some cases, clathrin-mediated endocytosis is required for GPCR activation of the ERK/MAPK cascade, whereas in others it is not. Accordingly, we compared ERK activation mediated by a GPCR that does not undergo agonist-stimulated endocytosis, the α
2A
adrenergic receptor (α
2A
AR), with ERK activation mediated by the β
2
adrenergic receptor (β
2
AR), which is endocytosed. Surprisingly, we found that in COS-7 cells, ERK activation by the α
2A
AR, like that mediated by both the β
2
AR and the epidermal growth factor receptor (EGFR), is sensitive to mechanistically distinct inhibitors of clathrin-mediated endocytosis, including monodansylcadaverine, a mutant dynamin I, and a mutant β-arrestin 1. Moreover, we determined that, as has been shown for many other GPCRs, both α
2A
and β
2
AR-mediated ERK activation involves transactivation of the EGFR. Using confocal immunofluorescence microscopy, we found that stimulation of the β
2
AR, the α
2A
AR, or the EGFR each results in internalization of a green fluorescent protein-tagged EGFR. Although β
2
AR stimulation leads to redistribution of both the β
2
AR and EGFR, activation of the α
2A
AR leads to redistribution of the EGFR but the α
2A
AR remains on the plasma membrane. These findings separate GPCR endocytosis from the requirement for clathrin-mediated endocytosis in EGFR transactivation-mediated ERK activation and suggest that it is the receptor tyrosine kinase or another downstream effector that must engage the endocytic machinery.
Some transformations of color information from lateral geniculate nucleus to striate cortex
Authors: De Valois Russell L. , Cottaris Nicolas P. , Elfar Sylvia D. , Mahon Luke E. , Wilson J. Anthony
Published: July 2002

We have recorded the responses of single cells in the lateral geniculate nucleus (LGN) and striate cortex of the macaque monkey. The response characteristics of neurons at these successive visual processing levels were examined with isoluminant gratings, cone-isolating gratings, and luminance-varying gratings. The main findings were: (
i
) Whereas almost all parvo- and konio-cellular LGN cells are of just two opponent-cell types, either differencing the L and M cones (L
o
and M
o
cells), or the S vs. L + M cones (S
o
cells), relatively few striate cortex simple cells show chromatic responses along these two cardinal LGN axes. Rather, most are shifted away from these LGN chromatic axes as a result of combining the outputs (or the transformed outputs) of S
o
with those of L
o
and/or M
o
cells. (
ii
) LGN cells on average process color information linearly, exhibiting sinusoidal changes in firing rate to isoluminant stimuli that vary sinusoidally in cone contrast as a function of color angle. Some striate cortex simple cells also give linear responses, but most show an expansive response nonlinearity, resulting in narrower chromatic tuning on average at this level. (
iii
) There are many more +S
o
than −S
o
LGN cells, but at the striate cortex level −S
o
input to simple cells is as common as +S
o
input. (
iv
) Overall, the contribution of the S-opponent path is doubled at the level of the striate cortex, relative to that at the LGN.
Function–structure analysis of proteins using covarion-based evolutionary approaches: Elongation factors
Authors: Gaucher Eric A. , Miyamoto Michael M. , Benner Steven A.
Published: July 2002

The divergent evolution of protein sequences from genomic
databases can be analyzed by the use of different mathematical models.
The most common treat all sites in a protein sequence as equally
variable. More sophisticated models acknowledge the fact that
purifying selection generally tolerates variable amounts of amino acid
replacement at different positions in a protein sequence. In their
“stationary” versions, such models assume that the replacement
rate at individual positions remains constant throughout evolutionary
history. “Nonstationary” covarion versions, however, allow the
replacement rate at a position to vary in different branches of the
evolutionary tree. Recently, statistical methods have been developed
that highlight this type of variation in replacement rates. Here, we
show how positions that have variable rates of divergence in different
regions of a tree (“covarion behavior”), coupled with analyses of
experimental three-dimensional structures, can provide experimentally
testable hypotheses that relate individual amino acid residues to
specific functional differences in those branches. We illustrate this
in the elongation factor family of proteins as a paradigm for
applications of this type of analysis in functional genomics generally.
The Well‐being of Salvadoran Refugees
Authors: Young Marta Y. , Evans David R.
Published: July 2002

Recently arrived and settled Salvadoran refugees and Anglo‐Canadians in London, Ontario were compared with respect to psychological distress, quality of life, and life satisfaction. A matched sample of 60 participants in each group completed a demographic questionnaire, the Brief Symptom Inventory, a shortened version of the Quality of Life Questionnaire, and the Satisfaction with Life Scale in either English or Spanish. Significant group differences were obtained on measures of quality of life and life satisfaction, but not of psychological distress. The results are discussed in relation to findings with other refugee groups.
Turks and Tatars in Bulgaria and the Balkans
Authors: Eminov Ali
Published: July 2002

The Koran and the Bible are God's grace Which is what all four holy Books embrace; To scorn and segregate this or that race Would be the darkest stains on one's face.Aşik Veysel

Nationalist movements everywhere aim to create “territorially bounded political units (states) out of homogeneous cultural communities (nations).” Unfortunately, ethnic, linguistic, religious, political, and personal identities rarely coincide with geographical boundaries that enclose nation states. There are always groups within nation states whose identities are different from the majority. The leaders of nation states often see the presence of multiple ethnic communities within a single nation state as a sign of tension and instability, a threat to the integrity and indeed the very survival of a nation state. Consequently, they seek ways to culturally homogenize the nation so that the state and the nation come to coincide with one another.
Research on memory development: Historical trends and current themes
Authors: Schneider Wolfgang
Published: July 2002

This article presents a survey of research on memory development conducted within the last 120 years. It begins with an examination of historical trends and then focuses on developmental trends over the last three decades. The article concludes with some predictions of future research activities and trends in this classic domain of cognitive development.
Rules of Everyday Family Life: The Development of Social Rules in Mother-Child and Sibling Relationships
Authors: Piotrowski Caroline
Published: July 2002

The aims of this study were: (1) to characterise changes in the frequency and kind of social rules used in mother-child and sibling relationships over a 14-month period; (2) to assess the degree to which conflict functioned as a forum for social rules; and (3) to describe the nature of the association between references to social rules and the quality of family relationships. Naturalistic home observations of 47 families were conducted when focal younger siblings were 33 and 47 months of age. Family conversations were coded for a variety of rules. Over time, mothers referred to rules less frequently per hour, whereas elder siblings referred to rules more frequently per hour. Proportionately, mothers employed rules relatively more often than children; however, conventional rules were utilised by mothers more than moral rules. As hypothesised, moral rules were more symmetrical in sibling relationships and more complementary in mother-child dyads. Mothers mentioned social rules more often during conflicts, whereas siblings did not. Finally, moral rules mentioned by both younger and elder siblings were associated with less harmonious sibling relationships. Results were discussed with regard to theories of moral socialisation and the nature of family relationships.
Inhibitory Effect of Ferulic Acid and Isoferulic Acid on the Production of Macrophage Inflammatory Protein‐2 in Response to Respiratory Syncytial Virus Infection in RAW264.7 Cells
Authors: Sakai S. , Kawamata H. , Kogure T. , Mantani N. , Terasawa K. , Umatake M. , Ochiai H.
Published: July 2002

We investigated the effect of ferulic acid (FA) and isoferulic acid (IFA), which are the main active components of the rhizoma of Cimicifuga heracleifolia (CH), an anti‐inflammatory drug used frequently in Japanese traditional medicine, on the production of macrophage inflammatory protein‐2 (MIR‐2) in a murine macrophage cell line, RAW264.7, in response to respiratory syncytial virus (RSV) infection. Following the exposure of cells to RSV for 20 h, the MIP‐2 level in condition medium was increased to about 20 ng/ml, although this level in mock‐infected cells was negligible. In the presence of either FA or IFA, RSV‐infected cells reduced MIP‐2 production in a dose‐dependent manner. These data suggest that FA and IFA might be responsible, at least in part, for the anti‐inflammatory drug effect of CH extract through the inhibition of MIP‐2 production.
Tuberculin skin testing in intravenous drug users: differences between HIV‐seropositive and HIV‐seronegative subjects
Authors: PORTU JOSÉ J. , ALDAMIZ‐ETXEBARRIA MIKEL , AGUD JOSÉ M. , ARÉVALO JOSÉ M. , ALMARAZ MARía J. , AYENSA CÁNDIDO
Published: July 2002

Abstract The prevalence of tuberculin skin test reactions among intravenous drug abusers and differences in tuberculin skin test positivity between HIV‐seropositive and HIV‐seronegative subjects were evaluated in a cross‐sectional study of 1131 subjects. They were recruited from a therapeutic community, from those who attended the centre for the treatment of drug addiction and from those who visited for any reason an acute tertiary‐care hospital in Vitoria‐Gasteiz, Basque Country (Spain). All subjects underwent skin testing with purified protein derivative (PPD) tuberculin and testing for HIV antibodies. CD4 + T‐lymphocyte count was determined in HIV‐seropositive individuals. Positive PPD tests were recorded in 35% of drug users who were HIV‐seropositive and in 65% in those who were HIV‐seronegative. In the HIV‐infected group, there was a significant association between results of the tuberculin test and CD4 + T‐lymphocyte count. When the CD4 + T‐lymphocyte count was ≥ 500 cells/mm 3, percentages of positive PPD tests were similar in HIV‐seropositives and HIV‐seronegatives (47% versus 65%) but when the CD4 + count was < 500 cells/mm 3, positive PPD tests occurred in only 21% of HIV‐seropositives. The PPD test showed a decreased sensitivity for detecting tuberculosis infection in HIV‐infected intravenous drug users with CD4 + T‐lymphocyte counts fewer than 500 cells/mm 3.