bluuebunny/crossref_metadata_2025 huggingface dataset:
Activation of NF-κB via a Src-dependent Ras-MAPK-pp90rsk pathway is required for
Pseudomonas aeruginosa
-induced mucin overproduction in epithelial cells
Authors: Li Jian-Dong , Feng Weijun , Gallup Marianne , Kim Jae-Ho , Gum James , Kim Young , Basbaum Carol
Published: July 2002

Cystic fibrosis (CF) is an autosomal recessive disorder, the most common lethal genetic disease in Caucasians. Respiratory disease is the major cause of morbidity and mortality. Indeed, 95% of CF patients die of respiratory failure.
Pseudomonas aeruginosa
, an opportunistic pathogen, chronically infects the lungs of over 85% of CF patients. It is ineradicable by antibiotics and responsible for airway mucus overproduction that contributes to airway obstruction and death. The molecular mechanisms underlying this pathology are unknown. Here we show that
P. aeruginosa
activates a c-Src-Ras-MEK1/2-MAPK-pp90rsk signaling pathway that leads to activation of nuclear factor NF-κB (p65/p50). Activated NF-κB binds to a κB site in the 5′-flanking region of the
MUC2
gene and activates
MUC2
mucin transcription. These studies bring new insight into bacterial–epithelial interactions and more specifically into the molecular pathogenesis of cystic fibrosis. Understanding these signaling and gene regulatory mechanisms opens up new therapeutic targets for cystic fibrosis.
Tobacco mosaic virus infection induces severe morphological changes of the endoplasmic reticulum
Authors: Reichel Christoph , Beachy Roger N.
Published: July 2002

The tobacco mosaic virus (TMV) movement protein (MP) facilitates transport of virus infection between adjacent cells by modifying plasmodesmata. Previous studies suggested that the cytoskeleton and the endomembrane system are involved in this transport. We examined the effects of TMV infection on the endoplasmic reticulum (ER) in transgenic
Nicotiana benthamiana
that accumulate the green fluorescent protein (GFP) in the ER. Fluorescence microscopy was used to show that early in infection the ER undergoes dramatic morphological changes that include the conversion of tubular ER into large aggregates that revert to tubular ER in later stages of infection. These changes parallel MP accumulation and degradation. Furthermore, a fusion protein comprising MP fused to GFP accumulates in or on these large aggregates of ER. Expression of MP-GFP in the absence of virus infection led to the production of fluorescent aggregates of the same apparent form and size. Microsomes isolated from infected leaves contain MP. We show that the MP appears to behave as an integral ER membrane protein and is exposed on the cytosolic face of the ER. The importance of the association of MP with ER and its possible role in intracellular and intercellular spread of infection is discussed.
Evidence that the nucleotide exchange and hydrolysis cycle of G proteins causes acute desensitization of G-protein gated inward rectifier K
+
channels
Authors: Chuang Huai-hu , Yu Mei , Jan Yuh Nung , Jan Lily Yeh
Published: July 2002

The G-protein gated inward rectifier K
+
channel (GIRK) is activated
in vivo
by the Gβγ subunits liberated upon G
i
-coupled receptor activation. We have recapitulated the acute desensitization of receptor-activated GIRK currents in heterologous systems and shown that it is a membrane-delimited process. Its kinetics depends on the guanine nucleotide species available and could be accounted for by the nucleotide exchange and hydrolysis cycle of G proteins. Indeed, acute desensitization is abolished by nonhydrolyzable GTP analogues. Whereas regulators of G-protein signaling (RGS) proteins by their GTPase-activating protein activities are regarded as negative regulators, a positive regulatory function of RGS4 is uncovered in our study; the opposing effects allow RGS4 to potentiate acute desensitization without compromising GIRK activation.
Is the molten globule a third phase of proteins?
Authors: Pande Vijay S. , Rokhsar Daniel S.
Published: July 2002

The equilibrium properties of proteins are studied by Monte Carlo simulation of two simplified models of protein-like heteropolymers. These models emphasize the polymeric entropy of the fluctuating polypeptide chain. Our calculations suggest a generic phase diagram that contains a thermodynamically distinct “molten globule” state in addition to a rigid native state and a nontrivial unfolded state. The roles of side-chain packing and loop entropy are discussed.
Hypoalgesia in mice with a targeted deletion of the tachykinin 1 gene
Authors: Zimmer Andreas , Zimmer Anne M. , Baffi Judith , Usdin Ted , Reynolds Kay , König Monika , Palkovits Miklós , Mezey Éva
Published: July 2002

The tachykinin neuropeptides, substance P and substance K, are produced in nociceptive primary sensory neurons and in many brain regions involved in pain signaling. However, the precise role and importance of these neuropeptides in pain responses has been debated. We now show that mice that cannot produce these peptides display no significant pain responses following formalin injection and have an increased pain threshold in the hotplate test. On the other hand, the mutant mice react normally in the tail flick assay and acetic acid-induced writhing tests. These results demonstrate that substance P and/or substance K have essential functions in specific responses to pain.
A read-ahead function in archaeal DNA polymerases detects promutagenic template-strand uracil
Authors: Greagg Martin A. , Fogg Mark J. , Panayotou George , Evans Steven J. , Connolly Bernard A. , Pearl Laurence H.
Published: July 2002

Deamination of cytosine to uracil is the most common promutagenic change in DNA, and it is greatly increased at the elevated growth temperatures of hyperthermophilic archaea. If not repaired to cytosine prior to replication, uracil in a template strand directs incorporation of adenine, generating a G⋅C → A⋅U transition mutation in half the progeny. Surprisingly, genomic analysis of archaea has so far failed to reveal any homologues of either of the known families of uracil-DNA glycosylases responsible for initiating the base-excision repair of uracil in DNA, which is otherwise universal. Here we show that DNA polymerases from several hyperthermophilic archaea (including Vent and
Pfu
) specifically recognize the presence of uracil in a template strand and stall DNA synthesis before mutagenic misincorporation of adenine. A specific template-checking function in a DNA polymerase has not been observed previously, and it may represent the first step in a pathway for the repair of cytosine deamination in archaea.
Reverse homeosis in homeotically reconstructed ribbonworms
Authors: Tarpin Michel , Gehring Walter J. , Bièrne Jacques
Published: July 2002

Homeosis is the replacement of one body part by another, which may be caused by either developmental or genetic variations. It is particularly obvious in segmented animals, like insects, in which one body segment may be transformed into another. However, homeosis also occurs in animals without overt segmentation that also have detailed positional information specifying their body plan. By grafting, we have artificially generated homeotic ribbonworms of the species
Lineus ruber
with a duplicated ocellar region replacing the postocellar region anterior to the brain. Such chimeric animals are capable of complete morphogenetic regulation of the anterior–posterior (A–P) pattern. The missing postocellar region is restored by intercalary regeneration, and the anterior duplicated ocellar region is eliminated by a process called transgeneration. Thus, homeosis is reversed, and a completely normal pattern along the A–P axis is restored. This reverse homeosis involves the elimination of the syngeneic eyes and the survival of the grafted allogeneic eye region.
LsPax-6
, the
Lineus sanguineus
ortholog of the mammalian
Pax-6
gene, which is considered to be a master control gene for eye morphogenesis, is expressed specifically in regenerating, regenerated, and intact eye regions. Our data show that ribbonworm eyes are either maintained or they regress according to their position along the A–P axis, even though there are no obvious segmental boundaries. This system allows us to test the function of LsPax-6 protein not only during eye regeneration but also during maintenance and regression of the eyes.
Gβ5 prevents the RGS7-Gαo interaction through binding to a distinct Gγ-like domain found in RGS7 and other RGS proteins
Authors: Levay Konstantin , Cabrera Jorge L. , Satpaev Daulet K. , Slepak Vladlen Z.
Published: July 2002

The G protein β subunit Gβ5 deviates significantly from the other four members of Gβ-subunit family in amino acid sequence and subcellular localization. To detect the protein targets of Gβ5
in vivo
, we have isolated a native Gβ5 protein complex from the retinal cytosolic fraction and identified the protein tightly associated with Gβ5 as the regulator of G protein signaling (RGS) protein, RGS7. Here we show that complexes of Gβ5 with RGS proteins can be formed
in vitro
from the recombinant proteins. The reconstituted Gβ5-RGS dimers are similar to the native retinal complex in their behavior on gel-filtration and cation-exchange chromatographies and can be immunoprecipitated with either anti-Gβ5 or anti-RGS7 antibodies. The specific Gβ5-RGS7 interaction is determined by a distinct domain in RGS that has a striking homology to Gγ subunits. Deletion of this domain prevents the RGS7-Gβ5 binding, although the interaction with Gα is retained. Substitution of the Gγ-like domain of RGS7 with a portion of Gγ1 changes its binding specificity from Gβ5 to Gβ1. The interaction of Gβ5 with RGS7 blocked the binding of RGS7 to the Gα subunit Gαo, indicating that Gβ5 is a specific RGS inhibitor.
Multiple endocytic pathways of G protein-coupled receptors delineated by GIT1 sensitivity
Authors: Claing Audrey , Perry Stephen J. , Achiriloaie Mircea , Walker Julia K. L. , Albanesi Joseph P. , Lefkowitz Robert J. , Premont Richard T.
Published: July 2002

Recently, we identified a GTPase-activating protein for the ADP ribosylation factor family of small GTP-binding proteins that we call GIT1. This protein initially was identified as an interacting partner for the G protein-coupled receptor kinases, and its overexpression was found to affect signaling and internalization of the prototypical β
2
-adrenergic receptor. Here, we report that GIT1 overexpression regulates internalization of numerous, but not all, G protein-coupled receptors. The specificity of the GIT1 effect is not related to the type of G protein to which a receptor couples, but rather to the endocytic route it uses. GIT1 only affects the function of G protein-coupled receptors that are internalized through the clathrin-coated pit pathway in a β-arrestin- and dynamin-sensitive manner. Furthermore, the GIT1 effect is not limited to G protein-coupled receptors because overexpression of this protein also affects internalization of the epidermal growth factor receptor. However, constitutive agonist-independent internalization is not regulated by GIT1, because transferrin uptake is not affected by GIT1 overexpression. Thus, GIT1 is a protein involved in regulating the function of signaling receptors internalized through the clathrin pathway and can be used as a diagnostic tool for defining the endocytic pathway of a receptor.
Rhodopsin kinase: Two mAbs binding near the carboxyl terminus cause time-dependent inactivation
Authors: Bruel Christophe , Cha Kiweon , Niu Li , Reeves Philip J. , Khorana H. Gobind
Published: July 2002

Two mAbs generated against rhodopsin kinase (RK) were characterized
for their epitopes. Both antibodies recognize short peptide sequences,
overlapping but distinct, close to the carboxyl terminus. Binding of RK
to the antibodies is slow. Attempts were made to use the antibodies
immobilized on protein A-Sepharose beads to bind and purify the enzyme.
Time-dependent inactivation of the enzyme occurred after its binding to
the antibodies. Studies using different conditions to maintain the
enzyme in the active form during binding or to reactivate the purified
inactivated enzyme were unsuccessful.
Analysis of a complete homeobox gene repertoire: Implications for the evolution of diversity
Authors: Kappen Claudia
Published: July 2002

The completion of sequencing projects for various organisms has already advanced our insight into the evolution of entire genomes and the role of gene duplications. One multigene family that has served as a paradigm for the study of gene duplications and molecular evolution is the family of homeodomain-encoding genes. I present here an analysis of the homeodomain repertoire of an entire genome, that of the nematode
Caenorhabditis elegans
, in relation to our current knowledge of these genes in plants, arthropods, and mammals. A methodological framework is developed that proposes approaches for the analysis of homeodomain repertoires and multigene families in general.
The development of contemporary Spanish psychology
Authors: Carpintero Helio
Published: July 2002

Spanish psychology has developed greatly in the last decades, both in academic and professional aspects. But its history during the 20th century includes not only positive periods but also others clearly regressive, which impeded scientific work and tried to return to a more philosophical orientation. Since the establishment of a psychology degree in the country, a great expansion has taken place, and other significant achievements have been made by some émigré Spanish psychologists in Latin‐American countries after being reinstalled there after a civil war that strongly affected the evolution of contemporary Spain. The main lines of the picture are drawn here.
Psychology in an Asian country: A report from Sri Lanka
Authors: de Zoysa Piyanjali , Ismail Chandrika
Published: July 2002

This paper describes the history and present status of psychology in Sri Lanka, including facilities for teaching psychology and legal issues in registration provisions for psychologists. The future directions in improving the status of psychology in the country are delineated.
Resource Allocation by Greek and Dutch Students: A Test of Three Models
Authors: van den Heuvel Koen , Poortinga Ype H.
Published: July 2002

Three models are presented that explain cross‐cultural differences in the allocation of resources to various social categories. (1) Individualism‐collectivism predicts, for all kinds of resources, a higher allocation in collectivistic than in individualistic societies to members of one's in group. (2) Kagitçibasi's model of family change makes a distinction between material and nonmaterial resources. Collectivistic value patterns should lead to higher allocation of nonmaterial resources than found in individualistic societies, but no such difference is expected for material investments. Both these models assume cross‐cultural differences in value orientations at a high level of generality. (3) An alternative is proposed, predicting universal patterns of exchange with only context‐specific cross‐cultural variations at a low level of generality. Hypotheses derived from the three models are tested with data from Greece and The Netherlands. It is concluded that the results are most compatible with the third model.
International and Domestic Adoption: Child, Parents, and Family Adjustment
Authors: Levy-Shiff Rachel , Zoran Naomi , Shulman Shmuel
Published: July 2002

Adjustment of school-aged children, parents, and families in international versus domestic adoption was studied in 100 Israeli families. No significant differences were found between the two groups of children in school adjustment, grades, IQ level, psychological adjustment (i.e. anxiety, depression, hyperactivity, self-concept), observed behaviours at home, and coping with adoption issues. However, they perceived their parents to be more supportive but controlling. Marked differences were found in parents’ behaviours, especially fathers’, and family functioning: In the international adoption group, compared with the domestic adoption group, parents used more problem-focused and support-seeking ways of coping, viewed parenting more as a challenge, were more involved with their children; but were more intrusive, reported more cohesive family relations and better marital adjustment, and coped differently with adoption issues. Results are discussed in terms of the differences in the two types of adoption.
Naltrexone in the treatment of heroin dependence: a literature review
Authors: TUCKER THAMIZAN K , RITTER ALISON J.
Published: July 2002

Naltrexone is an opioid antagonist which binds competitively at the opioid receptors and blocks the effects of exogenous opioids. Naltrexone has been used to facilitate withdrawal (in using rapid and ultra rapid withdrawal techniques). The research on the efficacy of rapid and ultra‐rapid withdrawal to date is extremely limited. The predominant use of naltrexone is as a post‐withdrawal relapse prevention intervention. Research has revealed that naltrexone's acceptability is generally poor, mainly appealing to those highly motivated towards achieving and maintaining abstinence. In spite of poor acceptibility, naltrexone‐treated participants often produce lower levels of craving for opiates, significantly higher rates and longer periods of abstinence and significantly greater improvement in psychosocial functioning than those treated with placebos and standard treatment programmes. The studies of highly motivated participants demonstrated the highest retention and completion rates. Being employed and having good social support have been identified as being associated with successful outcome. There also appears to be a trend towards greater success being associated with older participants, and those with a greater number of previous treatment attempts. The combined use of naltrexone and psychosocial therapy has proved to be more effective than psychosocial therapy alone, and more effective than naltrexone treatment alone, in improving post treatment outcome. Future research directions are summarized.
Cannabis policy and the burden of proof: is it now beyond reasonable doubt that cannabis prohibition is not working?
Authors: LENTON SIMON
Published: July 2002

‘Innocent until proven guilty’ is how many policy makers have thought about the prohibition of cannabis in Western Australia and elsewhere. Comparisons between so called ‘decriminalized’ and prohibitionist states show decriminalization has not led to higher rates of current cannabis use. Under prohibition significant numbers of Western Australian citizens receive a criminal record for no more serious offence than the possession of a small amount of cannabis for personal use. Recent research has shown that such a conviction can have a real and detrimental impact on people's lives, reinforces disrespect for the cannabis laws, but appears not to deter cannabis use among those so convicted. This paper suggests that it is now beyond reasonable doubt that applying criminal sanctions for minor cannabis offences is not in the best interests of the community and recommends an alternative.
Drug and alcohol problems: the users' perspective on family members' coping
Authors: KRISHNAN MYA , ORFORD JIM , BRADBURY COLIN , COPELLO ALEX , VELLEMAN RICHARD
Published: July 2002

The aim of the study was to explore problem drug and alcohol users' perceptions of family members' coping in a cross‐sectional interview study of problem drug and alcohol users, using a long semi‐structured interview. Participants comprised nine problem drinkers, five problem drug users and one problem drug and alcohol user (two‐thirds male). Detailed qualitative analysis of interview reports suggested that drug and alcohol users have clear ideas regarding family members' coping. Coping strategies which encompass clear elements of concern for them appear to be well‐received, in particular supportive coping and, in some cases, assertive coping. Controlling coping may be viewed positively by drug and alcohol users provided it is used in conjunction with supportive coping, thereby indicating concern. Furthermore, results suggested that users found all other forms of coping such as emotional, tolerant, inactive and avoidance to be unsupportive, the only exception being tolerant coping, which was found to be supportive when drug and alcohol users had no intention of altering their use. Drug and alcohol users' perceptions of family members' coping may be viewed in terms of the stress‐coping model. Drug and alcohol users appreciate supportive coping actions, while coping strategies which involve family members withdrawing from them, making them feel guilty or forcing them to take actions against their will are felt to be unsupportive. The findings have implications for the way in which family members are advised to cope, not only in terms of their own physical and psychological well‐being, but also in terms of how their actions are perceived by the users themselves.
Subchronic Toxicity of 1,3,5-Trinitrobenzene in Fischer 344 Rats
Authors: Reddy Tirumuru V. , Olson Greg R. , Wiechman Barry , Reddy Gunda , Torsella Joni , Daniel F. Bernard
Published: July 2002

The subchronic toxicity of 1,3,5-trinitrobenzene (TNB) in male and female Fischer 344 rats was evaluated by feeding a powdered certified laboratory diet containing 0, 66.7, 400 and 800 mg TNBl kg diet for 90 days. The calculated average TNB intake was 4.29, 24.70, and 49.28 mg/kg body weight (BW)day for females and 3.91, 22.73, and 44.16 mg/ kg BWI day for males. Food intake in the 400 and 800 mg/kg diet dose groups of both sexes was decreased throughout the study and resulted in a significant decrease in absolute body weights. A significant decrease in relative testicular weights and a significant increase in the relative liver weight were observed in male rats receiving 400 or 800 mg TNB/kg diet. A significant increase in the relative spleen weights of both sexes receiving 400 or 800 mg TNB diet was noted. The relative liver weight was also increased only in female rats maintained on the 800 mg TNB diet. Histopathological examinations revealed that the susceptible organs for TNB toxicity were kidney (hyaline droplets) in all male dose groups and testes (seminiferous tubular degeneration) in rats receiving 400 and 800 mg TNB diet groups. The spleen was also affected (extramedullary hematopoiesis) in both sexes in the 400 and 800 mg dose groups. Hematological studies at both 45 (data not given) and 90 days in the 400 and 800 mg dose groups indicated decreased values for red blood cell counts and hemoglobin content, while reticulocytes and methemoglobin levels were increased. Clinical chemistry parameters were unaffected. Based on kidney toxicity and hematological effects, a Low Observed Adverse Effect Level (LOAEL) of 3.91 mg/kg BWI day was suggested for subchronic toxicity studies on TNB.
Effects of volatile inhalants on sensorimotor reactivity in rats
Authors: Wiley Jenny L. , Bowen Scott E. , Balster Robert L.
Published: July 2002

Abusers of inhaled solvents may show impaired cognitive functioning as reflected in problems with short‐term memory and attention. In the present study, we examined the effects of the abused solvents toluene, m‐xylene and 1,1,1‐trichloroethane in an acoustic startle paradigm which included tests for pre‐pulse inhibition (PPI). This procedure has been used to model sensorimotor reactivity and gating, pre‐attentional processes that may be disrupted by mental illness, substance abuse or drug withdrawal. Following exposure to an inhalant vapor or to air, rats were placed in startle chambers in which they were exposed to loud: [120 dB] acoustic pulses presented alone or preceded by a less intense pre‐pulse: [85 dB]. Results were compared to those obtained with the depressants ethanol, pentobarbital and methoxyflurane, which have many acute behavioral effects in common with those of abused solvents. As expected, these depressants only decreased startle amplitudes during pulse alone trials and had no effect on PPI. Although solvent vapors also had no effect on PPI, they did not decrease startle amplitudes in pulse alone trials, at least at the concentrations tested. In fact, toluene robustly increased startle amplitude. The lack of effect of the abused solvents on PPI does not provide support for an hypothesis that acute effects of vapors on attention and early information processing is a basis for cognitive deficits in solvent abusers.
Effects of glutamine and ethanol in vitro on lymphocytes from human alcohol abusers and non‐abusers
Authors: Greig J. E. , Keast D. , Palmer T. N.
Published: July 2002

The amino acid L‐glutamine is essential for the function of both resting and stimulated lymphocytes, and its uptake is significantly increased after stimulation. The effects of ethanol on the immune system appear to be widespread and significant, and its role as an osmolyte has recently been proposed to be important. The in vitro effect of concurrent glutamine deprivation and ethanol exposure in different osmotic conditions was investigated with respect to the proliferative capability of lymphocytes in vitro from both alcohol abusers and non‐abusers, and also the kinetics of glutamine transport across peripheral blood lymphocytes (PBL) membranes from non‐abusers. In a physiologically normal situation ethanol increases osmolality, and tonicity remains balanced. However, in situations of electrolyte imbalance ethanol addition may result in osmolality remaining near normal levels, but with hypotonicity. It was found that a lack of glutamine and iso‐osmotic hypotonic high ethanol concentrations caused a greater inhibition of mitogen‐stimulated lymphocyte proliferation than either factor alone. This was true for PBL from both non‐abusers and abusers of alcohol. There was no change in membrane permeability and active transport of glutamine into lymphocytes from non‐abusers that were stimulated in vitro in various conditions of ethanol, osmolality and tonicity. While lymphocyte proliferation may be inhibited in certain conditions and this is further affected by a lack of glutamine, the capacity for active glutamine transport was maintained, perhaps due to its essential role in lymphocyte metabolism.
Hepatitis in alcohol and drug misusers: practical issues
Authors: CRAMP MATTHEW E. , WILLIAMS ROGER
Published: July 2002

Viral hepatitis, particularly that due to HBV or HCV, is a significant problem in the UK population of IVDUs and alcohol misusers. Patients attending drug rehabilitation clinics should be screened for HBV and HCV infection. All those found to be HBcAb negative should be vaccinated against HBV. Those found to be HBsAg or HCV antibody positive should have their liver function tests (LFTs) checked and be referred to a specialist liver clinic for further evaluation. Interferon treatment of HBV or HCV infection is effective in a proportion of cases but it is an expensive agent that requires careful monitoring. Current management regimes are centred around the modification of risk behaviour and the identification of those most in need of, and most likely to respond to, anti‐viral treatment. Several new anti‐viral agents are currently undergoing clinical evaluation and this together with the prospect of vaccines designed to clear an established infection means that the next few years are likely to see some major advances in the treatment of chronic viral hepatitis.
Carbohydrate‐deficient transferrin in the assessment of harmful alcohol consumption: diagnostic performance and clinical significance
Authors: BEAN PAMELA
Published: July 2002

The last decade saw the emergence of carbohydrate‐deficient transferrin (CDT) as the most promising marker for the diagnosis of alcohol abuse. Daily alcohol consumption of four beers, four glasses of wine or three standard drinks causes increased concentrations of CDT in serum. CDT is serum transferrin with a reduced content of oligosaccharides due to the detrimental effects of alcohol metabolism on the glycosylation pathway of hepatocytes and/or the increased activity of circulating glycosidases in serum. Most current CDT procedures entail separation of normal transferrin from CDT in a charged matrix‐like isoelectric focusing and ion exchange chromatography, followed by detection/quantitation of CDT by a myriad of immunoassays: immunoblotting, radioimmunoassay, enzyme immunoassay and nephelometry. New CDT procedures present the advantages of improved performance, inexpensive automation and CDT results expressed as a percentage of total serum transferrin. CDT's major asset is its high specificity in well‐defined populations i.e. individuals ingesting 60 g alcohol daily for at least a week.The sensitivity rates, which vary between 22% and 81%, depend on the amount of alcohol ingested, time of sample collection after the cessation of drinking, age, gender and the cut‐off point chosen for analysis of tests' results. Regarding clinical applications, best outcome is achieved when the test is used to confirm a suspicion of alcohol abuse and when monitoring abstinence and relapses.The low prevalence of alcohol abuse in the general population challenges its use as a screening test.With the advent of inexpensive automation and the constant emergence of innovative, improved tests, we are seeing the rise of a new era in alcohol abuse diagnosis as affordability and education allows widespread use of CDT in a variety of settings.
The Calpain Small Subunit Gene Is Essential: Its Inactivation Results in Embryonic Lethality
Authors: Zimmerman Un‐Jin P. , Boring Landin , Pak UJhang Ho , Mukerjee Neeta , Wang Kevin K. W.
Published: July 2002

Creation of transgenic (knockout) mice deficient in calpain small (30 kDa) subunit gene was undertaken to clarify the proposed role of the small subunit for calpain proteolytic activity and to gain insight into the importance of the gene in the whole animal. The gene was targeted and disrupted in embryonic stem cells by homologous recombination, and chimeric mice were generated. Heterozygous F1 generation mice were crossed to obtain F2 generation. Among F2 generation mice, we found only wild‐type and heterozygous animals in the 80 pups genotyped to date; no homozygous mice have been found, although 20 were expected. The heterozygotes had no apparent phenotypic abnormalities. Analysis of their tissues revealed no significant difference in mRNA expression, protein content, or proteolytic activity in comparison with their wild‐type littermates. Genotyping of fetuses at different stages of development also revealed only wild‐type and normal heterozygous fetuses. No moribund embryos or resorption sites were observed in the uterine cavity. The results indicate that at least one normal allele is essential for postnatal survival. Disruption of both alleles appears to be lethal in very early fetal development.
As Others See Us: Accomplishments Over 25 Years Reflect the Symbiotic Link Between Biology and Chemistry
Authors: Berg Jeremy M.
Published: July 2002

Under the title “Biochemistry, Revised Edition,” this article was published in the March 26, 2001 issue of Chemical and Engineering News, p. 130, celebrating the 125th anniversary of the American Chemical Society. Written from the viewpoint of a chemist, it looks to how “many of the most intriguing biological problems can be posed in chemical terms.” It is reproduced here by permission of the author and publisher.
Mitotic Spindle Positioning in Saccharomyces cerevisiae Is Accomplished by Antagonistically Acting Microtubule Motor Proteins
Authors: Cottingham Frank R. , Hoyt M. Andrew
Published: July 2002

Proper positioning of the mitotic spindle is often essential for cell division and differentiation processes. The asymmetric cell division characteristic of budding yeast, Saccharomyces cerevisiae, requires that the spindle be positioned at the mother–bud neck and oriented along the mother–bud axis. The single dynein motor encoded by the S. cerevisiae genome performs an important but nonessential spindle-positioning role. We demonstrate that kinesin-related Kip3p makes a major contribution to spindle positioning in the absence of dynein. The elimination of Kip3p function in dyn1Δ cells severely compromised spindle movement to the mother–bud neck. In dyn1Δ cells that had completed positioning, elimination of Kip3p function caused spindles to mislocalize to distal positions in mother cell bodies. We also demonstrate that the spindle-positioning defects exhibited by dyn1 kip3 cells are caused, to a large extent, by the actions of kinesin- related Kip2p. Microtubules in kip2Δ cells were shorter and more sensitive to benomyl than wild-type, in contrast to the longer and benomyl-resistant microtubules found in dyn1Δ and kip3Δ cells. Most significantly, the deletion of KIP2 greatly suppressed the spindle localization defect and slow growth exhibited by dyn1 kip3 cells. Likewise, induced expression of KIP2 caused spindles to mislocalize in cells deficient for dynein and Kip3p. Our findings indicate that Kip2p participates in normal spindle positioning but antagonizes a positioning mechanism acting in dyn1 kip3 cells. The observation that deletion of KIP2 could also suppress the inviability of dyn1Δ kar3Δ cells suggests that kinesin-related Kar3p also contributes to spindle positioning.
Cadherins Promote Skeletal Muscle Differentiation in Three-dimensional Cultures
Authors: Redfield Ann , Nieman Marvin T. , Knudsen Karen A.
Published: July 2002

The cell–cell adhesion molecule N-cadherin, with its associated catenins, is expressed by differentiating skeletal muscle and its precursors. Although N-cadherin's role in later events of skeletal myogenesis such as adhesion during myoblast fusion is well established, less is known about its role in earlier events such as commitment and differentiation. Using an in vitro model system, we have determined that N-cadherin– mediated adhesion enhances skeletal muscle differentiation in three-dimensional cell aggregates. We transfected the cadherin-negative BHK fibroblastlike cell line with N-cadherin. Expression of exogenous N-cadherin upregulated endogenous β-catenin and induced strong cell–cell adhesion. When BHK cells were cultured as three-dimensional aggregates, N-cadherin enhanced withdrawal from the cell cycle and stimulated differentiation into skeletal muscle as measured by increased expression of sarcomeric myosin and the 12/101 antigen. In contrast, N-cadherin did not stimulate differentiation of BHK cells in monolayer cultures. The effect of N-cadherin was not unique since E-cadherin also increased the level of sarcomeric myosin in BHK aggregates. However, a nonfunctional mutant N-cadherin that increased the level of β-catenin failed to promote skeletal muscle differentiation suggesting an adhesion-competent cadherin is required. Our results suggest that cadherin-mediated cell–cell interactions during embryogenesis can dramatically influence skeletal myogenesis.
Synergistic effects of MAP2 and MAP1B knockout in neuronal migration, dendritic outgrowth, and microtubule organization
Authors: Teng Junlin , Takei Yosuke , Harada Akihiro , Nakata Takao , Chen Jianguo , Hirokawa Nobutaka
Published: July 2002

MAP1B and MAP2 are major members of neuronal microtubule-associated proteins (MAPs). To gain insights into the function of MAP2 in vivo, we generated MAP2-deficient (map2−/−) mice. They developed without any apparent abnormalities, which indicates that MAP2 is dispensable in mouse survival. Because previous reports suggest a functional redundancy among MAPs, we next generated mice lacking both MAP2 and MAP1B to test their possible synergistic functions in vivo. Map2−/−map1b−/− mice died in their perinatal period. They showed not only fiber tract malformations but also disrupted cortical patterning caused by retarded neuronal migration. In spite of this, their cortical layer maintained an “inside-out” pattern. Detailed observation of primary cultures of hippocampal neurons from map2−/−map1b−/− mice revealed inhibited microtubule bundling and neurite elongation. In these neurons, synergistic effects caused by the loss of MAP2 and MAP1B were more apparent in dendrites than in axons. The spacing of microtubules was reduced significantly in map2−/−map1b−/− mice in vitro and in vivo. These results suggest that MAP2 and MAP1B have overlapping functions in neuronal migration and neurite outgrowth by organizing microtubules in developing neurons both for axonal and dendritic morphogenesis but more dominantly for dendritic morphogenesis.
The role of cell cycle–regulated expression in the localization of spatial landmark proteins in yeast
Authors: Schenkman Laura R. , Caruso Carlo , Pagé Nicolas , Pringle John R.
Published: July 2002

In Saccharomyces cerevisiae, Bud8p and Bud9p are homologous plasma membrane glycoproteins that appear to mark the distal and proximal cell poles, respectively, as potential sites for budding in the bipolar pattern. Here we provide evidence that Bud8p is delivered to the presumptive bud site (and thence to the distal pole of the bud) just before bud emergence, and that Bud9p is delivered to the bud side of the mother-bud neck (and thence to the proximal pole of the daughter cell) after activation of the mitotic exit network, just before cytokinesis. Like the delivery of Bud8p, that of Bud9p is actin dependent; unlike the delivery of Bud8p, that of Bud9p is also septin dependent. Interestingly, although the transcription of BUD8 and BUD9 appears to be cell cycle regulated, the abundance of BUD8 mRNA peaks in G2/M and that of BUD9 mRNA peaks in late G1, suggesting that the translation and/or delivery to the cell surface of each protein is delayed and presumably also cell cycle regulated. The importance of time of transcription in localization is supported by promoter-swap experiments: expression of Bud8p from the BUD9 promoter leads to its localization predominantly to the sites typical for Bud9p, and vice versa. Moreover, expression of Bud8p from the BUD9 promoter fails to rescue the budding-pattern defect of a bud8 mutant but fully rescues that of a bud9 mutant. However, although expression of Bud9p from the BUD8 promoter fails to rescue a bud9 mutant, it also rescues only partially the budding-pattern defect of a bud8 mutant, suggesting that some feature(s) of the Bud8p protein is also important for Bud8p function. Experiments with chimeric proteins suggest that the critical element(s) is somewhere in the extracytoplasmic domain of Bud8p.
Evidence that Singlet Oxygen-induced Human T Helper Cell Apoptosis Is the Basic Mechanism of Ultraviolet-A Radiation Phototherapy
Authors: Morita Akimichi , Werfel Thomas , Stege Helger , Ahrens Constanze , Karmann Karin , Grewe Markus , Grether-Beck Susanne , Ruzicka Thomas , Kapp Alexander , Klotz Lars-Oliver , Sies Helmut , Krutmann Jean
Published: July 2002

Ultraviolet A (UVA) irradiation is effectively used to treat patients with atopic dermatitis and other T cell mediated, inflammatory skin diseases. In the present study, successful phototherapy of atopic dermatitis was found to result from UVA radiation-induced apoptosis in skin-infiltrating T helper cells, leading to T cell depletion from eczematous skin. In vitro, UVA radiation-induced human T helper cell apoptosis was mediated through the FAS/FAS-ligand system, which was activated in irradiated T cells as a consequence of singlet oxygen generation. These studies demonstrate that singlet oxygen is a potent trigger for the induction of human T cell apoptosis. They also identify singlet oxygen generation as a fundamental mechanism of action operative in phototherapy.
Mitochondrial Basis for Immune Deficiency
Authors: Arpaia Enrico , Benveniste Patricia , Di Cristofano Antonio , Gu Yiping , Dalal Ilan , Kelly Susan , Hershfield Michael , Pandolfi Pier Paolo , Roifman Chaim M. , Cohen Amos
Published: July 2002

We generated purine nucleoside phosphorylase (PNP)-deficient mice to gain insight into the mechanism of immune deficiency disease associated with PNP deficiency in humans. Similar to the human disease, PNP deficiency in mice causes an immunodeficiency that affects T lymphocytes more severely than B lymphocytes. PNP knockout mice exhibit impaired thymocyte differentiation, reduced mitogenic and allogeneic responses, and decreased numbers of maturing thymocytes and peripheral T cells. T lymphocytes of PNP-deficient mice exhibit increased apoptosis in vivo and higher sensitivity to gamma irradiation in vitro. We propose that the immune deficiency in PNP deficiency is a result of inhibition of mitochondrial DNA repair due to the accumulation of dGTP in the mitochondria. The end result is increased sensitivity of T cells to spontaneous mitochondrial DNA damage, leading to T cell depletion by apoptosis.
Involvement of Tweak in Interferon γ–Stimulated Monocyte Cytotoxicity
Authors: Nakayama Masafumi , Kayagaki Nobuhiko , Yamaguchi Noriko , Okumura Ko , Yagita Hideo
Published: July 2002

TWEAK, a new member of the tumor necrosis factor (TNF) family, induces cell death in some tumor cell lines, but its physiological functions are largely unknown. In this study, we investigated the expression and function of TWEAK in human peripheral blood mononuclear cells (PBMCs) by using newly generated anti–human TWEAK mAbs. Although freshly isolated PBMCs expressed no detectable level of TWEAK on their surfaces, a remarkable TWEAK expression was rapidly observed on monocytes upon stimulation with interferon (IFN)-γ but not with IFN-α or lipopolysaccharide. Cytotoxic activity of IFN-γ–stimulated monocytes against human squamous carcinoma cell line HSC3 was inhibited partially by anti-TWEAK mAb alone and almost completely by combination with anti-TRAIL (TNF-related apoptosis-inducing ligand) mAb. These results revealed a novel pathway of monocyte cytotoxicity against tumor cells that is mediated by TWEAK and potentiated by IFN-γ.
Identification of Regulatory T Cells in Tolerated Allografts
Authors: Graca Luis , Cobbold Stephen P. , Waldmann Herman
Published: July 2002

Induction of transplantation tolerance with certain therapeutic nondepleting monoclonal antibodies can lead to a robust state of peripheral “dominant” tolerance. Regulatory CD4+ T cells, which mediate this form of “dominant” tolerance, can be isolated from spleens of tolerant animals. To determine whether there were any extra-lymphoid sites that might harbor regulatory T cells we sought their presence in tolerated skin allografts and in normal skin. When tolerated skin grafts are retransplanted onto T cell–depleted hosts, graft-infiltrating T cells exit the graft and recolonize the new host. These colonizing T cells can be shown to contain members with regulatory function, as they can prevent nontolerant lymphocytes from rejecting fresh skin allografts, without hindrance of rejection of third party skin. Our results suggest that T cell suppression of graft rejection is an active process that operates beyond secondary lymphoid tissue, and involves the persistent presence of regulatory T cells at the site of the tolerated transplant.
On fourth-order elliptic boundary value problems
Authors: Pao C.
Published: July 2002

This paper is concerned with the existence and uniqueness of a solution for a class of fourth-order elliptic boundary value problems. The existence of a solution is proven by the method of upper and lower solutions without any monotone nondecreasing or nonincreasing property of the nonlinear function. Sufficient conditions for the uniqueness of a solution and some techniques for the construction of upper and lower solutions are given. All the existence and uniqueness results are directly applicable to fourth-order two-point boundary value problems.
Commutative quantum current operators, semi-infinite construction and functional models
Authors: Ding Jintai , Feigin Boris
Published: July 2002

We construct the commutative current operator

x
¯

+

(
z
)

\bar x^+(z)

inside

U
q

(

s
l

^

(
2
)
)

U_q(\hat {\mathfrak {sl}}(2))

. With this operator and the condition of quantum integrability on the quantum currents of

U
q

(

s
l

^

(
2
)
)

U_q(\hat {\mathfrak {sl}}(2))

, we derive the quantization of the semi-infinite construction of integrable modules of

s
l

^

(
2
)

\hat {\mathfrak {sl}}(2)

which has been previously obtained by means of the current operator

e
(
z
)

e(z)

of

s
l

^

(
2
)

\hat {\mathfrak {sl}}(2)

. The quantization of the functional models for

s
l

^

(
2
)

\hat {\mathfrak {sl}}(2)

is also given.
Irreducible Genuine Characters of the Metaplectic Group: Kazhdan-Lusztig Algorithm and Vogan Duality
Authors: Renard David , Trapa Peter
Published: July 2002

We establish a Kazhdan-Lusztig algorithm to compute characters of irreducible genuine representations of the (nonlinear) metaplectic group with half-integral infinitesimal character. We then prove a character multiplicity duality theorem for representations of

M
p
(
2
n
,

R

)

Mp(2n,\mathbb R)

at fixed half-integral infinitesimal character. This allows us to extend some of Langlands’ ideas to

M
p
(
2
n
,

R

)

Mp(2n,\mathbb R)

.
Extracellular Signal-regulated Kinase Phosphorylates Tumor Necrosis Factor α-converting Enzyme at Threonine 735: A Potential Role in Regulated Shedding
Authors: Dı́az-Rodrı́guez Elena , Montero Juan Carlos , Esparı́s-Ogando Azucena , Yuste Laura , Pandiella Atanasio
Published: July 2002

The ectodomain of certain transmembrane proteins can be released by the action of cell surface proteases, termed secretases. Here we have investigated how mitogen-activated protein kinases (MAPKs) control the shedding of membrane proteins. We show that extracellular signal-regulated kinase (Erk) acts as an intermediate in protein kinase C-regulated TrkA cleavage. We report that the cytosolic tail of the tumor necrosis factor α-converting enzyme (TACE) is phosphorylated by Erk at threonine 735. In addition, we show that Erk and TACE associate. This association is favored by Erk activation and by the presence of threonine 735. In contrast to the Erk route, the p38 MAPK was able to stimulate TrkA cleavage in cells devoid of TACE activity, indicating that other proteases are also involved in TrkA shedding. These results demonstrate that secretases are able to discriminate between the different stimuli that trigger membrane protein ectodomain cleavage and indicate that phosphorylation by MAPKs may regulate the proteolytic function of membrane secretases.
Systematic Structure-Function Analysis of the Small GTPase Arf1 in Yeast
Authors: Click Eleanor S. , Stearns Tim , Botstein David
Published: July 2002

Members of the ADP-ribosylation factor (Arf) family of small GTPases are implicated in vesicle traffic in the secretory pathway, although their precise function remains unclear. We generated a series of 23 clustered charge-to-alanine mutations in the Arf1 protein ofSaccharomyces cerevisiae to determine the portions of this protein important for its function in cells. These mutants display a number of phenotypes, including conditional lethality at high or low temperature, defects in glycosylation of invertase, dominant lethality, fluoride sensitivity, and synthetic lethality with thearf2 null mutation. All mutations were mapped onto the available crystal structures for Arf1p: Arf1p bound to GDP, to GTP, and complexed with the regulatory proteins ArfGEF and ArfGAP. From this systematic structure-function analysis we demonstrate that all essential mutations studied map to one hemisphere of the protein and provide strong evidence in support of the proposed ArfGEF contact site on Arf1p but minimal evidence in support of the proposed ArfGAP-binding site. In addition, we describe the isolation of a spatially distant intragenic suppressor of a dominant lethal mutation in the guanine nucleotide-binding region of Arf1p.
Caenorhabditis elegansInositol 5-Phosphatase Homolog Negatively Regulates Inositol 1,4,5-Triphosphate Signaling in Ovulation
Authors: Bui Yen Kim , Sternberg Paul W.
Published: July 2002

Ovulation in Caenorhabditis elegans requires inositol 1,4,5-triphosphate (IP3) signaling activated by the epidermal growth factor (EGF)-receptor homolog LET-23. We generated a deletion mutant of a type I 5-phosphatase,ipp-5, and found a novel ovulation phenotype whereby the spermatheca hyperextends to engulf two oocytes per ovulation cycle. The temporal and spatial expression of IPP-5 is consistent with its proposed inhibition of IP3signaling in the adult spermatheca. ipp-5 acts downstream oflet-23, and interacts withlet-23–mediated IP3signaling pathway genes. We infer that IPP-5 negatively regulates IP3signaling to ensure proper spermathecal contraction.