bluuebunny/crossref_metadata_2025 huggingface dataset:
Sequestration of cAMP response element-binding proteins by transcription factor decoys causes collateral elaboration of regenerating
Aplysia
motor neuron axons
Authors: Dash Pramod K. , Tian Lian-Ming , Moore Anthony N.
Published: July 2002

Axonal injury increases intracellular Ca
2+
and cAMP and has been shown to induce gene expression, which is thought to be a key event for regeneration. Increases in intracellular Ca
2+
and/or cAMP can alter gene expression via activation of a family of transcription factors that bind to and modulate the expression of CRE (Ca
2+
/cAMP response element) sequence-containing genes. We have used
Aplysia
motor neurons to examine the role of CRE-binding proteins in axonal regeneration after injury. We report that axonal injury increases the binding of proteins to a CRE sequence-containing probe. In addition, Western blot analysis revealed that the level of ApCREB2, a CRE sequence-binding repressor, was enhanced as a result of axonal injury. The sequestration of CRE-binding proteins by microinjection of CRE sequence-containing plasmids enhanced axon collateral formation (both number and length) as compared with control plasmid injections. These findings show that Ca
2+
/cAMP-mediated gene expression via CRE-binding transcription factors participates in the regeneration of motor neuron axons.
Isolation of a protein Z-dependent plasma protease inhibitor
Authors: Han Xin , Fiehler Ryan , Broze George J.
Published: July 2002

Human protein Z (PZ) is a 62,000-
M
r
, vitamin K-dependent plasma protein whose structure is similar to coagulation factors VII, IX, X, protein C, and protein S, but whose function is not known. The procoagulant activity of factor Xa in a one-stage plasma coagulation assay is reduced when factor Xa is first incubated with PZ. This apparent inhibitory effect is time dependent, requires the presence of calcium ions and procoagulant phospholipids (rabbit brain cephalin), and appears predominantly related to the incubation period of PZ with cephalin. In serum the initial rate of inhibition of factor Xa with calcium ions and cephalin also is enhanced in the presence PZ. A PZ-dependent protease inhibitor (ZPI) has been isolated from plasma. ZPI is a 72,000-
M
r
single-chain protein with an N-terminal amino acid sequence of LAPSPQSPEXXA (X = indeterminate) and an estimated concentration in citrate-treated plasma of 1.0–1.6 μg/ml. In systems using purified components, the factor Xa inhibition produced by ZPI is rapid (>95% within 1 min by coagulation assay) and requires the presence of PZ, calcium ions, and cephalin. The inhibitory process appears to involve the formation of a factor Xa–PZ–ZPI complex at the phospholipid surface.
ATM binds to β-adaptin in cytoplasmic vesicles
Authors: Lim Dae-Sik , Kirsch David G. , Canman Christine E. , Ahn Jin-Hyun , Ziv Yael , Newman Lori S. , Darnell Robert B. , Shiloh Yosef , Kastan Michael B.
Published: July 2002

Inherited mutations in the
ATM
gene lead to a complex clinical phenotype characterized by neuronal degeneration, oculocutaneous telangiectasias, immune dysfunction, and cancer predisposition. Using the yeast two-hybrid system, we demonstrate that ataxia telangiectasia mutated (ATM) binds to β-adaptin, one of the components of the AP-2 adaptor complex, which is involved in clathrin-mediated endocytosis of receptors. The interaction between ATM and β-adaptin was confirmed
in vitro,
and coimmunoprecipitation and colocalization studies show that the proteins also associate
in vivo
. ATM also interacts
in vitro
with β-NAP, a neuronal-specific β-adaptin homolog that was identified as an autoantigen in a patient with cerebellar degeneration. Our data describing the association of ATM with β-adaptin in vesicles indicate that ATM may play a role in intracellular vesicle and/or protein transport mechanisms.
CuAand CuZare variants of the electron transfer center in nitrous oxide reductase
Authors: Farrar J. A. , Zumft W. G. , Thomson A. J.
Published: July 2002

Nitrous oxide reductase (N2OR) is a dimeric copper-dependent bacterial enzyme that catalyzes the reduction of N2O to N2as part of the denitrification pathway. In the absence of an x-ray crystal structure, the current model of the nature of the copper sites within the enzyme is based on four copper atoms per monomer and assigns two copper atoms to an electron transfer center, CuA, abis-thiolate-bridged dinuclear copper center found to date only in N2OR and cytochromecoxidase, and two copper atoms to a second dinuclear center, CuZ, presumed to be the site of catalysis. Based on detailed analysis of the low temperature magnetic CD spectra of N2OR, this paper revises the current model and proposes that both CuAand CuZare variants of an electron transfer center and hence that all of the observed optical features are due to this electron transfer center. It is proposed further that the presence of these different forms provides a mechanism for the delivery of two electrons to an active site comprising copper ions lacking thiolate coordination.
The adenomatous polyposis coli-binding protein EB1 is associated with cytoplasmic and spindle microtubules
Authors: Berrueta Lisbeth , Kraeft Stine-Katherine , Tirnauer Jennifer S. , Schuyler Scott C. , Chen Lan Bo , Hill David E. , Pellman David , Bierer Barbara E.
Published: July 2002

The evolutionarily conserved protein EB1 originally was identified by its physical association with the carboxyl-terminal portion of the adenomatous polyposis coli (APC) tumor suppressor protein, an APC domain commonly mutated in familial and sporadic forms of colorectal neoplasia. The subcellular localization of EB1 in epithelial cells was studied by using immunofluorescence and biochemical techniques. EB1 colocalized both to cytoplasmic microtubules in interphase cells and to spindle microtubules during mitosis, with pronounced centrosome staining. The cytoskeletal array detected by anti-EB1 antibody was abolished by incubation of the cells with nocodazole, an agent that disrupts microtubules; upon drug removal, EB1 localized to the microtubule-organizing center. Immunofluorescence analysis of SW480, a colon cancer cell line that expresses only carboxyl-terminal-deleted APC unable to interact with EB1, demonstrated that EB1 remained localized to the microtubule cytoskeleton, suggesting that this pattern of subcellular distribution is not mediated by its interaction with APC.
In vitro
cosedimentation with taxol-stabilized microtubules demonstrated that a significant fraction of EB1 associated with microtubules. Recent studies of the yeast EB1 homologues Mal3 and Bim1p have demonstrated that both proteins localize to microtubules and are important
in vivo
for microtubule function. Our results demonstrate that EB1 is a novel component of the microtubule cytoskeleton in mammalian cells. Associating with the mitotic apparatus, EB1 may play a physiologic role connecting APC to cellular division, coordinating the control of normal growth and differentiation processes in the colonic epithelium.
Isolation of a high-affinity stable single-chain Fv specific for mesothelin from DNA-immunized mice by phage display and construction of a recombinant immunotoxin with anti-tumor activity
Authors: Chowdhury Partha S. , Viner Jaye L. , Beers Richard , Pastan Ira
Published: July 2002

Mesothelin is a differentiation antigen present on the surface of ovarian cancers, mesotheliomas, and several other types of human cancers. Because among normal tissues, mesothelin is present only on mesothelial cells, it represents a good target for antibody-mediated delivery of cytotoxic agents. In the present study mice were immunized with an eukaryotic expression vector coding for mesothelin. When high serum antibody titers were obtained, a phage display library was made from the splenic mRNA of these mice. After three rounds of panning on recombinant mesothelin, a single-chain Fv (scFv)-displaying phage was selected that bound specifically to recombinant mesothelin and mesothelin-positive cells. The scFv was used to construct an immunotoxin by genetically fusing it with a truncated mutant of
Pseudomonas
exotoxin A. The purified immunotoxin binds mesothelin with high affinity (Kd 11 nm), is stable for over 40 hr at 37°C and is very cytotoxic to cells expressing mesothelin. It also produces regressions of tumors expressing mesothelin. This combination of selective cytotoxicity, high activity, and stability makes the immunotoxin a good candidate for development as a therapeutic agent. This work also shows that DNA immunization can be used to isolate and clone antibodies against epitopes present on human proteins in their native conformation.
A mAb recognizing a surface antigen ofMycobacterium tuberculosisenhances host survival
Authors: Teitelbaum Rachel , Glatman-Freedman Aharona , Chen Bing , Robbins John B. , Unanue Emil , Casadevall Arturo , Bloom Barry R.
Published: July 2002

Murine mAbs reactive with the surface ofMycobacterium tuberculosiswere assayed for their ability to affect the course of infection in mice challenged with virulent organisms. An IgG3 mAb (9d8) specific for arabinomannan and reactive with purified antigen from a clinical isolate ofM. tuberculosisconferred partial protection on mice after respiratory challenge (30–60% survival >75 days;P≤ 0.05). Control mice pretreated with an irrelevant mAb of the same isotype succumbed to tuberculosis within 30 days. Mice with gene disruptions in interferon γ and major histocompatibility complex Class II also were partially protected from challenge. The protective mAb was neither bactericidal nor inhibitory of infection or bacterial replication. Nevertheless, it profoundly altered the nature of the granulomas in the infected lungs. Mice treated with mAb 9d8 and challenged withM. tuberculosislocalized the pathogen within granuloma centers, suggesting that the mAb conferred protection by enhancing a cellular immune response.
Structure-function analysis of integrase interactor 1/hSNF5L1 reveals differential properties of two repeat motifs present in the highly conserved region
Authors: Morozov Alexei , Yung Eric , Kalpana Ganjam V.
Published: July 2002

Retroviral integrase (IN) catalyzes the integration of retroviral cDNA into host chromosome. Ini1 (integrase interactor 1) is a host protein that specifically binds and stimulates
in vitro
joining activity of HIV-1 IN. Ini1 has homology to yeast transcription factor SNF5 and is a component of the analogous mammalian SWI/SNF complex that can remodel chromatin. Little is known about the function of Ini1 in mammalian cells. To gain insight into the functional domains of Ini1, and to understand the details of protein–protein interactions of IN and Ini1, a structure-function analysis of Ini1 was initiated. By means of the yeast two-hybrid system, the minimal IN binding domain of Ini1 was characterized. One of the two repeat motifs present in the highly conserved regions of Ini1 was found necessary and sufficient to bind to IN in yeast as well as
in vitro
. Because IN binds to only one of the two repeat motifs in this conserved region of Ini1, it appears that the IN-Ini1 interaction is very specific and functionally significant. Characterization of DNA-binding properties of Ini1 revealed that Ini1 can bind to plasmid DNA, binding more readily to supercoiled DNA than to the relaxed circular DNA. The minimal domain for DNA binding was localized to a region upstream of repeat 1. The DNA binding activity of Ini1 is not required for its ability to interact with IN. The finding that the two repeat motifs of Ini1 display differential binding to HIV-1 IN and that this discrete component of mammalian SWI/SNF complex binds to DNA will help understand the role of Ini1 in HIV-1 integration and in cellular process.
The effects of practice on the functional anatomy of task performance
Authors: Petersen Steven E. , van Mier Hanneke , Fiez Julie A. , Raichle Marcus E.
Published: July 2002

The effects of practice on the functional anatomy observed in two
different tasks, a verbal and a motor task, are reviewed in this paper.
In the first, people practiced a verbal production task, generating an
appropriate verb in response to a visually presented noun. Both
practiced and unpracticed conditions utilized common regions such as
visual and motor cortex. However, there was a set of regions that was
affected by practice. Practice produced a shift in activity from left
frontal, anterior cingulate, and right cerebellar hemisphere to
activity in Sylvian-insular cortex. Similar changes were also observed
in the second task, a task in a very different domain, namely the
tracing of a maze. Some areas were significantly more activated during
initial unskilled performance (right premotor and parietal cortex and
left cerebellar hemisphere); a different region (medial frontal cortex,
“supplementary motor area”) showed greater activity during
skilled performance conditions. Activations were also found in regions
that most likely control movement execution irrespective of skill level
(e.g., primary motor cortex was related to velocity of movement). One
way of interpreting these results is in a “scaffolding-storage”
framework. For unskilled, effortful performance, a scaffolding set of
regions is used to cope with novel task demands. Following practice, a
different set of regions is used, possibly representing storage of
particular associations or capabilities that allow for skilled
performance. The specific regions used for scaffolding and storage
appear to be task dependent.
Bcl-xL protects adult septal cholinergic neurons from axotomized cell death
Authors: Blömer U. , Kafri T. , Randolph-Moore L. , Verma I. M. , Gage F. H.
Published: July 2002

Bcl-xL suppresses apoptotic cell death induced by diverse stimuli in cell lines
in vitro
. To examine the mechanism by which axotomized cholinergic neurons die
in vivo
, lentiviral vectors expressing Bcl-xL, human nerve growth factor (hNGF), or green fluorescent protein were injected into the septum 3 weeks before transection of the fimbria fornix. Three weeks after transection, Bcl-xL- and hNGF-injected animals showed significantly higher numbers of spared cholinergic neurons compared with control (green fluorescent protein) injected animals. These results provide evidence that adult axotomized cholinergic neurons die of apoptotic death that can be prevented by local delivery of hNGF or intracellular delivery of Bcl-xL.
Multiple event activation of a generic prodrug trigger by antibody catalysis
Authors: Shabat Doron , Rader Christoph , List Benjamin , Lerner Richard A. , Barbas Carlos F.
Published: July 2002

Chemotherapeutic regimes are typically limited by nonspecific toxicity. To address this problem we have developed a broadly applicable drug-masking chemistry that operates in conjunction with a unique broad-scope catalytic antibody. This masking chemistry is applicable to a wide range of drugs because it is compatible with virtually any heteroatom. We demonstrate that generic drug-masking groups may be selectively removed by sequential retro-aldol–retro-Michael reactions catalyzed by antibody 38C2. This reaction cascade is not catalyzed by any known natural enzyme. Application of this masking chemistry to the anticancer drugs doxorubicin and camptothecin produced prodrugs with substantially reduced toxicity. These prodrugs are selectively unmasked by the catalytic antibody when it is applied at therapeutically relevant concentrations. We have demonstrated the efficacy of this approach by using human colon and prostate cancer cell lines. The antibody demonstrated a long
in vivo
half-life after administration to mice. Based on these findings, we believe that the system described here has the potential to become a key tool in selective chemotherapeutic strategies.
Proteolytic release and nuclear translocation of Notch-1 are induced by presenilin-1 and impaired by pathogenic presenilin-1 mutations
Authors: Song Weihong , Nadeau Philip , Yuan Menglan , Yang Xudong , Shen Jie , Yankner Bruce A.
Published: July 2002

The Notch family of proteins consists of transmembrane receptors that play a critical role in the determination of cell fate. Genetic studies in
Caenorhabditis elegans
suggest that the presenilin proteins, which are associated with familial Alzheimer’s disease, regulate Notch signaling. Here we show that proteolytic release of the Notch-1 intracellular domain (NICD), an essential step in the activation of Notch signaling, is markedly reduced in presenilin-1 (PS1)-deficient cells and is restored by PS1 expression. Nuclear translocation of the NICD is also markedly reduced in PS1-deficient cells, resulting in reduced transcriptional activation. Mutations in PS1 that are associated with familial Alzheimer’s disease impair the ability of PS1 to induce proteolytic release of the NICD and nuclear translocation of the cleaved protein. These results suggest that PS1 plays a central role in the proteolytic activation of the Notch-1-signaling pathway and that this function is impaired by pathogenic PS1 mutations. Thus, dysregulation of proteolytic function may underlie the mechanism by which presenilin mutations cause Alzheimer’s disease.
Committee proposals and restrictive rules
Authors: Banks Jeffrey S.
Published: July 2002

I analyze a game-theoretic model of committee–legislature interaction in which a majority decision to adopt either an open or closed amendment rule occurs following the committee’s proposal of a bill. I find that, in equilibrium, the closed rule is almost always chosen when the dimension of the policy space is >1. Furthermore, the difference between the equilibrium outcome and that which would have occurred under the open rule can be arbitrarily small.
Dimerization of zinc fingers mediated by peptides evolved
in vitro
from random sequences
Authors: Wang Bryan S. , Pabo Carl O.
Published: July 2002

Peptides that mediate dimerization of attached zinc finger DNA-binding domains have been evolved
in vitro
starting from random sequences. We first used phage display to select dimerization elements from libraries of random 15-residue polypeptides that were fused to the N terminus of the zinc finger domains. We then reoptimized these peptides by sequentially randomizing five-residue blocks (proceeding across the peptide in three steps) and selecting variant peptides that further stabilized the protein–DNA complex. Biochemical experiments confirmed that the selected peptides promote dimerization of the zinc fingers on an appropriate DNA target site. These results demonstrate that dimerization units can be obtained readily from random polypeptide libraries of moderate complexity. Our success reemphasizes the utility of searching random peptide libraries in protein design projects, and the sequences presented here may be useful when designing novel transcription factors.
Role of the type 5 adenovirus gene encoding the early region 1B 55-kDa protein in pulmonary pathogenesis
Authors: Ginsberg Harold S. , Moldawer Lyle L. , Prince Gregory A.
Published: July 2002

Comparison of the inflammatory response of
Sigmodon hispidus
cotton rats to pulmonary infection with wild-type 5 adenovirus (Ad5) or with a viral mutant, in which the early region 1B gene encoding a 55-kDa protein, Ad5dl110 (dl110), was deleted, indicated that the inflammation in animals infected with dl110 was markedly reduced compared with the inflammation in animals infected with wild-type Ad5, although both viruses replicated to the same extent. Comparable experiments done with C57BL/6 mice yielded identical results, even though only the early phase of gene expression essential for viral replication occurs in mice. Cytokine analysis of infected mouse lungs indicated that tumor necrosis factor-α and IL-6 were produced in relatively large quantities in wild-type Ad5-infected mice and at significantly lower levels in dl110-infected mice during the early stages of infection.
Imaging the functioning human brain
Authors: Weng Xuchu , Ding Yu-Shin , Volkow Nora D.
Published: July 2002

One of the most exciting methodological advances for brain
research field arises in functional brain imaging, which enables us to
localize and characterize neural activity and biochemical events in the
living human brain. Recently developed event-related functional MRI
makes it possible to visualize the brain activity associated with
cognitive processes with the temporal resolution of the hemodynamic
response. In addition, the high sensitivity and selectivity of
positron-emission tomography allow us to probe the neurochemical
processes at the molecular level. Positron-emission tomography also has
been applied to investigate the effects of therapeutic drugs as well as
the effects of drugs of abuse.
Dynamic force spectroscopy of single DNA molecules
Authors: Strunz Torsten , Oroszlan Krisztina , Schäfer Rolf , Güntherodt Hans-Joachim
Published: July 2002

To explore the analytic relevance of unbinding force measurements between complementary DNA strands with an atomic force microscope, we measured the forces to mechanically separate a single DNA duplex under physiological conditions by pulling at the opposite 5′-ends as a function of the loading rate (dynamic force spectroscopy). We investigated DNA duplexes with 10, 20, and 30 base pairs with loading rates in the range of 16–4,000 pN/s. Depending on the loading rate and sequence length, the unbinding forces of single duplexes varied from 20 to 50 pN. These unbinding forces are found to scale with the logarithm of the loading rate, which is interpreted in terms of a single energy barrier along the mechanical separation path. The parameters describing the energy landscape, i.e., the distance of the energy barrier to the minimum energy along the separation path and the logarithm of the thermal dissociation rate, are found to be proportional to the number of base pairs of the DNA duplex. These single molecule results allow a quantitative comparison with data from thermodynamic ensemble measurements and a discussion of the analytic applications of unbinding force measurements for DNA.
Genetic and environmental conditions that increase longevity in
Caenorhabditis elegans
decrease metabolic rate
Authors: Van Voorhies Wayne A. , Ward Samuel
Published: July 2002

Mutations that increase the longevity of the soil nematode
Caenorhabditis elegans
could define genes involved in a process specific for aging. Alternatively, these mutations could reduce animal metabolic rate and increase longevity as a consequence. In ectotherms, longevity is often negatively correlated with metabolic rate. Consistent with these observations, environmental conditions that reduce the metabolic rate of
C. elegans
also extend longevity. We found that the metabolic rate of long-lived
C. elegans
mutants is reduced compared with that of wild-type worms and that a genetic suppressor that restored normal longevity to long-lived mutants restored normal metabolic rate. Thus, the increased longevity of some long-lived
C. elegans
mutants may be a consequence of a reduction in their metabolic rate, rather than an alteration of a genetic pathway that leads to enhanced longevity while maintaining normal physiology. The actual mechanism responsible for the inverse correlation between metabolic rate and longevity remains unknown.
Cholesterol accumulation in tissues of the Niemann-Pick type C mouse is determined by the rate of lipoprotein-cholesterol uptake through the coated-pit pathway in each organ
Authors: Xie Chonglun , Turley Stephen D. , Dietschy John M.
Published: July 2002

Niemann-Pick type C (NPC) disease is associated with the accumulation of unesterified cholesterol in nearly all tissues and with progressive neurodegeneration. A murine model of this disease, the NPC mouse, was used to determine whether this sequestered cholesterol represented sterol carried in low density lipoprotein (LDL) and chylomicrons (CMs) taken up into the tissues through the coated-pit pathway. By 7 weeks of age, the sterol pool in the NPC mice had increased from 2,165 to 5,669 mg/kg body weight because of the daily sequestration of 67 mg of cholesterol per kg in the various organs. This was 7-fold greater than the rate of accumulation in control mice. The rate of LDL clearance in the NPC mouse was normal (523 ml/day per kg) and accounted for the uptake of 78 mg/day per kg of cholesterol in LDL whereas 8 mg/day per kg was taken up from CMs. Deletion of the LDL receptor in NPC mice altered the concentration of unesterified cholesterol in every organ in a manner consistent with the changes also observed in the rate of LDL cholesterol uptake in those tissues. Similarly, altering the flow of cholesterol to the liver through the CM pathway changed the concentration of unesterified cholesterol in that organ. Together, these observations strongly support the conclusion that, in NPC disease, it is cholesterol carried in LDL and CMs that is sequestered in the tissues and not sterol that is newly synthesized and carried in high density lipoprotein.
Intracellular localization of P1 ParB protein depends on ParA and
parS
Authors: Erdmann Natalie , Petroff Tamara , Funnell Barbara E.
Published: July 2002

The P1 partition system promotes faithful plasmid segregation during the
Escherichia coli
cell cycle. This system consists of two proteins, ParA and ParB, that act on a plasmid site called
parS
. By immunofluorescence microscopy, we observed that ParB localizes to discrete foci that are most often located close to the one-quarter and three-quarters positions of cell length. The visualization of ParB foci depended completely on the presence of
parS
, although their visualization was independent of the chromosomal context of
parS
(in P1 or the bacterial chromosome). In integration host factor-defective mutants, in which ParB binding to
parS
is weakened, only a fraction of the total pool of ParB had converged into foci. Taken together, these results indicate that
parS
recruits a pool of ParB into foci and that the resulting ParB–
parS
complexes serve as substrates for the segregation reaction. In the absence of ParA, the position of ParB foci in cells is perturbed, indicating that at least one of the roles of ParA is to direct ParB–
parS
complexes to the proper one-quarter positions from a cell pole. Finally, inhibition of cell division did not inhibit localization of ParB foci in cells, indicating that the positioning signals in the
E. coli
host that are needed for P1 partition do not depend on early division events.
Cooperative binding of ATP and MgADP in the sulfonylurea receptor is modulated by glibenclamide
Authors: Ueda Kazumitsu , Komine Jun , Matsuo Michinori , Seino Susumu , Amachi Teruo
Published: July 2002

The ATP-sensitive potassium (K
ATP
) channels in
pancreatic β cells are critical in the regulation of glucose-induced
insulin secretion. Although electrophysiological studies provide clues
to the complex control of K
ATP
channels by ATP, MgADP, and
pharmacological agents, the molecular mechanism of
K
ATP
-channel regulation remains unclear. The
K
ATP
channel is a heterooligomeric complex of SUR1 subunits
of the ATP-binding-cassette superfamily with two nucleotide-binding
folds (NBF1 and NBF2) and the pore-forming Kir6.2 subunits. Here, we
report that MgATP and MgADP, but not the Mg salt of γ-thio-ATP,
stabilize the binding of prebound 8-azido-[α-
32
P]ATP to
SUR1. Mutation in the Walker A and B motifs of NBF2 of SUR1 abolished
this stabilizing effect of MgADP. These results suggest that SUR1 binds
8-azido-ATP strongly at NBF1 and that MgADP, either by direct binding
to NBF2 or by hydrolysis of bound MgATP at NBF2, stabilizes prebound
8-azido-ATP binding at NBF1. The sulfonylurea glibenclamide caused
release of prebound 8-azido-[α-
32
P]ATP from SUR1 in the
presence of MgADP or MgATP in a concentration-dependent manner. This
direct biochemical evidence of cooperative interaction in nucleotide
binding of the two NBFs of SUR1 suggests that glibenclamide both blocks
this cooperative binding of ATP and MgADP and, in cooperation with the
MgADP bound at NBF2, causes ATP to be released from NBF1.
Mouse steroid receptor coactivator-1 is not essential for peroxisome proliferator-activated receptor α-regulated gene expression
Authors: Qi Chao , Zhu Yijun , Pan Jie , Yeldandi Anjana V. , Rao M. Sambasiva , Maeda Nobuyo , Subbarao V. , Pulikuri Sujata , Hashimoto Takashi , Reddy Janardan K.
Published: July 2002

Peroxisome proliferator-activated receptors (PPARs) are
ligand-dependent transcription factors, and it is assumed that the
biological effects of these receptors depend on interactions with
recently identified coactivators, including steroid receptor
coactivator-1 (SRC-1). We assessed the
in vivo
function
of SRC-1 on the PPARα-regulated gene expression in liver by
generating mice in which the SRC-1 gene was inactivated by gene
targeting. The homozygous (SRC-1
−/−
) mice were viable and
fertile and exhibited no detectable gross phenotypic defects. When
challenged with a PPARα ligand, such as ciprofibrate or Wy-14,643,
the SRC-1
−/−
mice displayed typical pleiotropic
responses, including hepatomegaly, peroxisome proliferation in
hepatocytes, and increased mRNA and protein levels of genes that are
regulated by PPARα. These alterations were indistinguishable from
those exhibited by SRC-1
+/+
wild-type mice fed either
ciprofibrate- or Wy-14,643-containing diets. These results indicate
that SRC-1 is not essential for PPARα-mediated transcriptional
activation
in vivo
and suggest redundancy in
nuclear receptor coactivators.
Tissue-specific processing of cocaine- and amphetamine-regulated transcript peptides in the rat
Authors: Thim Lars , Kristensen Peter , Nielsen Per F. , Wulff Birgitte S. , Clausen Jes T.
Published: July 2002

Cocaine- and amphetamine-regulated transcript (CART) is a recently discovered hypothalamic peptide regulated by leptin and with a potent appetite-suppressing activity. In the rat, the CART gene encodes a peptide of 116 amino acid residues (or a splice variant 13 residues longer). The predicted signal sequence is 27 amino acid residues, resulting in a prohormone of 89 residues. The CART prohormone contains several potential posttranslational processing sites in the form of mono- and dibasic sequences. In the present study we have purified CART peptides from extracts of adrenal gland, hypothalamus, nucleus accumbens, and pituitary gland (anterior and neurointermediate lobe) of the rat and determined the peptide structures by using microsequencing and mass spectrometry. In none of the tissues examined the long splice variant was found. From the adrenal gland, the CART(1–89) and CART(10–89) peptides were isolated, in contrast to the hypothalamus and nucleus accumbens, from which the shorter form peptides CART(42–89) and CART(49–89) were purified. From the anterior lobe of the pituitary gland, CART(42–89) was isolated, in contrast to the neurointermediate lobe, which contains only CART(49–89). This tissue-specific processing indicates that CART peptides may have different biological functions in the periphery and in the central nervous system.
Plaque-induced neurite abnormalities: Implications for disruption of neural networks in Alzheimer’s disease
Authors: Knowles Roger B. , Wyart Claire , Buldyrev Sergey V. , Cruz Luis , Urbanc Brigita , Hasselmo Michael E. , Stanley H. Eugene , Hyman Bradley T.
Published: July 2002

The brains of Alzheimer’s disease patients contain extracellular Aβ amyloid deposits (senile plaques). Although genetic evidence causally links Aβ deposition to the disease, the mechanism by which Aβ disrupts cortical function is unknown. Using triple immunofluorescent confocal microscopy and three-dimensional reconstructions, we found that neuronal processes that cross through an Aβ deposit are likely to have a radically changed morphology. We modeled the electrophysiological effect of this changed morphology and found a predicted delay of several milliseconds over an average plaque. We propose that this type of delay, played out among thousands of plaques throughout neocortical areas, disrupts the precise temporal firing patterns of action potentials, contributing directly to neural system failure and dementia.
Identification of
bdm-1
, a gene involved in G protein β-subunit function and α-subunit accumulation
Authors: Kasahara Shin , Wang Ping , Nuss Donald L.
Published: July 2002

Targeted disruption of Gα and Gβ genes has established the requirement of an intact G protein signaling pathway for optimal execution of several important physiological processes, including pathogenesis, in the chestnut blight fungus
Cryphonectria parasitica
. We now report the identification of a G protein signal transduction component,
b
eta
d
isruption
m
imic factor-
1
, BDM-1. Disruption of the corresponding gene,
bdm-1
, resulted in a phenotype indistinguishable from that previously observed after disruption of the Gβ subunit gene,
cpgb-1
. The BDM-1 deduced amino acid sequence contained several significant clusters of identity with mammalian phosducin, including a domain corresponding to a highly conserved 11-amino acid stretch that has been implicated in binding to the Gβγ dimer and two regions of defined Gβ/phosducin contact points. Unlike the negative regulatory function proposed for mammalian phosducin, the genetic data presented in this report suggest that BDM-1 is required for or facilitates Gβ function. Moreover, disruption of either
bdm-1
or
cpgb-1
resulted in a significant, posttranscriptional reduction in the accumulation of CPG-1, a key Gα subunit required for a range of vital physiological processes.
Direct activation of capsaicin receptors by products of lipoxygenases: Endogenous capsaicin-like substances
Authors: Hwang Sun Wook , Cho Hawon , Kwak Jiyeon , Lee Soon-Youl , Kang Chang-Joong , Jung Jooyoung , Cho Soohyun , Min Kyung Hoon , Suh Young-Ger , Kim Donghee , Oh Uhtaek
Published: July 2002

Capsaicin, a pungent ingredient of hot peppers, causes excitation of small sensory neurons, and thereby produces severe pain. A nonselective cation channel activated by capsaicin has been identified in sensory neurons and a cDNA encoding the channel has been cloned recently. However, an endogenous activator of the receptor has not yet been found. In this study, we show that several products of lipoxygenases directly activate the capsaicin-activated channel in isolated membrane patches of sensory neurons. Among them, 12- and 15-(
S
)-hydroperoxyeicosatetraenoic acids, 5- and 15-(
S
)-hydroxyeicosatetraenoic acids, and leukotriene B
4
possessed the highest potency. The eicosanoids also activated the cloned capsaicin receptor (VR1) expressed in HEK cells. Prostaglandins and unsaturated fatty acids failed to activate the channel. These results suggest a novel signaling mechanism underlying the pain sensory transduction.
NMR structure of the bovine prion protein
Authors: López García Francisco , Zahn Ralph , Riek Roland , Wüthrich Kurt
Published: July 2002

The NMR structures of the recombinant 217-residue polypeptide chain of the mature bovine prion protein, bPrP(23–230), and a C-terminal fragment, bPrP(121–230), include a globular domain extending from residue 125 to residue 227, a short flexible chain end of residues 228–230, and an N-terminal flexibly disordered “tail” comprising 108 residues for the intact protein and 4 residues for bPrP(121–230), respectively. The globular domain contains three α-helices comprising the residues 144–154, 173–194, and 200–226, and a short antiparallel β-sheet comprising the residues 128–131 and 161–164. The best-defined parts of the globular domain are the central portions of the helices 2 and 3, which are linked by the only disulfide bond in bPrP. Significantly increased disorder and mobility is observed for helix 1, the loop 166–172 leading from the β-strand 2 to helix 2, the end of helix 2 and the following loop, and the last turn of helix 3. Although there are characteristic local differences relative to the conformations of the murine and Syrian hamster prion proteins, the bPrP structure is essentially identical to that of the human prion protein. On the other hand, there are differences between bovine and human PrP in the surface distribution of electrostatic charges, which then appears to be the principal structural feature of the “healthy” PrP form that might affect the stringency of the species barrier for transmission of prion diseases between humans and cattle.
Voltage-sensor sodium channel mutations cause hypokalemic periodic paralysis type 2 by enhanced inactivation and reduced current
Authors: Jurkat-Rott Karin , Mitrovic Nenad , Hang Chao , Kouzmenkine Alexei , Iaizzo Paul , Herzog Jürgen , Lerche Holger , Nicole Sophie , Vale-Santos Jose , Chauveau Dominique , Fontaine Bertrand , Lehmann-Horn Frank
Published: July 2002

The pathomechanism of familial hypokalemic periodic paralysis
(HypoPP) is a mystery, despite knowledge of the underlying
dominant point mutations in the dihydropyridine receptor (DHPR) voltage
sensor. In five HypoPP families without DHPR gene defects, we
identified two mutations, Arg-672→His and →Gly, in the voltage
sensor of domain 2 of a different protein: the skeletal muscle sodium
channel α subunit, known to be responsible for hereditary muscle
diseases associated with myotonia. Excised skeletal muscle fibers from
a patient heterozygous for Arg-672→Gly displayed depolarization and
weakness in low-potassium extracellular solution. Slowing and smaller
size of action potentials were suggestive of excitability of the
wild-type channel population only. Heterologous expression of the two
sodium channel mutations revealed a 10-mV left shift of the
steady-state fast inactivation curve enhancing inactivation and a
sodium current density that was reduced even at potentials at which
inactivation was removed. Decreased current and small action potentials
suggested a low channel protein density. The alterations are decisive
for the pathogenesis of episodic muscle weakness by reducing the number
of excitable sodium channels particularly at sustained membrane
depolarization. The results prove that SCN4A, the gene encoding the
sodium channel α subunit of skeletal muscle is responsible for
HypoPP-2 which does not differ clinically from DHPR-HypoPP. HypoPP-2
represents a disease caused by enhanced channel inactivation and
current reduction showing no myotonia.
Cloning and characterization of IL-17B and IL-17C, two new members of the IL-17 cytokine family
Authors: Li Hanzhong , Chen Jian , Huang Arthur , Stinson Jeremy , Heldens Sherry , Foster Jessica , Dowd Patrick , Gurney Austin L. , Wood William I.
Published: July 2002

IL-17 is a T cell-derived cytokine that may play an important role in the initiation or maintenance of the proinflammatory response. Whereas expression of IL-17 is restricted to activated T cells, the IL-17 receptor is found to be widely expressed, a finding consistent with the pleiotropic activities of IL-17. We have cloned and expressed two novel human cytokines, IL-17B and IL-17C, that are related to IL-17 (≈27% amino acid identity). IL-17B mRNA is expressed in adult pancreas, small intestine, and stomach, whereas IL-17C mRNA is not detected by RNA blot hybridization of several adult tissues. No expression of IL-17B or IL-17C mRNA is found in activated T cells. In a survey of cytokine induction, IL-17B and IL-17C stimulate the release of tumor necrosis factor α and IL-1β from the monocytic cell line, THP-1, whereas IL-17 has only a weak effect in this system. No induction of IL-1α, IL-6, IFN-γ, or granulocyte colony-stimulating factor is found in THP-1 cells. Fluorescence-activated cell sorter analysis shows that IL-17B and IL-17C bind to THP-1 cells. Conversely, IL-17B and IL-17C are not active in an IL-17 assay or the stimulation of IL-6 release from human fibroblasts and do not bind to the human IL-17 receptor extracellular domain. These data show that there is a family of IL-17-related cytokines differing in patterns of expression and proinflammatory responses that may be transduced through a cognate set of cell surface receptors.
Direct isolation of human central nervous system stem cells
Authors: Uchida Nobuko , Buck David W. , He Dongping , Reitsma Michael J. , Masek Marilyn , Phan Thinh V. , Tsukamoto Ann S. , Gage Fred H. , Weissman Irving L.
Published: July 2002

Stem cells, which are clonogenic cells with self-renewal and
multilineage differentiation properties, have the potential to replace
or repair damaged tissue. We have directly isolated clonogenic human
central nervous system stem cells (hCNS-SC) from fresh human fetal
brain tissue, using antibodies to cell surface markers and
fluorescence-activated cell sorting. These hCNS-SC are phenotypically
5F3 (CD133)
+
, 5E12
+
, CD34
−
,
CD45
−
, and CD24
−/lo
. Single
CD133
+
CD34
−
CD45
−
sorted cells
initiated neurosphere cultures, and the progeny of clonogenic cells
could differentiate into both neurons and glial cells. Single cells
from neurosphere cultures initiated from CD133
+
CD34
−
CD45
−
cells were again replated as
single cells and were able to reestablish neurosphere cultures,
demonstrating the self-renewal potential of this highly enriched
population. Upon transplantation into brains of immunodeficient
neonatal mice, the sorted/expanded hCNS-SC showed potent engraftment,
proliferation, migration, and neural differentiation.
Role of AMP-activated protein kinase in the regulation by glucose of islet beta cell gene expression
Authors: da Silva Xavier Gabriela , Leclerc Isabelle , Salt Ian P. , Doiron Bruno , Hardie D. Grahame , Kahn Axel , Rutter Guy A.
Published: July 2002

Elevated glucose concentrations stimulate the transcription of the pre-proinsulin (PPI), L-type pyruvate kinase (L-PK), and other genes in islet beta cells. In liver cells, pharmacological activation by 5-amino-4-imidazolecarboxamide riboside (AICAR) of AMP-activated protein kinase (AMPK), the mammalian homologue of the yeast SNF1 kinase complex, inhibits the effects of glucose, suggesting a key signaling role for this kinase. Here, we demonstrate that AMPK activity is inhibited by elevated glucose concentrations in MIN6 beta cells and that activation of the enzyme with AICAR prevents the activation of the L-PK gene by elevated glucose. Furthermore, microinjection of antibodies to the α2- (catalytic) or β2-subunits of AMPK complex, but not to the α1-subunit or extracellular stimulus-regulated kinase, mimics the effects of elevated glucose on the L-PK and PPI promoter activities as assessed by single-cell imaging of promoter luciferase constructs. In each case, injection of antibodies into the nucleus and cytosol, but not the nucleus alone, was necessary, indicating the importance of either a cytosolic phosphorylation event or the subcellular localization of the α2-subunits. Incubation with AICAR diminished, but did not abolish, the effect of glucose on PPI transcription. These data suggest that glucose-induced changes in AMPK activity are necessary and sufficient for the regulation of the L-PK gene by the sugar and also play an important role in the regulation of the PPI promoter.
Mega-urban Regions and World City Formation: Globalisation, the Economic Crisis and Urban Policy Issues in Pacific Asia
Authors: Douglass Mike
Published: July 2002

In Pacific Asia, the globalisation of trade, production and finance underlies an accelerated urban transition focusing on a limited number of mega-urban regions. Intercity competition for world city status among these regions has intensified following the 1997 economic crisis. With governments compelled to devote greater amounts of public resources to creating a built environment to host global investment, a number of key policy issues are emerging. These include demands for inclusive governance and more livable cities; the appearance of new forms of urban poverty; low economic resilience in the face of growing global economic turbulence and the spatial unevenness of global economic growth. With the rise of civil society as a political force, addressing these issues calls for a sharper focus on cities not simply as economic agglomerations or collectivities of consumers in the world market, but also as arenas for the formation of political communities.
From Goddess Spirituality to Irigaray's Angel: The Politics of the Divine
Authors: Ingram Penelope
Published: July 2002

This article argues that the act of conceptualizing a female divine, whether by so-called low-brow Goddess Spiritualists or high-brow French philosophers, rather than being a mere spiritual exercise, has enormous political significance for feminisms. In particular, I demonstrate that Irigaray's concept of the sensible transcendental, by refiguring a god which is both male and female, transcendent and immanent, theorizes a potential dissolution of the binary logic which forms the basis of western philosophy. The second half of the article looks at the complex role of the angel in Irigaray's theory of the divine in order to demonstrate that, as a result of having misinterpreted the angel's position, critics have failed to recognize the significance and relevance of Irigaray's sensible transcendental for feminist politics.
Growing up Young, Asian and Female in Britain: A Report on Self-harm and Suicide
Authors: Bhardwaj Anita
Published: July 2002

It is now a well-documented statistic that Asian women between the ages of 15–35 are two to three times more vulnerable to suicide and self-harm than their non-Asian counterparts (Soni-Raleigh, 1996). The article will summarize the findings of the research report Growing Up Young, Asian and Female in Britain (Newham Asian Women's Project, 1998), which aimed to explore the reasons why young Asian women self-harmed and to evaluate the service responses they were given. It also made recommendations aimed at decreasing the high prevalence of suicide and self-harm among young Asian women. The article concludes by reflecting upon the impact and service developments that have been made since the report was published.
Embodying Transition: FGC, Displacement, and Gender-making for Sudanese in Cairo
Authors: Fábos Anita Häusermann
Published: July 2002

In this article I analyze both generalized propriety as a boundary marker of Sudanese identity in Cairo, and gendered attitudes toward morality and female genital cutting (FGC) as a fundamental aspect of that boundary. Sudanese have been profoundly affected by the ongoing political crisis in their home country, by the displacement triggered by political and economic collapse, and by their deteriorating legal and social status in Egypt. The dramatic changes in the circumstances of Sudanese residence in Cairo have challenged the cultural norm of gender complementarity as men ‘stay at home’ for want of work while women seek and find new opportunities for themselves. This unstable situation has led Sudanese to place more emphasis on ‘proper’ ways of behaving and being, an assertion that helps define the ethnic boundaries of the Sudanese community in Cairo. I demonstrate the inconsistencies between discourse and reality through ethnographic data while analyzing how Sudanese have found new ways of asserting their identity and resisting the practice of FGC.
Theorizing Gender Systems and the Project of Modernity in the Twentieth-Century Caribbean
Authors: Barriteau Eudine
Published: July 2002

A central thesis of this paper is that the philosophical contradictions of liberal ideologies predispose states to institute unjust gender systems. I argue that postcolonial Caribbean states have inherited a complexity of social relations and structures from the Enlightenment discourses of Liberalism, yet they seem unaware that the discourses which created colonialism and Western expansion were themselves part of the Enlightenment project of modernity. In this paper I apply this theoretical framework to a historical analysis of gender systems in the twentieth-century Caribbean. The paper examines three distinct periods: 1900–37, 1937–50s and 1950s-90s, the transition from colonial to postcolonial modernizing societies, and attempts to generate a gendered analytical model which can be widely applied both within and outside of the region.