bluuebunny/crossref_metadata_2025 huggingface dataset:
Our ancestral physiological phenotype: An adaptation for hypoxia tolerance and for endurance performance?
Authors: Hochachka Peter W. , Gunga Hanns Christian , Kirsch Karl
Published: July 2002

There are well known mechanistic similarities in human physiology between adaptations for endurance performance and hypoxia tolerance. By using background principles arising from recent studies of the evolution of the diving response in marine mammals, here we analyze human responses to hypobaric hypoxia based on studies with several different low and high altitude human lineages. As in the evolution of the diving response in pinnipeds, we found “conservative” and “adaptable” physiological characters involved in human responses to hypoxia. Because the analysis concerns traits within a single species, conservative characters dominate the picture (they define basic human physiology and largely are independent of environmental parameters). Most notably, we also found evidence for adaptable characters forming the foundations for a fairly unique physiological phenotype—a low capacity version favored under hypobaric hypoxia and a high capacity one favored for endurance performance. Because current evidence implies that the human species arose under conditions that were getting colder, drier, and higher (situations in which these traits would have been advantageous), we hypothesize that this physiology is our “ancestral” condition.
Response regulators implicated in His-to-Asp phosphotransfer signaling in
Arabidopsis
Authors: Imamura Aya , Hanaki Naoto , Umeda Hiroyuki , Nakamura Ayako , Suzuki Tomomi , Ueguchi Chiharu , Mizuno Takeshi
Published: July 2002

The His to Asp phosphotransfer signal transduction mechanism involves three common signaling domains: the transmitter (or His-kinase), the receiver, and the histidine-containing phototransfer (HPt) domain. Typically, a sensor kinase has a His-kinase domain and a response regulator has a receiver domain containing a phosphoaccepting aspartate, whereas a histidine-containing phototransfer domain serves as a mediator of the histidine-to-aspartate phosphotransfer. This signal transduction mechanism was thought to be restricted to prokaryotes. However, many examples have been discovered in diverse eukaryotic species including higher plants. In
Arabidopsis
, three sensor kinases have been characterized, namely, ETR1, ERS, and CKI1, which were suggested to be involved in ethylene- and cytokinin-dependent signal transduction pathways, respectively. To date, no response regulator has been discovered in higher plants. We identify five distinct
Arabidopsis
response regulator genes, each encoding a protein containing a receiver-like domain.
In vivo
and
in vitro
evidence that ARRs can function as phosphoaccepting response regulators was obtained by employing the
Escherichia coli
His-Asp phosphotransfer signaling system.
A single-headed dimer of
Escherichia coli
ribosomal protein L7/L12 supports protein synthesis
Authors: Oleinikov Andrew V. , Jokhadze George G. , Traut Robert R.
Published: July 2002

During protein synthesis, the two elongation factors Tu and G alternately bind to the 50S ribosomal subunit at a site of which the protein L7/L12 is an essential component. L7/L12 is present in each 50S subunit in four copies organized as two dimers. Each dimer consists of distinct domains: a single N-terminal (“tail”) domain that is responsible for both dimerization and binding to the ribosome via interaction with the protein L10 and two independent globular C-terminal domains (“heads”) that are required for binding of elongation factors to ribosomes. The two heads are connected by flexible hinge sequences to the N-terminal domain. Important questions concerning the mechanism by which L7/L12 interacts with elongation factors are posed by us in response to the presence of two dimers, two heads per dimer, and their dynamic, mobile properties. In an attempt to answer these questions, we constructed a single-headed dimer of L7/L12 by using recombinant DNA techniques and chemical cross-linking. This chimeric molecule was added to inactive core particles lacking wild-type L7/L12 and shown to restore activity to a level approaching that of wild-type two-headed L7/L12.
Disruption of the CD4–major histocompatibility complex class II interaction blocks the development of CD4+T cellsin vivo
Authors: Riberdy Janice M. , Mostaghel Elahe , Doyle Carolyn
Published: July 2002

The experiments presented in this report were designed to specifically examine the role of CD4–major histocompatibility complex (MHC) class II interactions during T cell developmentin vivo. We have generated transgenic mice expressing class II molecules that cannot interact with CD4 but that are otherwise competent to present peptides to the T cell receptor. MHC class II expression was reconstituted in Aβ gene knock-out mice by injection of a transgenic construct encoding either the wild-type I-Aβbprotein or a construct encoding a mutation designed to specifically disrupt binding to the CD4 molecule. We demonstrate that the mutation, EA137 and VA142 in the β2 domain of I-Ab, is sufficient to disrupt CD4–MHC class II interactionsin vivo. Furthermore, we show that this interaction is critical for the efficient selection of a complete repertoire of mature CD4+T helper cells as evidenced by drastically reduced numbers of conventional CD4+T cells in animals expressing the EA137/VA142 mutant I-Aband by the failure to positively select the transgenic AND T cell receptor on the mutated I-Ab. These results underscore the importance of the CD4–class II interaction in the development of mature peripheral CD4+T cells.
Dehydroepiandrosterone: A potential signalling molecule for neocortical organization during development
Authors: Compagnone Nathalie A. , Mellon Synthia H.
Published: July 2002

Dehydroepiandrosterone (DHEA) and its sulfate derivative (DHEAS) are the most abundant steroids produced by the human adrenal, but no receptors have been identified for these steroids, and no function for them has been established, other than as precursors for sex steroid synthesis. DHEA and DHEAS are found in brains from many species, and we have shown that enzymes crucial for their synthesis, especially P450c17 (17α-hydroxylase/c17,20 lyase), are expressed in a developmentally regulated, region-specific fashion in the developing rodent brain. One region of embryonic expression of P450c17, the neocortical subplate, has been postulated to play a role in guiding cortical projections to their appropriate targets. We therefore determined if products of P450c17 activity, DHEA and DHEAS, regulated the motility and/or growth of neocortical neurons. In primary cultures of mouse embryonic neocortical neurons, DHEA increased the length of neurites containing the axonal marker Tau-1, and the incidence of varicosities and basket-like process formations in a dose-dependent fashion. These effects could be seen at concentrations normally found in the brain. By contrast, DHEAS had no effect on Tau-1 axonal neurites but increased the length of neurites containing the dendritic marker microtubule-associated protein-2. DHEA rapidly increased free intracellular calcium via activation of
N
-methyl-
d
-aspartate (NMDA) receptors. These studies provide evidence of mechanisms by which DHEA and DHEAS exert biological actions, show that they have specific functions other than as sex steroid precursors, mediate their effects
via
non-classic steroid hormone receptors, and suggest that their developmentally regulated synthesis
in vivo
may play crucial and different roles in organizing the neocortex.
Evolution of rhizobia by acquisition of a 500-kb symbiosis island that integrates into a phe-tRNA gene
Authors: Sullivan John T. , Ronson Clive W.
Published: July 2002

Nodulation and nitrogen fixation genes of
Mesorhizobium loti
are encoded on the chromosome of the bacterium. Nevertheless, there is strong evidence that these genes can be transferred from an inoculant strain to nonsymbiotic mesorhizobia in the field environment. Here we report that the chromosomal symbiotic element of
M. loti
strain ICMP3153 is transmissible in laboratory matings to at least three genomic species of nonsymbiotic mesorhizobia. The element is 500 kb in size, integrates into a phe-tRNA gene, and encodes an integrase of the phage P4 family just within its left end. The entire phe-tRNA gene is reconstructed at the left end of the element upon integration, whereas the 3′ 17 nucleotides of the tRNA gene are present as a direct repeat at the right end. We termed the element a symbiosis island on the basis of its many similarities to pathogenicity islands. It may represent a class of genetic element that contributes to microbial evolution by acquisition.
Self-assembly of chlorophenols in water
Authors: Rogalska Ewa , Rogalski Marek , Gulik-Krzywicki Tadeusz , Gulik Annette , Chipot Christophe
Published: July 2002

In saturated solutions of some di- and trichlorophenols, structures with complex morphologies, consisting of thin, transparent sheets often coiling into helices and ultimately twisting into filaments, were observed under the optical microscope. Freeze-fracture electron microscopy, x-ray diffraction, phase diagrams, and molecular modeling were performed to elucidate the observed phenomena. Here, we present evidence that the chlorophenols studied, when interacting with water, self-assemble into bilayers. The fact that some chlorophenols form the same supramolecular structures as those described previously for structurally nonrelated surfactants sheds light on the mechanisms of self-assembly.
Asymmetric DNA bending in the Cre-
loxP
site-specific recombination synapse
Authors: Guo Feng , Gopaul Deshmukh N. , Van Duyne Gregory D.
Published: July 2002

Cre recombinase catalyzes site-specific recombination between two 34-bp
loxP
sites in a variety of DNA substrates. At the start of the recombination pathway, the
loxP
sites are each bound by two recombinase molecules, and synapsis of the sites is mediated by Cre–Cre interactions. We describe the structures of synaptic complexes formed between a symmetrized
loxP
site and two Cre mutants that are defective in strand cleavage. The DNA in these complexes is bent sharply at a single base pair step at one end of the crossover region in a manner that is atypical of protein-induced DNA bends. A large negative roll (−49°) and a positive tilt (16°) open the major groove toward the center of the synapse and compress the minor groove toward the protein–DNA interface. The bend direction of the site appears to determine which of the two DNA substrate strands will be cleaved and exchanged in the initial stages of the recombination pathway. These results provide a structural basis for the observation that exchange of DNA strands proceeds in a defined order in some tyrosine recombinase systems. The Cre-loxS synaptic complex structure supports a model in which synapsis of the
loxP
sites results in formation of a Holliday junction-like DNA architecture that is maintained through the initial cleavage and strand exchange steps in the site-specific recombination pathway.
The C-terminal region of
Schizosaccaromyces pombe
proliferating cell nuclear antigen is essential for DNA polymerase activity
Authors: Kelman Zvi , Zuo Shaojun , Arroyo Martha P. , Wang Teresa S.-F. , Hurwitz Jerard
Published: July 2002

Proliferating cell nuclear antigen (PCNA), the processivity factor (sliding clamp) of DNA polymerases (Pols), plays essential roles in DNA metabolism. In this report, we examined the functional role of the C-terminal region of
Schizosaccaromyces pombe
PCNA both
in vitro
and
in vivo
. The deletion or Ala substitution of the last 9 aa (252–260A), as well as Ala replacement of only 4 aa (252–255A) at the C terminus, failed to substitute for the wild-type PCNA protein for cell growth in
S. pombe
. Two other PCNA mutant proteins, A251V and K253E, exhibited cold-sensitive phenotypes. Several yeast strains harboring mutations, including those at the acidic C-terminal region, showed elevated sensitivity to DNA damage. The ability of the mutant PCNA proteins to stimulate DNA synthesis by Polδ and Polɛ also was studied
in vitro
. The mutant proteins that did not support cell growth and a mutant protein containing a single amino acid substitution at position 252, where Pro is replaced by Ala, stimulated Polδ and Polɛ activities poorly. All mutant PCNA proteins, however, were assembled around DNA by the clamp loader, replication factor C, efficiently. Thus, the C-terminal region of PCNA is important for interactions with both Polδ and Polɛ and for cell survival after DNA damage. The C terminus of sliding clamps from other organisms has been shown to be important for clamp loading as well as polymerase interactions. The relationship between the conserved sequence in this region in different organisms is discussed.
Role for yeast inhibitor of apoptosis (IAP)-like proteins in cell division
Authors: Uren Anthony G. , Beilharz Traude , O’Connell Matthew J. , Bugg Sarah J. , van Driel Rosemary , Vaux David L. , Lithgow Trevor
Published: July 2002

Inhibitors of apoptosis (IAPs) are a family of proteins that bear baculoviral IAP repeats (BIRs) and regulate apoptosis in vertebrates and
Drosophila melanogaster
. The yeasts
Saccharomyces cerevisiae
and
Schizosaccharomyces pombe
both encode a single IAP, designated
BIR1
and
bir1
, respectively, each of which bears two BIRs. In rich medium,
BIR1
mutant
S. cerevisiae
underwent normal vegetative growth and mitosis. Under starvation conditions, however,
BIR1
mutant diploids formed spores inefficiently, instead undergoing pseudohyphal differentiation. Most spores that did form failed to survive beyond two divisions after germination.
bir1
mutant
S. pombe
spores also died in the early divisions after spore germination and became blocked at the metaphase/anaphase transition because of an inability to elongate their mitotic spindle. Rather than inhibiting caspase-mediated cell death, yeast IAP proteins have roles in cell division and appear to act in a similar way to the IAPs from
Caenorhabditis elegans
and the mammalian IAP Survivin.
Thermal avoidance in
Caenorhabditis elegans
: An approach to the study of nociception
Authors: Wittenburg Nicole , Baumeister Ralf
Published: July 2002

Upon perception of a noxious stimulus, an organism executes defense mechanisms, such as escape responses. The molecular basis of these mechanisms is poorly understood. In this paper we show that upon exposure to noxious temperature,
Caenorhabditis elegans
reacts by a withdrawal reflex. To analyze this thermal avoidance behavior, we developed a laser-based assay to quantify the response. The escape reflex can be observed in 98% of the adult animals, but is not executed in animals in diapause. The thermal avoidance response differs significantly from the thermotaxis behavior that is based on the perception of physiological temperature. It involves different neurons and is influenced by mutations in distinct genes. As in mammals, the strength of the thermal avoidance response is increased by application of capsaicin, the pungent ingredient in chili peppers. We find that thermal avoidance is strongly reduced in mutants affecting the neural transmission modulated by glutamate and neuropeptides as well as in mutants affecting the structure and function of sensory neurons. We suggest that the study of this nociceptive behavior in
C. elegans
can be used to understand the genetic and molecular basis of thermal nociception.
Abortive expansion of the cumulus and impaired fertility in mice lacking the prostaglandin E receptor subtype EP
2
Authors: Hizaki Hiroko , Segi Eri , Sugimoto Yukihiko , Hirose Masaya , Saji Tomomi , Ushikubi Fumitaka , Matsuoka Toshiyuki , Noda Yoichi , Tanaka Takashi , Yoshida Nobuaki , Narumiya Shuh , Ichikawa Atsushi
Published: July 2002

Female mice lacking the gene encoding the prostaglandin (PG) E
2
receptor subtype EP
2
(EP
2
−/−
) become pregnant and deliver their pups at term, but with a much reduced litter size. A decrease in ovulation number and a much reduced fertilization rate were observed in EP
2
−/−
females without difference of the uterus to support implantation of wild-type embryos. Treatment with gonadotropins induced EP
2
mRNA expression in the cumulus cells of ovarian follicles of wild-type mice. The immature cumuli oophori from wild-type mice expanded
in vitro
in response to both follicle-stimulating hormone and PGE
2
, but the response to PGE
2
was absent in those from EP
2
−/−
mice. Cumulus expansion proceeded normally in preovulatory follicles but became abortive in a number of ovulated complexes in EP
2
−/−
mice, indicating that EP
2
is involved in cumulus expansion in the oviduct
in vivo
. No difference in the fertilization rate between wild-type and EP
2
−/−
mice was found in
in vitro
studies using cumulus-free oocytes. These results indicate that PGE
2
cooperates with gonadotropin to complete cumulus expansion for successful fertilization.
Vacuole acidification is required for trans-SNARE pairing, LMA1 release, and homotypic fusion
Authors: Ungermann Christian , Wickner William , Xu Zuoyu
Published: July 2002

Vacuole fusion occurs in three stages: priming, in which Sec18p mediates Sec17p release, LMA1 (low
M
r
activity 1) relocation, and cis
-
SNARE (soluble
N
-ethylmaleimide-sensitive factor attachment protein receptor) complex disassembly; docking, mediated by Ypt7p and trans-SNARE association; and fusion of docked vacuoles. Ca
2+
and calmodulin regulate late stages of the reaction. We now show that the vacuole proton gradient, generated by the vacuolar proton ATPase, is needed for trans-SNARE complex formation during docking and hence for the subsequent LMA1 release. Though neither the vacuolar Pmc1p Ca
2+
-ATPase nor the Vcx1p Ca
2+
/H
+
exchanger are needed for the fusion reaction, they participate in Ca
2+
and Δμ
H
+
homeostasis. Fusion itself does not require the maintenance of trans-SNARE pairs.
Basolateral amygdala noradrenergic influence enables enhancement of memory consolidation induced by hippocampal glucocorticoid receptor activation
Authors: Roozendaal Benno , Nguyen Bichngoc T. , Power Ann E. , McGaugh James L.
Published: July 2002

Previously, we reported that bilateral excitotoxic lesions of the basolateral nucleus of the amygdala (BLA) block the enhancing effects of posttraining systemic or intrahippocampal glucocorticoid administration on memory for inhibitory avoidance training. The present study further examined the basis of this permissive influence of the BLA on hippocampal memory functioning. Immediate posttraining unilateral infusions of the specific glucocorticoid receptor agonist RU 28362 (11β,17β-dihydroxy-6,21-dimethyl-17α-pregna-4,6-trien-20-yn-3-one; 3.0, 10.0, or 30.0 ng in 0.5 μl) administered into the dorsal hippocampus of male Sprague–Dawley rats induced dose-dependent enhancement of 48-h inhibitory avoidance retention. Infusions of the β-adrenoceptor antagonist atenolol (0.5 μg in 0.2 μl) into the ipsilateral, but not the contralateral, BLA 10 min prior to training blocked the hippocampal glucocorticoid effects on memory consolidation. Infusions of the muscarinic cholinergic antagonist atropine (0.5 μg in 0.2 μl) into either the ipsilateral or contralateral BLA before training did not block the hippocampal glucocorticoid effects. These findings provide further evidence that β-adrenergic activity in the BLA is essential in enabling glucocorticoid-induced modulation of memory consolidation and are consistent with the hypothesis that the BLA regulates the strength of memory consolidation in other brain structures. The ipsilateral nature of the BLA–hippocampus interaction indicates that BLA influences on hippocampal memory processes are mediated through neural pathways rather than by influences by means of the activation of peripheral stress responses.
RAG2is regulated differentially in B and T cells by elements 5′ of the promoter
Authors: Monroe Robert J. , Chen Feng , Ferrini Roger , Davidson Laurie , Alt Frederick W.
Published: July 2002

To studyRAG2gene regulationin vivo, we developed a blastocyst complementation method in whichRAG2-deficient embryonic stem cells were transfected with genomic clones containingRAG2and then assessed for their ability to generate lymphocytes. ARAG2genomic clone that contained only theRAG2promoter sequences rescued V(D)J recombination inRAG2-deficient pro-B cell lines, but did not rescue development ofRAG2-deficient lymphocytesin vivo. However, inclusion of varying lengths of sequences 5′ of theRAG2promoter generated constructs capable of rescuing onlyin vivoB cell development, as well as other constructs that rescued both B and T cell development. In particular, the 2-kb 5′ region starting just upstream of the RAG2 promoter, as well as the region from 2–7 kb 5′, could independently drive B cell development, but not efficient T cell development. Deletion of the 2-kb 5′ region from the murine germ line demonstrated that this region was not required forRAGexpression sufficient to generate normal B or T cell numbers, implying redundancy among 5′ elements. We conclude thatRAG2expressionin vivorequires elements beyond the core promoter, that such elements contribute to differential regulation in the B vs. T lineages, and that sequences sufficient to direct B cell expression are located in the promoter-proximal 5′ region.
Interactions among enzymes of the
Arabidopsis
flavonoid biosynthetic pathway
Authors: Burbulis Ian E. , Winkel-Shirley Brenda
Published: July 2002

Flavonoids are secondary metabolites derived from phenylalanine and acetate metabolism that perform a variety of essential functions in higher plants. Studies over the past 30 years have supported a model in which flavonoid metabolism is catalyzed by an enzyme complex localized to the endoplasmic reticulum [Hrazdina, G. & Wagner, G. J. (1985)
Arch. Biochem. Biophys.
237, 88–100]. To test this model further we assayed for direct interactions between several key flavonoid biosynthetic enzymes in developing
Arabidopsis
seedlings. Two-hybrid assays indicated that chalcone synthase, chalcone isomerase (CHI), and dihydroflavonol 4-reductase interact in an orientation-dependent manner. Affinity chromatography and immunoprecipitation assays further demonstrated interactions between chalcone synthase, CHI, and flavonol 3-hydroxylase in lysates from
Arabidopsis
seedlings. These results support the hypothesis that the flavonoid enzymes assemble as a macromolecular complex with contacts between multiple proteins. Evidence was also found for posttranslational modification of CHI. The importance of understanding the subcellular organization of elaborate enzyme systems is discussed in the context of metabolic engineering.
Abnormal neurotransmission in mice lacking synaptic vesicle protein 2A (SV2A)
Authors: Crowder Kelly M. , Gunther Jane M. , Jones Theresa A. , Hale Brian D. , Zhang Hai Zhuan , Peterson Michael R. , Scheller Richard H. , Chavkin Charles , Bajjalieh Sandra M.
Published: July 2002

Synaptic vesicle protein 2 (SV2) is a membrane glycoprotein common to all synaptic and endocrine vesicles. Unlike many proteins involved in synaptic exocytosis, SV2 has no homolog in yeast, indicating that it performs a function unique to secretion in higher eukaryotes. Although the structure and protein interactions of SV2 suggest multiple possible functions, its role in synaptic events remains unknown. To explore the function of SV2 in an
in vivo
context, we generated mice that do not express the primary SV2 isoform, SV2A, by using targeted gene disruption. Animals homozygous for the SV2A gene disruption appear normal at birth. However, they fail to grow, experience severe seizures, and die within 3 weeks, suggesting multiple neural and endocrine deficits. Electrophysiological studies of spontaneous inhibitory neurotransmission in the CA3 region of the hippocampus revealed that loss of SV2A leads to a reduction in action potential-dependent γ-aminobutyric acid (GABA)ergic neurotransmission. In contrast, action potential-independent neurotransmission was normal. Analyses of synapse ultrastructure suggest that altered neurotransmission is not caused by changes in synapse density or morphology. These findings demonstrate that SV2A is an essential protein and implicate it in the control of exocytosis.
The
FEZ1
gene at chromosome 8p22 encodes a leucine-zipper protein, and its expression is altered in multiple human tumors
Authors: Ishii Hideshi , Baffa Raffaele , Numata Shin-Ichiro , Murakumo Yoshiki , Rattan Shashi , Inoue Hiroshi , Mori Masaki , Fidanza Vincenzo , Alder Hansjuerg , Croce Carlo M.
Published: July 2002

Alterations of human chromosome 8p occur frequently in many tumors. We identified a 1.5-Mb common region of allelic loss on 8p22 by allelotype analysis. cDNA selection allowed isolation of several genes, including
FEZ1
. The predicted Fez1 protein contained a leucine-zipper region with similarity to the DNA-binding domain of the cAMP-responsive activating-transcription factor 5. RNA blot analysis revealed that
FEZ1
gene expression was undetectable in more than 60% of epithelial tumors. Mutations were found in primary esophageal cancers and in a prostate cancer cell line. Transcript analysis from several
FEZ1
-expressing tumors revealed truncated mRNAs, including a frameshift. Alteration and inactivation of the
FEZ1
gene may play a role in various human tumors.
Dedifferentiation of adenocarcinomas by activation of phosphatidylinositol 3-kinase
Authors: Kobayashi Michimoto , Nagata Satoshi , Iwasaki Takuya , Yanagihara Kazuyoshi , Saitoh Izumu , Karouji Yutaka , Ihara Sayoko , Fukui Yasuhisa
Published: July 2002

Signet ring cell carcinoma is a malignant type of poorly differentiated adenocarcinomas in stomach, which is characterized by the occasional presence of signet ring-like cancer cells. We found that expression of constitutively active phosphatidylinositol 3-kinase (PI 3-kinase) in well differentiated adenocarcinoma cell lines induced the loss of cell–cell contact and some of the cells changed their shapes to signet ring cell-like, characterized by appearance of mucus droplets in the cytoplasm with well developed endplasmic reticulum and Golgi complexes. The active PI 3-kinase-expressing cells formed poorly differentiated tumors in nude mice, which were clearly different from those of the original cell lines. The PI 3-kinase activities detected in anti-phosphotyrosine immunoprecipitates were higher in several signet ring cell carcinoma-derived cell lines than in other adenocarcinoma cell lines. In addition, PI 3-kinase was found to be associated with a 200-kDa protein phosphorylated in tyrosine in 4 of 6 signet ring cells but not in other cell lines, suggesting that PI 3-kinase is possibly activated in these cells by binding to the 200-kDa protein. The 200-kDa protein–PI 3-kinase complex was exclusively fractionated in the membrane fractions. The specific activity of the PI 3-kinase immunoprecipitated with anti-phosphotyrosine antibody was ≈3-fold higher than that with anti-PI 3-kinase antibody. These results suggest that PI 3-kinase in signet ring cell carcinoma is recruited to the membrane and activated by the binding to the 200-kDa protein.
Selective depletion of heat shock protein 70 (Hsp70) activates a tumor-specific death program that is independent of caspases and bypasses Bcl-2
Authors: Nylandsted Jesper , Rohde Mikkel , Brand Karsten , Bastholm Lone , Elling Folmer , Jäättelä Marja
Published: July 2002

Heat shock protein 70 is an antiapoptotic chaperone protein highly expressed in human breast tumors and tumor cell lines. Here, we demonstrate that the mere inhibition of its synthesis by adenoviral transfer or classical transfection of antisense Hsp70 cDNA (asHsp70) results in massive death of human breast cancer cells (MDA-MB-468, MCF-7, BT-549, and SK-BR-3), whereas the survival of nontumorigenic breast epithelial cells (HBL-100) or fibroblasts (WI-38) is not affected. Despite the apoptotic morphology as judged by electron microscopy, the asHsp70-induced death was independent of known caspases and the p53 tumor suppressor protein. Furthermore, Bcl-2 and Bcl-X
L
, which protect tumor cells from most forms of apoptosis, failed to rescue breast cancer cells from asHsp70-induced death. These results show that tumorigenic breast cancer cells depend on the constitutive high expression of Hsp70 to suppress a transformation-associated death program. Neutralization of Hsp70 may open new possibilities for treatment of cancers that have acquired resistance to therapies activating the classical apoptosis pathway.
Evolutionary consequences of changes in species' geographical distributions driven by Milankovitch climate oscillations
Authors: Dynesius Mats , Jansson Roland
Published: July 2002

We suggest Milankovitch climate oscillations as a common cause for geographical patterns in species diversity, species' range sizes, polyploidy, and the degree of specialization and dispersability of organisms. Periodical changes in the orbit of the Earth cause climatic changes termed Milankovitch oscillations, leading to large changes in the size and location of species' geographical distributions. We name these recurrent changes “orbitally forced species' range dynamics” (ORD). The magnitude of ORD varies in space and time. ORD decreases gradual speciation (attained by gradual changes over many generations), increases range sizes and the proportions of species formed by polyploidy and other “abrupt” mechanisms, selects against specialization, and favor dispersability. Large ORD produces species prone neither to extinction nor gradual speciation. ORD increases with latitude. This produces latitudinal patterns, among them the gradient in species diversity and species' range sizes (Rapoport's rule). Differential ORD and its evolutionary consequences call for new conservation strategies on the regional to global scale.
Inactivation of catecholamines by superoxide gives new insights on the pathogenesis of septic shock
Authors: Macarthur Heather , Westfall Thomas C. , Riley Dennis P. , Misko Thomas P. , Salvemini Daniela
Published: July 2002

A major feature of septic shock is the development of a vascular crisis characterized by nonresponsiveness to sympathetic vasoconstrictor agents and the subsequent irreversible fall in blood pressure. In addition, sepsis, like other inflammatory conditions, results in a large increase in the production of free radicals, including superoxide anions (O
2⨪
) within the body. Here we show that O
2⨪
reacts with catecholamines deactivating them
in vitro
. Moreover, this deactivation would appear to account for the hyporeactivity to exogenous catecholamines observed in sepsis, because administration of a superoxide dismutase (SOD) mimetic to a rat model of septic shock to remove excess O
2⨪
restored the vasopressor responses to norepinephrine. This treatment with the SOD mimetic also reversed the hypotension in these animals; suggesting that deactivation of endogenous norepinephrine by O
2⨪
contributes significantly to this aspect of the vascular crisis. Indeed, the plasma concentrations of both norepinephrine and epinephrine in septic rats treated with the SOD mimetic were significantly higher than in untreated rats. Interestingly, the plasma concentrations for norepinephrine and epinephrine were inversely related to the plasma concentrations of adrenochromes, the product of the autoxidation of catecholamines initiated by O
2⨪
. We propose, therefore, that the use of a SOD mimetic represents a new paradigm for the treatment of septic shock. By removing O
2⨪
, exogenous and endogenous catecholamines are protected from autoxidation. As a result, both hyporeactivity and hypotension are reversed, generation of potentially toxic adrenochromes is reduced, and survival rate is improved.
Nonglassy kinetics in the folding of a simple single-domain protein
Authors: Gillespie Blake , Plaxco Kevin W.
Published: July 2002

Theory suggests that the otherwise rapid folding of simple
heteropolymer models becomes “glassy”—dominated by multiple
kinetically trapped misfolded states—at low temperatures or when the
overall bias toward the native state is reduced relative to the depth
of local minima. Experimental observations of nonsingle-exponential
protein-folding kinetics have been taken as evidence that the
protein-folding free energy landscape is similarly rough. No equivalent
analysis, however, has been reported for a simple single-domain protein
lacking prolines, disulfide bonds, prosthetic groups, or other gross
structural features that might complicate folding. In an effort to
characterize the glassiness of a folding free energy landscape in the
absence of these potentially complicating factors, we have monitored
the folding of a kinetically simple protein, peptostreptococcal protein
L (protein L). We observe no statistically significant deviation from
homogeneous single-exponential relaxation kinetics across temperatures
ranging from near the protein's melting temperature to as low as
−15°C. On the basis of these observations, we estimate that, if
there is a glass transition in the folding of protein L, it occurs at
least 45°C and possibly more than 145°C below the freezing point of
water. Apparently the folding free energy landscape of protein L is
extremely smooth, which may be indicative of a rate-limiting step in
folding that is, effectively, a nonglassy process.
A nuclear export signal in the N-terminal regulatory domain of IκBα controls cytoplasmic localization of inactive NF-κB/IκBα complexes
Authors: Huang Tony T. , Kudo Nobuaki , Yoshida Minoru , Miyamoto Shigeki
Published: July 2002

Appropriate subcellular localization is crucial for regulation of NF-κB function. Herein, we show that latent NF-κB complexes can enter and exit the nucleus in preinduction states. The nuclear export inhibitor leptomycin B (LMB) sequestered NF-κB/IκBα complexes in the nucleus. Using deletion and site-directed mutagenesis, we identified a previously uncharacterized nuclear export sequence in residues 45–54 of IκBα that was required for cytoplasmic localization of inactive complexes. This nuclear export sequence also caused nuclear exclusion of heterologous proteins in a LMB-sensitive manner. Importantly, a LMB-insensitive CRM1 mutant (Crm1-K1) abolished LMB-induced nuclear accumulation of the inactive complexes. Moreover, a cell-permeable p50 NF-κB nuclear localization signal peptide also blocked these LMB effects. These results suggest that NF-κB/IκBα complexes shuttle between the cytoplasm and nucleus by a nuclear localization signal-dependent nuclear import and a CRM1-dependent nuclear export. The LMB-induced nuclear complexes could not bind DNA and were inaccessible to signaling events, because LMB inhibited NF-κB activation without affecting the subcellular localization of upstream kinases IKKβ and NIK. Our findings indicate that the dominant nuclear export over nuclear import contributes to the largely cytoplasmic localization of the inactive complexes to achieve efficient NF-κB activation by extracellular signals.
Aberrant development of motor axons and neuromuscular synapses in erbB2-deficient mice
Authors: Lin Weichun , Sanchez Hugo B. , Deerinck Tom , Morris Jacqueline K. , Ellisman Mark , Lee Kuo-Fen
Published: July 2002

Receptor tyrosine kinase erbB2, which is activated by neuregulin, is expressed in Schwann and muscle cells in the developing neuromuscular junction (NMJ).
In vitro
studies have shown that neuregulin promotes the survival and migration of Schwann cells and stimulates acetylcholine receptor gene transcription in cultured muscle cells. These findings suggest an important role for erbB2 in the development of the NMJ. Here we examine erbB2-deficient mice to determine whether erbB2 is required for NMJ development
in vivo
. Our analysis shows that there are pre- and postsynaptic defects of developing NMJ in erbB2-deficient embryos. The presynaptic defects include defasciculation and degeneration of the motor nerves, and an absence of Schwann cells. The postsynaptic defect features an impairment of junctional folds at the neuromuscular synapse in the mutants. These results demonstrate that erbB2 is essential for
in vivo
development of the NMJ.
Nonglacial rapid climate events: Past and future
Authors: Overpeck Jonathan , Webb Robert
Published: July 2002

The paleoclimate record makes it clear that rapid climate
shifts of the 20th century are only a subset of possible climate system
behavior that might occur in the absence of glacial conditions, and
that climatic surprises could be a challenge for society even in the
absence of significant greenhouse warming.
Can ozone depletion and global warming interact to produce rapid climate change?
Authors: Hartmann Dennis L. , Wallace John M. , Limpasuvan Varavut , Thompson David W. J. , Holton James R.
Published: July 2002

The atmosphere displays modes of variability whose structures exhibit a strong longitudinally symmetric (annular) component that extends from the surface to the stratosphere in middle and high latitudes of both hemispheres. In the past 30 years, these modes have exhibited trends that seem larger than their natural background variability, and may be related to human influences on stratospheric ozone and/or atmospheric greenhouse gas concentrations. The pattern of climate trends during the past few decades is marked by rapid cooling and ozone depletion in the polar lower stratosphere of both hemispheres, coupled with an increasing strength of the wintertime westerly polar vortex and a poleward shift of the westerly wind belt at the earth's surface. Annular modes of variability are fundamentally a result of internal dynamical feedbacks within the climate system, and as such can show a large response to rather modest external forcing. The dynamics and thermodynamics of these modes are such that strong synergistic interactions between stratospheric ozone depletion and greenhouse warming are possible. These interactions may be responsible for the pronounced changes in tropospheric and stratospheric climate observed during the past few decades. If these trends continue, they could have important implications for the climate of the 21st century.
Lipotoxic heart disease in obese rats: Implications for human obesity
Authors: Zhou Yan-Ting , Grayburn Paul , Karim Asad , Shimabukuro Michio , Higa Moritake , Baetens Dany , Orci Lelio , Unger Roger H.
Published: July 2002

To determine the mechanism of the cardiac dilatation and reduced contractility of obese Zucker Diabetic Fatty rats, myocardial triacylglycerol (TG) was assayed chemically and morphologically. TG was high because of underexpression of fatty acid oxidative enzymes and their transcription factor, peroxisome proliferator-activated receptor-α. Levels of ceramide, a mediator of apoptosis, were 2–3 times those of controls and inducible nitric oxide synthase levels were 4 times greater than normal. Myocardial DNA laddering, an index of apoptosis, reached 20 times the normal level. Troglitazone therapy lowered myocardial TG and ceramide and completely prevented DNA laddering and loss of cardiac function. In this paper, we conclude that cardiac dysfunction in obesity is caused by lipoapoptosis and is prevented by reducing cardiac lipids.
Conversion of α-lactalbumin to a protein inducing apoptosis
Authors: Svensson M. , Håkansson A. , Mossberg A.-K. , Linse S. , Svanborg C.
Published: July 2002

In this study α-lactalbumin was converted from the regular, native state to a folding variant with altered biological function. The folding variant was shown to induce apoptosis in tumor cells and immature cells, but healthy cells were resistant to this effect. Conversion to HAMLET (human α-lactalbumin made lethal to tumor cells) required partial unfolding of the protein and a specific fatty acid, C18:1, as a necessary cofactor. Conversion was achieved with α-lactalbumin derived from human milk whey and with recombinant protein expressed in
Escherichia coli
. We thus have identified the folding change and the fatty acid as two key elements that define HAMLET, the apoptosis-inducing functional state of α-lactalbumin. Although the environment in the mammary gland favors the native conformation of α-lactalbumin that serves as a specifier in the lactose synthase complex, the conditions under which HAMLET was formed resemble those in the stomach of the nursing child. Low pH is known to release Ca
2+
from the high-affinity Ca
2+
-binding site and to activate lipases that hydrolyze free fatty acids from milk triglycerides. We propose that this single amino acid polypeptide chain may perform vastly different biological functions depending on its folding state and the
in vivo
environment. It may be speculated that molecules like HAMLET can aid in lowering the incidence of cancer in breast-fed children by purging of tumor cells from the gut of the neonate.
Cellular and molecular responses of the uterus to embryo implantation can be elicited by locally applied growth factors
Authors: Paria Bibhash C. , Ma Wen-ge , Tan Jian , Raja Shefali , Das Sonjoy K. , Dey Sudhansu K. , Hogan Brigid L. M.
Published: July 2002

The implantation of a blastocyst into a receptive uterus is
associated with a series of events, namely the attachment reaction
followed by decidualization of the stroma. Previous studies established
that the gene encoding heparin-binding EGF-like growth factor (HB-EGF)
is expressed in the luminal epithelium solely at the site of blastocyst
apposition preceding the attachment reaction. We report here the
expression during implantation of 21 genes encoding other signaling
proteins, including those belonging to the Bone morphogenetic protein
(BMP), fibroblast growth factor (FGF), WNT, and Hedgehog (HH) pathways.
We find that the attachment reaction is associated with a localized
stromal induction of genes encoding BMP-2, FGF-2, and WNT-4. Despite
efforts by many investigators, a simple
in vitro
model
of implantation is not yet available to study either the hierarchy of
the events triggered in the uterus by the embryo or the function of
individual signaling proteins. We have therefore approached these
questions by introducing beads loaded with purified factors into the
receptive uterus. We show that beads soaked in HB-EGF or insulin-like
growth factor-1 (IGF-1), but not other proteins, induce many of the
same discrete local responses elicited by the blastocyst, including
increased localized vascular permeability, decidualization, and
expression of
Bmp2
at the sites of the beads. By
contrast, the expression domains of
Indian hedgehog
(
Ihh
),
patched
, and
noggin
become restricted as decidualization proceeds. Significantly, beads
containing BMP-2 do not themselves elicit an implantation response but
affect the spacing of implantation sites induced by blastocysts
cotransferred with the beads.
High-resolution minisatellite-based typing as a portable approach to global analysis ofMycobacterium tuberculosismolecular epidemiology
Authors: Mazars Edith , Lesjean Sarah , Banuls Anne-Laure , Gilbert Michèle , Vincent Véronique , Gicquel Brigitte , Tibayrenc Michel , Locht Camille , Supply Philip
Published: July 2002

The worldwide threat of tuberculosis to human health emphasizes the need to develop novel approaches to a global epidemiological surveillance. The current standard forMycobacterium tuberculosistyping based on IS6110restriction fragment length polymorphism (RFLP) suffers from the difficulty of comparing data between independent laboratories. Here, we propose a high-resolution typing method based on variable number tandem repeats (VNTRs) of genetic elements named mycobacterial interspersed repetitive units (MIRUs) in 12 human minisatellite-like regions of theM. tuberculosisgenome. MIRU-VNTR profiles of 72 differentM. tuberculosisisolates were established by PCR analysis of all 12 loci. From 2 to 8 MIRU-VNTR alleles were identified in the 12 regions in these strains, which corresponds to a potential of over 16 million different combinations, yielding a resolution power close to that of IS6110-RFLP. All epidemiologically related isolates tested were perfectly clustered by MIRU-VNTR typing, indicating that the stability of these MIRU-VNTRs is adequate to track outbreak episodes. The correlation between genetic relationships inferred from MIRU-VNTR and IS6110-RFLP typing was highly significant. Compared with IS6110-RFLP, high-resolution MIRU-VNTR typing has the considerable advantages of being fast, appropriate for allM. tuberculosisisolates, including strains that have a few IS6110copies, and permitting easy and rapid comparison of results from independent laboratories. This typing method opens the way to the construction of digital global databases for molecular epidemiology studies ofM. tuberculosis.
The myth of the best argument: power, deliberation and reason1
Authors: Pellizzoni Luigi
Published: July 2002

Power in communication takes two main forms. As ‘external’ power, it consists in the ability to acknowledge or disregard a speaker or a discourse. As ‘internal’ power, it is the ability of an argument to eliminate other arguments by demonstrating its superiority. A positive or negative value may be ascribed to these forms of power. Four ideal‐typical positions are discussed – strategy, technocracy, constructionism, and deliberation.

Public deliberation has three virtues – civic virtue, governance virtue and cognitive virtue. Deliberation lowers the propensity to, and the benefit of, strategic behaviour. It also increases knowledge, enhancing the quality of decisions.

For Habermas, the unity of reason is expressed in the possibility of agreement on the most convincing argument. However, sometimes conflicts are deeplying, principles and factual descriptions are profoundly different, and uncertainty is radical. The best argument cannot be found. There is no universal reason. The question is whether non‐strategic agreement may spring from the incommensurability of languages.

In search of an answer, Rawls's concept of overlapping consensus, the feminist theory of the public sphere, and the idea of deliberation as co‐operation are discussed. The argument developed is that the approach to deliberative democracy may be renewed by rethinking its motivational and cognitive elements. Public deliberation is grounded on a pre‐political level of co‐operation. Intractable controversies may be faced at the level of practices, looking for local, contextual answers.
Suicidal ideation, parenting style, and family climate among Hong Kong adolescents
Authors: Lai Ka Wai , McBride‐Chang Catherine
Published: July 2002

The prevalence of suicidal ideation and its relations with perceived parenting treatment and family climate was examined in 120 Hong Kong students aged 15–19 years. Fifty‐two per cent of the participants reported suicide ideation. Suicide ideation was found to be significantly associated with perceived authoritarian parenting, low parental warmth, high maternal over‐control, negative child‐rearing practices, and a negative family climate. A positive family climate may act as a buffer against developing suicide ideation in adolescents.
Culture and Gender Differences in the Perception of Friendship by Adolescents
Authors: Verkuyten Maykel , Masson Kees
Published: July 2002

This study examines personal collectivism and individualism (or allocentrism and idiocentrism) in relation to the perception of same‐sex friendships among adolescents living in a multi‐ethnic context in the Netherlands. Respondents originally from collectivist cultures were more allocentric than respondents originally from individualist cultures. Among the former group allocentrism was unrelated to idiocentrism, whereas a negative relation was found among the latter group. Allocentrism was related to a greater sensitivity to friends, using more ascribed features in describing friends, having fewer friends but seeing their relationship as closer, perceiving less intimacy with other‐than‐best‐friends, and endorsing rules about relations with third parties more. Idiocentrism was related to less sensitivity to friends, using more personal characteristics in describing friends, but also to having fewer friends, talking less intimately with others, and endorsing friendship rules about intimacy less. Additionally, gender had independent effects on the perception of friendship, suggesting that cultural and gender differences cannot be characterized by the same set of features.