bluuebunny/crossref_metadata_2025 huggingface dataset:
Pneumonia: The Demented Patient's Best Friend? Discomfort After Starting or Withholding Antibiotic Treatment
Authors: Van Der Steen Jenny T. , Ooms Marcel E. , Van Der Wal Gerrit , Ribbe Miel W.
Published: October 2002

OBJECTIVES: To assess suffering in demented nursing home patients with pneumonia treated with antibiotics or without antibiotics. This study should provide the first empirical data on whether pneumonia is a “friend” or an “enemy” of demented patients and promote a debate on appropriate palliative care.

DESIGN: Prospective cohort study.

SETTING: Psychogeriatric wards of 61 nursing homes in the Netherlands.

PARTICIPANTS: Six hundred sixty‐two demented patients with pneumonia treated with (77%) or without (23%) antibiotics.

MEASUREMENTS: Using an observational scale (Discomfort Scale—Dementia of Alzheimer Type), discomfort was assessed at the time of the pneumonia treatment decision and periodically thereafter for 3 months or until death. (Thirty‐nine percent of patients treated with antibiotics and 93% of patients treated without antibiotics died within 3 months.) Physicians also offered a retrospective judgment of discomfort 2 weeks before the treatment decision. In addition, pneumonia symptoms were assessed at baseline and on follow‐up. Linear regression was performed with discomfort shortly before death as an outcome.

RESULTS: A peak in discomfort was observed at baseline. Compared with surviving patients treated with antibiotics, the level of discomfort was generally higher in patients in whom antibiotic treatment was withheld and in nonsurvivors. However, these same patients had more discomfort before the pneumonia. Breathing problems were most prominent. Shortly before death from pneumonia, discomfort increased. Discomfort was higher shortly before death when pneumonia was the final cause of death than with death from other causes.

CONCLUSION: Irrespective of antibiotic treatment, pneumonia causes substantial suffering in demented patients. Adequate symptomatic treatment deserves priority attention.
Smoking Patterns Among Seniors and the Medicare Stop Smoking Program
Authors: Arday David R. , Lapin Pauline , Chin Joseph , Preston Jeanette A.
Published: October 2002

OBJECTIVES: To characterize smoking patterns in the older U.S. community‐dwelling Medicare population at the national level and in states chosen to participate in the new Medicare Stop Smoking Program (MSSP) demonstration. To describe the MSSP.

DESIGN: Data from the Behavioral Risk Factor Surveillance System (BRFSS) 1996 to 1999 were analyzed.

SETTING: The BRFSS is a cross‐sectional random‐digit‐dialed telephone survey conducted in all states plus the District of Columbia and Puerto Rico.

PARTICIPANTS: BRFSS respondents aged 65 and older who self‐identified as receiving Medicare benefits.

MEASUREMENTS: Using BRFSS core questionnaire variables, recent trends in prevalence of current smoking and smoking cessation were estimated, as were prevalences by various demographic characteristics, for both the nation and the MSSP states as a group.

RESULTS: As of 1999, an estimated 10.2% of this population were current smokers, with those aged 65 to 74 smoking at twice the rate (12.9%) of those aged 75 and older (6.1%) and blacks (14.7%) smoking more than whites (10.0%). Between 1996 and 1999, the prevalence of everyday smokers indicating they had attempted to quit for 1 day or longer in the past year rose from 37.1% to 42.2%. National patterns were mirrored in the states chosen to participate in the MSSP.

CONCLUSIONS: Young‐old Medicare recipients have a higher smoking prevalence, although interest in quitting appears to be rising. The chosen MSSP states appear to be a representative of national smoking patterns in the older Medicare population.
Preoperative Supraphysiological Testosterone in Older Men Undergoing Knee Replacement Surgery
Authors: Amory John K. , Chansky Howard A. , Chansky Kari L. , Camuso Matthew R. , Hoey Chris T. , Anawalt Bradley D. , Matsumoto Alvin M. , Bremner William J.
Published: October 2002

OBJECTIVES: Older patients undergoing knee replacement surgery can recover more slowly than younger patients and require extended rehabilitation. Because administration of supraphysiological testosterone (T) dramatically increases strength, we hypothesized that preoperative T therapy would improve functional recovery and reduce hospital stay in older men undergoing knee replacement surgery.

DESIGN: Double‐blinded, placebo‐controlled pilot trial.

SETTING: A Veterans Affairs orthopedics clinic and inpatient postoperative unit.

PARTICIPANTS: Twenty‐five men, mean age 70, undergoing elective knee replacement.

INTERVENTION: Preoperative, supraphysiological T administration (600 mg T enanthate intramuscularly weekly for 4 weeks) or sesame oil placebo.

MEASUREMENTS: Length of hospital stay and functional ability by Functional Independence Measure (FIM) score.

RESULTS: Mean length of hospital stay ± standard deviation was nonsignificantly reduced in the T group (5.9 ± 2.4 days vs 6.8 ± 2.5 days; P = .15). At postoperative Day 3, there was a significant improvement in ability to stand (mean FIM score 5.2 ± 1.0 vs 4.0 ± 1.1; P = .04) and trends towards improvements in walking and stair climbing in the T group. There were no complications attributable to T therapy.

CONCLUSIONS: In older men undergoing knee replacement surgery, preoperative supraphysiological T administration may confer some clinical benefit. Future studies using longer courses of preoperative T administration in larger numbers of older men undergoing knee replacement surgery are warranted.
Floor‐Rise Strategy Training in Older Adults
Authors: Hofmeyer Mark R. , Alexander Neil B. , Nyquist Linda V. , Medell Jodi L. , Koreishi Aaleya
Published: October 2002

OBJECTIVES: To determine the effect of a 2‐week (six‐session) training intervention to improve the ability of disabled older adults to rise from the floor.

DESIGN: Prospective intervention trial.

SETTING: Congregate housing in Michigan.

PARTICIPANTS: Subjects aged 65 and older who admitted to requiring assistance (such as from a person, equipment, or device) in performing at least one of the following mobility‐related activities of daily living: transferring, walking, bathing, and toileting.

INTERVENTION: Participants were randomly allocated to individual training (n = 17, mean age 81) in strategies to rise from the floor (using for example, certain key intermediate body positions) or a control chair‐based flexibility intervention (n = 18, mean age 80).

MEASUREMENTS: At baseline and postintervention, residents were queried regarding their rise difficulty (difficulty scale) and symptoms (symptoms scale) associated with the rise and were tested in their ability to perform timed floor‐rise tasks. These tasks varied in starting position (supine vs all fours) and in use of a support to assist in rising (no support, use of an end table, use of a chair).

RESULTS: Using baseline performance as the covariate, by analysis of covariance (ANCOVA), the training group showed a significant (P < .05) improvement in mean number of rise tasks completed (baseline mean 6.6, postintervention mean 7.3) versus essentially no improvement in the controls. Similarly, by ANCOVA, the training group (compared with controls) showed a significant (P < .05) improvement on the difficulty and symptoms scales. There was no intervention effect for rise time.

CONCLUSIONS: A short‐term, strategy‐based intervention improved floor‐rise ability and perceived difficulty and symptoms associated with the rise. This approach, focusing on key intermediate body positions, may be useful in training floor‐rise skills, particularly in older adults at risk for falls.
Proton Magnetic Resonance Spectroscopy Reveals Similar White Matter Biochemical Changes in Patients with Chronic Hypertension and Early Alzheimer's Disease
Authors: Catani Marco , Mecocci Patrizia , Tarducci Roberto , Howard Robert , Pelliccioli Gian Piero , Mariani Elena , Metastasio Antonio , Benedetti Claudia , Senin Umberto , Cherubini Antonio
Published: October 2002

OBJECTIVES: Hypertension is a risk factor for dementia and is associated with some of the brain changes that are found in Alzheimer's disease and other neurodegenerative diseases, such as atrophy and neurofibrillary tangles. We evaluated the cerebral white matter biochemical pattern in healthy older subjects, older patients with chronic hypertension, and patients with Alzheimer's disease (AD) using proton magnetic resonance spectroscopy (1H‐MRS).

DESIGN: Cross‐sectional study.

SETTING: University‐affiliated outpatient clinic.

PARTICIPANTS: Ten healthy older subjects, 10 cognitively intact older patients with chronic hypertension, and 10 older patients with early AD.

MEASUREMENTS: All subjects underwent clinical examination, neuropsychological assessment, and 1H‐MRS to measure N‐acetylaspartate (NAA), myoinositol, choline, and creatine resonance signals in an 8‐cm3 voxel located in the paratrigonal white matter region bilaterally. NAA/creatine, myoinositol/creatine, and choline/creatine ratios were measured, and the mean values were compared using one‐way analysis of variance with Tukey test for post hoc analysis.

RESULTS: A significantly higher mean myoinositol/creatine (ratio ± standard deviation) was found in hypertensive patients (0.67 ± 0.05) and in AD patients (0.68 ± 0.08) than in controls (0.56 ± 0.04) (P < .001). Conversely neither NAA/creatine ratio nor choline/creatine ratio differed among the three groups.

CONCLUSIONS: In this study, cognitively intact chronic hypertensive older patients had a higher white matter myoinositol/creatine ratio compared with healthy older subjects, suggesting that myoinositol may be a sensitive marker of the effects of chronic hypertension on the brain. Moreover, the similar increase of myoinositol/creatine ratio in patients with hypertension and in those with early AD provides further evidence of common brain changes with these conditions.
Knowledge and Beliefs About Influenza, Pneumococcal Disease, and Immunizations Among Older People
Authors: Santibanez Tammy A. , Nowalk M. Patricia , Zimmerman Richard K. , Jewell Ilene Katz , Bardella Inis J. , Wilson Stephen A. , Terry Martha A.
Published: October 2002

OBJECTIVES: Despite the burden of disease caused by influenza and pneumococcus, immunization rates are moderate and have not reached national goals set for 2010. This study's objective was to identify patient knowledge, attitudes, and beliefs that serve as facilitators of and barriers to influenza and pneumococcal vaccination.

DESIGN: A survey conducted in 2000 by computer‐assisted telephone interviewing.

SETTING: To encounter a broad spectrum of patients and healthcare systems, we sampled patients at inner‐city health centers, Department of Veterans Affairs outpatient clinics, and rural and suburban practices.

PARTICIPANTS: Inclusion criteria were patients aged 66 and older and an office visit after September 30, 1998.

MEASUREMENTS: Responses to questionnaire.

RESULTS: Overall, 1,007 (82%) interviews were completed among 1,234 people contacted by phone. Vaccination against pneumococcal disease was significantly related to being able to accurately describe one or more classic symptoms of pneumonia (P = .05). Vaccination against influenza and pneumococcal disease was significantly related to belief that vaccination was the best way to prevent these diseases (P < .001). The unvaccinated reported that they felt they were not likely to contract influenza and that they did not know they needed the pneumococcal vaccine. Access was not related to vaccination status.

CONCLUSIONS: Educational campaigns to increase vaccination rates among older adults should focus on symptoms of, risk for, and severity of influenza and pneumococcal diseases and encouraging physicians to recommend the vaccines to their patients.
The Alcohol‐Related Problems Survey: Identifying Hazardous and Harmful Drinking in Older Primary Care Patients
Authors: Fink Arlene , Morton Sally C. , Beck John C. , Hays Ron D. , Spritzer Karen , Oishi Sabine , Moore Alison A.
Published: October 2002

OBJECTIVES: Older adults can incur problems at low levels of alcohol consumption because of age‐related physiological changes, declining health and functional status, and medication use. We have developed and tested a screening measure specifically for older people, the Alcohol‐Related Problems Survey (ARPS), to identify older adults with these risks.

DESIGN: Survey.

SETTING: Academic and community primary care clinics.

PARTICIPANTS: Five hundred forty‐nine current drinkers aged 65 and older, mostly white with high school or more education.

MEASURES: Alcohol use was classified as harmful, hazardous, or nonhazardous depending upon consumption alone or combined with selected comorbidities and medication use. Harmful drinking (including alcohol abuse or dependence) means the presence of problems (e.g., hypertension, adverse drug events, legal problems) due to drinking. Hazardous drinking means risks for problems are likely. Nonhazardous drinking poses no known risks for problems.

RESULTS: Eleven percent of subjects were harmful drinkers and 35% were hazardous drinkers. Harmful drinking was more common in men than women and in persons younger than 75 than those aged 75 and older. Similar proportions of men and women and younger and older age groups were hazardous drinkers. Most harmful drinkers were identified by their use of alcohol with their comorbidity, whereas most hazardous drinkers were identified by their use of alcohol with medications. Test‐retest reliability was substantial (κ = 0.65).

CONCLUSION: Physicians are urged to screen for alcohol‐related problems in older persons. The ARPS reliably identifies harmful, hazardous, and nonhazardous drinking in older adults resulting most often from the interaction between alcohol and disease and medication use.
Delirium Superimposed on Dementia: A Systematic Review
Authors: Fick Donna M. , Agostini Joseph V. , Inouye Sharon K.
Published: October 2002

Delirium in a patient with preexisting dementia is a common problem that may have serious complications and poor prognostic implications. The purpose of this paper was to conduct a systematic review of the medical literature on delirium superimposed on dementia, specifically to review studies on prevalence, associated features, outcomes, and management. Areas of controversy and gaps in our knowledge of this problem are highlighted. Finally, an agenda for future research is proposed. Fourteen studies were reviewed, including seven prospective studies, three retrospective studies, two cross‐sectional studies, and two clinical trials. For the review of the literature on delirium superimposed on dementia, we searched MEDLINE from January 1966 through February 2002 for research studies with primary sources of data. Selection criteria for inclusion of articles in this study were inclusion of data on subjects with delirium superimposed on dementia, inclusion of a validated operational definition/measures of dementia and delirium, actual data on persons with delirium and dementia reported in the paper, and reporting of primary data. MEDLINE was searched using the following key search terms: delirium, acute confusion, cognitive impairment, Alzheimer's disease, dementia, delirium superimposed on dementia, and elderly. The prevalence of delirium superimposed on dementia ranged from 22% to 89% of hospitalized and community populations aged 65 and older with dementia. To date, only one reported study systematically identified associated factors and interventions for delirium superimposed on dementia, but several studies examining outcomes have found that adverse events are associated with delirium in persons with dementia, including accelerated and long‐term cognitive and functional decline, need for institutionalization, rehospitalization, and increased mortality. This paper highlights the dearth of research on delirium superimposed on dementia and stresses the importance of early recognition and prevention of delirium in persons with dementia.
Interactions of climate change with biological invasions and land use in the Hawaiian Islands: Modeling the fate of endemic birds using a geographic information system
Authors: Benning Tracy L. , LaPointe Dennis , Atkinson Carter T. , Vitousek Peter M.
Published: October 2002

The Hawaiian honeycreepers (Drepanidae) represent a superb illustration of evolutionary radiation, with a single colonization event giving rise to 19 extant and at least 10 extinct species [Curnutt, J. & Pimm, S. (2001)Stud. Avian Biol.22, 15–30]. They also represent a dramatic example of anthropogenic extinction. Crop and pasture land has replaced their forest habitat, and human introductions of predators and diseases, particularly of mosquitoes and avian malaria, has eliminated them from the remaining low- and mid-elevation forests. Landscape analyses of three high-elevation forest refuges show that anthropogenic climate change is likely to combine with past land-use changes and biological invasions to drive several of the remaining species to extinction, especially on the islands of Kauai and Hawaii.
Uptake of the anticancer drug cisplatin mediated by the copper transporter Ctr1 in yeast and mammals
Authors: Ishida Seiko , Lee Jaekwon , Thiele Dennis J. , Herskowitz Ira
Published: October 2002

Cisplatin is a chemotherapeutic drug used to treat a variety of cancers. Both intrinsic and acquired resistance to cisplatin, as well as toxicity, limit its effectiveness. Molecular mechanisms that underlie cisplatin resistance are poorly understood. Here we demonstrate that deletion of the yeast
CTR1
gene, which encodes a high-affinity copper transporter, results in increased cisplatin resistance and reduced intracellular accumulation of cisplatin. Copper, which causes degradation and internalization of Ctr1 protein (Ctr1p), enhances survival of wild-type yeast cells exposed to cisplatin and reduces cellular accumulation of the drug. Cisplatin also causes degradation and delocalization of Ctr1p and interferes with copper uptake in wild-type yeast cells. Mouse cell lines lacking one or both mouse
Ctr1
(
mCtr1
) alleles exhibit increased cisplatin resistance and decreased cisplatin accumulation in parallel with
mCtr1
gene dosage. We propose that cisplatin uptake is mediated by the copper transporter Ctr1p in yeast and mammals. The link between Ctr1p and cisplatin transport may explain some cases of cisplatin resistance in humans and suggests ways of modulating sensitivity and toxicity to this important anticancer drug.
Genome sequence of
Streptococcus mutans
UA159, a cariogenic dental pathogen
Authors: Ajdić Dragana , McShan William M. , McLaughlin Robert E. , Savić Gorana , Chang Jin , Carson Matthew B. , Primeaux Charles , Tian Runying , Kenton Steve , Jia Honggui , Lin Shaoping , Qian Yudong , Li Shuling , Zhu Hua , Najar Fares , Lai Hongshing , White Jim , Roe Bruce A. , Ferretti Joseph J.
Published: October 2002

Streptococcus mutans
is the leading cause of dental caries (tooth decay) worldwide and is considered to be the most cariogenic of all of the oral streptococci. The genome of
S. mutans
UA159, a serotype c strain, has been completely sequenced and is composed of 2,030,936 base pairs. It contains 1,963 ORFs, 63% of which have been assigned putative functions. The genome analysis provides further insight into how
S. mutans
has adapted to surviving the oral environment through resource acquisition, defense against host factors, and use of gene products that maintain its niche against microbial competitors.
S. mutans
metabolizes a wide variety of carbohydrates via nonoxidative pathways, and all of these pathways have been identified, along with the associated transport systems whose genes account for almost 15% of the genome. Virulence genes associated with extracellular adherent glucan production, adhesins, acid tolerance, proteases, and putative hemolysins have been identified. Strain UA159 is naturally competent and contains all of the genes essential for competence and quorum sensing. Mobile genetic elements in the form of IS elements and transposons are prominent in the genome and include a previously uncharacterized conjugative transposon and a composite transposon containing genes for the synthesis of antibiotics of the gramicidin/bacitracin family; however, no bacteriophage genomes are present.
The innermost inner core of the earth: Evidence for a change in anisotropic behavior at the radius of about 300 km
Authors: Ishii Miaki , Dziewoński Adam M.
Published: October 2002

Since the discovery of the inner core in 1936, no additional spherical subshell of the Earth has been observed. Based on an extensive seismic data set, we propose the existence of an innermost inner core, with a radius of ∼300 km, that exhibits a distinct transverse isotropy relative to the bulk inner core. Specifically, within the innermost inner core, the slowest direction of wave propagation is ∼45° from the east-west direction. In contrast, the direction of the slowest wave propagation in the overlying inner core is east-west. The distinct anisotropy at the center of the Earth may represent fossil evidence of a unique early history of inner-core evolution.
Resistance of α-synuclein null mice to the parkinsonian neurotoxin MPTP
Authors: Dauer William , Kholodilov Nikolai , Vila Miquel , Trillat Anne-Cecile , Goodchild Rose , Larsen Kristin E. , Staal Roland , Tieu Kim , Schmitz Yvonne , Yuan Chao Annie , Rocha Marcelo , Jackson-Lewis Vernice , Hersch Steven , Sulzer David , Przedborski Serge , Burke Robert , Hen Rene
Published: October 2002

Parkinson's disease (PD) is most commonly a sporadic illness, and is characterized by degeneration of substantia nigra dopamine (DA) neurons and abnormal cytoplasmic aggregates of α-synuclein. Rarely, PD may be caused by missense mutations in α-synuclein. MPTP, a neurotoxin that inhibits mitochondrial complex I, is a prototype for an environmental cause of PD because it produces a pattern of DA neurodegeneration that closely resembles the neuropathology of PD. Here we show that α-synuclein null mice display striking resistance to MPTP-induced degeneration of DA neurons and DA release, and this resistance appears to result from an inability of the toxin to inhibit complex I. Contrary to predictions from
in vitro
data, this resistance is not due to abnormalities of the DA transporter, which appears to function normally in α-synuclein null mice. Our results suggest that some genetic and environmental factors that increase susceptibility to PD may interact with a common molecular pathway, and represent the first demonstration that normal α-synuclein function may be important to DA neuron viability.
Cell surface polarization during yeast mating
Authors: Bagnat Michel , Simons Kai
Published: October 2002

Exposure to mating pheromone in haploid
Saccharomyces cerevisiae
cells results in the arrest of the cell cycle, expression of mating-specific genes, and polarized growth toward the mating partner. Proteins involved in signaling, polarization, cell adhesion, and fusion are localized to the tip of the mating cell (shmoo) where fusion will eventually occur. The mechanisms ensuring the correct targeting and retention of these proteins are poorly understood. Here we show that in pheromone-treated cells, a reorganization of the plasma membrane involving lipid rafts results in the retention of proteins at the tip of the mating projection, segregated from the rest of the membrane. Sphingolipid and ergosterol biosynthetic mutants fail to polarize proteins to the tip of the shmoo and are deficient in mating. Our results show that membrane microdomain clustering at the mating projection is involved in the generation and maintenance of polarity during mating.
Identification of macromolecular complexes in cryoelectron tomograms of phantom cells
Authors: Frangakis Achilleas S. , Böhm Jochen , Förster Friedrich , Nickell Stephan , Nicastro Daniela , Typke Dieter , Hegerl Reiner , Baumeister Wolfgang
Published: October 2002

Electron tomograms of intact frozen-hydrated cells are essentially three-dimensional images of the entire proteome of the cell, and they depict the whole network of macromolecular interactions. However, this information is not easily accessible because of the poor signal-to-noise ratio of the tomograms and the crowded nature of the cytoplasm. Here, we describe a template matching algorithm that is capable of detecting and identifying macromolecules in tomographic volumes in a fully automated manner. The algorithm is based on nonlinear cross correlation and incorporates elements of multivariate statistical analysis. Phantom cells, i.e., lipid vesicles filled with macromolecules, provide a realistic experimental scenario for an assessment of the fidelity of this approach. At the current resolution of ≈4 nm, macromolecules in the size range of 0.5–1 MDa can be identified with good fidelity.
A probabilistic explanation of brightness scaling
Authors: Nundy Surajit , Purves Dale
Published: October 2002

The perceptions of lightness or brightness elicited by a visual target are linked to its luminance by a nonlinear function that varies according to the physical characteristics of the target and the background on which it is presented. Although no generally accepted explanation of this scaling relationship exists, it has long been considered a byproduct of low- or mid-level visual processing. Here we examine the possibility that brightness scaling is actually the signature of a biological strategy for dealing with inevitably ambiguous visual stimuli, in which percepts of lightness/brightness are determined by the probabilistic relationship between luminances in the image plane and their possible real-world sources.
The SQV-1 UDP-glucuronic acid decarboxylase and the SQV-7 nucleotide-sugar transporter may act in the Golgi apparatus to affect
Caenorhabditis elegans
vulval morphogenesis and embryonic development
Authors: Hwang Ho-Yon , Horvitz H. Robert
Published: October 2002

Recent findings indicate that glycosaminoglycans can play important roles in animal development. The genes
sqv
-
3
, -
7
, and -
8
, which are necessary for vulval morphogenesis in
Caenorhabditis elegans
, affect the biosynthesis of chondroitin and heparan sulfate glycosaminoglycans. We cloned
sqv
-
1
and showed that the SQV-1 protein is a type II transmembrane protein that functions as a UDP-glucuronic acid decarboxylase. SQV-1 localizes to punctate cytoplasmic compartments and colocalizes with the SQV-7 nucleotide-sugar transporter, which probably acts in the Golgi apparatus. SQV-1 and SQV-7 are both expressed in the vulva and in oocytes, where they likely act in vulval morphogenesis and embryonic development, respectively. Progeny of
sqv
-
7
and
sqv
-
1
null mutants fail to initiate cytokinesis, possibly because they are unable to separate the plasma membrane from the eggshell, a defect analogous to that of incomplete vulval invagination.
TheCaenorhabditis elegansvulval morphogenesis genesqv-4encodes a UDP-glucose dehydrogenase that is temporally and spatially regulated
Authors: Hwang Ho-Yon , Horvitz H. Robert
Published: October 2002

The development of theCaenorhabditis elegansvulva requires the involution of epithelial cells and provides a model for organ morphogenesis. Mutations inC. elegans sqv(squashedvulva) genes affect both vulval morphogenesis and embryonic development. We found thatsqv-4encodes a protein similar to UDP-glucose dehydrogenases and showed that the SQV-4 protein specifically catalyzes the conversion of UDP-glucose to UDP-glucuronic acid, which is essential for the biosynthesis of chondroitin and heparan sulfate proteoglycans. SQV-4 is expressed in the vulva and in oocytes, among many other cells, and SQV-4 levels are dramatically increased in a specific subset of vulval cells during vulval morphogenesis. We propose that the regulation of UDP-glucuronic acid production in a specific subset of vulval cells helps determine the shape of the vulva.
Measurement of microsecond dynamic motion in the intestinal fatty acid binding protein by using fluorescence correlation spectroscopy
Authors: Chattopadhyay Krishnananda , Saffarian Saveez , Elson Elliot L. , Frieden Carl
Published: October 2002

Fluorescence correlation spectroscopy (FCS) measurements have been carried out on the intestinal fatty acid binding protein (IFABP) to study microsecond dynamics of the protein in its native state as well as in pH-induced intermediates. IFABP is a small (15 kDa) protein that consists mostly of antiparallel β-strands enclosing a large central cavity into which the ligand binds. Because this protein does not contain cysteine, two cysteine mutants (Val60Cys and Phe62Cys) have been prepared and covalently modified with fluorescein. Based on fluorescence measurements, one of the mutants (Val60Flu) has the fluorescein moiety inside the cavity of the protein, whereas the fluorescein is exposed to solvent in the other (Phe62Flu). The protein modified at position 60 demonstrates the presence of a conformational event on the order of 35 μsec, which is not seen in the other mutant (Phe62Flu). The amplitude of this fast conformational event decreases sharply at low pH as the protein unfolds. Experiments measuring the diffusion as a function of pH indicate the formation of a compact state distinct from the native state at about pH 3.5. Steady state fluorescence and far-UV CD indicates that unfolding occurs at pH values below pH 3.
Pairs of dipeptides synergistically activate the binding of substrate by ubiquitin ligase through dissociation of its autoinhibitory domain
Authors: Du Fangyong , Navarro-Garcia Federico , Xia Zanxian , Tasaki Takafumi , Varshavsky Alexander
Published: October 2002

Protein degradation by the ubiquitin (Ub) system controls the concentrations of many regulatory proteins. The degradation signals (degrons) of these proteins are recognized by the system's Ub ligases (complexes of E2 and E3 enzymes). Two substrate-binding sites of UBR1, the E3 of the N-end rule pathway in the yeast
Saccharomyces cerevisiae
, recognize basic (type 1) and bulky hydrophobic (type 2) N-terminal residues of proteins or short peptides. A third substrate-binding site of UBR1 targets CUP9, a transcriptional repressor of the peptide transporter PTR2, through an internal (non-N-terminal) degron of CUP9. Previous work demonstrated that dipeptides with destabilizing N-terminal residues allosterically activate UBR1, leading to accelerated
in vivo
degradation of CUP9 and the induction of PTR2 expression. Through this positive feedback,
S. cerevisiae
can sense the presence of extracellular peptides and react by accelerating their uptake. Here, we show that dipeptides with destabilizing N-terminal residues cause dissociation of the C-terminal autoinhibitory domain of UBR1 from its N-terminal region that contains all three substrate-binding sites. This dissociation, which allows the interaction between UBR1 and CUP9, is strongly increased only if both type 1- and type 2-binding sites of UBR1 are occupied by dipeptides. An aspect of autoinhibition characteristic of yeast UBR1 also was observed with mammalian (mouse) UBR1. The discovery of autoinhibition in Ub ligases of the UBR family indicates that this regulatory mechanism may also control the activity of other Ub ligases.
A family of RIM-binding proteins regulated by alternative splicing: Implications for the genesis of synaptic active zones
Authors: Wang Yun , Liu Xinran , Biederer Thomas , Südhof Thomas C.
Published: October 2002

RIMs are presynaptic active zone proteins that regulate neurotransmitter release. We describe two related genes that encode proteins with identical C-terminal sequences that bind to the conserved PDZ domain of RIMs via an unusual PDZ-binding motif. These proteins were previously reported separately as
E
LKS,
R
ab6-interacting protein 2, and
C
AST, leading us to refer to them by the acronym ERC. Alternative splicing of the C terminus of ERC1 generates a longer ERC1a variant that does not bind to RIMs and a shorter ERC1b variant that binds to RIMs, whereas the C terminus of ERC2 is synthesized only in a single RIM-binding variant. ERC1a is expressed ubiquitously as a cytosolic protein outside of brain; ERC1b is detectable only in brain, where it is both a cytosolic protein and an insoluble active zone component; and ERC2 is brain-specific but exclusively localized to active zones. Only brain-specific ERCs bind to RIMs, but both ubiquitous and brain-specific ERCs bind to Rab6, a GTP-binding protein involved in membrane traffic at the Golgi complex. ERC1a and ERC1b/2 likely perform similar functions at distinct localizations, indicating unexpected connections between nonneuronal membrane traffic at the Golgi complex executed via Rab6 and neuronal membrane traffic at the active zone executed via RIMs.
Role of the lever arm in the processive stepping of myosin V
Authors: Purcell Thomas J. , Morris Carl , Spudich James A. , Sweeney H. Lee
Published: October 2002

Myosin V is a two-headed molecular motor that binds six light chains per heavy chain, which creates unusually long lever arms. This motor moves processively along its actin track in discrete 36-nm steps. Our model is that one head of the two-headed myosin V tightly binds to actin and swings its long lever arm through a large angle, providing a stroke. We created single-headed constructs with different-size lever arms and show that stroke size is proportional to lever arm length. In a two-headed molecule, the stroke provides the directional bias, after which the unbound head diffuses to find its binding site, 36 nm forward. Our two-headed construct with all six light chains per head reconstitutes the 36-nm processive step seen in tissue-purified myosin V. Two-headed myosin V molecules with only four light chains per head are still processive, but their step size is reduced to 24 nm. A further reduction in the length of the lever arms to one light chain per head results in a motor that is unable to walk processively. This motor produces single small ≈6-nm strokes, and ATPase and pyrene actin quench measurements show that only one of the heads of this dimer rapidly binds to actin for a given binding event. These data show that for myosin V with its normal proximal tail domain, both heads and a long lever arm are required for large, processive steps.
Does RNA polymerase help drive chromosome segregation in bacteria?
Authors: Dworkin Jonathan , Losick Richard
Published: October 2002

In contrast to eukaryotic cells, bacteria segregate their chromosomes without a conspicuous mitotic apparatus. Replication of bacterial chromosomes initiates bidirectionally from a single site (
oriC
), and visualization of the region of the chromosome containing
oriC
in living cells reveals that origins rapidly move apart toward opposite poles of the cell during the cell cycle. The motor that drives this poleward movement is unknown. An attractive candidate is RNA polymerase, which is a powerful and abundant molecular motor. If, as has been suggested for other macromolecular complexes, the movement of RNA polymerase is restricted in the cell, then transcription would translocate the DNA template, thereby providing the motive force to separate replicating chromosomes. A coordinated effect of many transcribing RNA polymerases could result from the widely conserved global bias of gene orientation away from
oriC
. By using fluorescence microscopy of living
Bacillus subtilis
cells, we demonstrate that an inhibitor of RNA polymerase acts to inhibit separation of newly duplicated DNAs near the origin of replication. We propose that the force exerted by RNA polymerase contributes to chromosome movement in bacteria, and that this force, coupled with the biased orientation of transcription units, helps to drive chromosome segregation.
On the simulation of protein folding by short time scale molecular dynamics and distributed computing
Authors: Fersht Alan R.
Published: October 2002

There are proposals to overcome the current incompatibilities between the time scales of protein folding and molecular dynamics simulation by using a large number of short simulations of only tens of nanoseconds (distributed computing). According to the principles of first-order kinetic processes, a sufficiently large number of short simulations will include,
de facto
, a small number of long time scale events that have proceeded to completion. But protein folding is not an elementary kinetic step: folding has a series of early conformational steps that lead to lag phases at the beginning of the kinetics. The presence of these lag phases can bias short simulations toward selecting minor pathways that have fewer or faster lag steps and so miss the major folding pathways. Attempts to circumvent the lags by using loosely coupled parallel simulations that search for first-order transitions are also problematic because of the difficulty of detecting transitions in molecular dynamics simulations. Nevertheless, the procedure of using parallel independent simulations is perfectly valid and quite feasible once the time scale of simulation proceeds past the lag phases into a single exponential region.
Small molecule modulation of Smoothened activity
Authors: Chen James K. , Taipale Jussi , Young Keith E. , Maiti Tapan , Beachy Philip A.
Published: October 2002

Smoothened (Smo), a distant relative of G protein-coupled receptors, mediates Hedgehog (Hh) signaling during embryonic development and can initiate or transmit ligand-independent pathway activation in tumorigenesis. Although the cellular mechanisms that regulate Smo function remain unclear, the direct inhibition of Smo by cyclopamine, a plant-derived steroidal alkaloid, suggests that endogenous small molecules may be involved. Here we demonstrate that SAG, a chlorobenzothiophene-containing Hh pathway agonist, binds to the Smo heptahelical bundle in a manner that antagonizes cyclopamine action. In addition, we have identified four small molecules that directly inhibit Smo activity but are structurally distinct from cyclopamine. Functional and biochemical studies of these compounds provide evidence for the small molecule modulation of Smo through multiple mechanisms and yield insights into the physiological regulation of Smo activity. The mechanistic differences between the Smo antagonists may be useful in the therapeutic manipulation of Hh signaling.
Despite ubiquitous autoantigen expression, arthritogenic autoantibody response initiates in the local lymph node
Authors: Mandik-Nayak Laura , Wipke Brian T. , Shih Fei F. , Unanue Emil R. , Allen Paul M.
Published: October 2002

K/BxN mice develop an inflammatory joint disease with many features characteristic of rheumatoid arthritis. In this model, the KRN transgenic T cells and nontransgenic B cells both recognize the glycolytic enzyme glucose-6-phosphate-isomerase (GPI) as an autoantigen. Here, we followed the anti-GPI B cell response that naturally arises in K/BxN mice. The anti-GPI B cell response was robust and arose at the same time as the development of serum anti-GPI autoantibody and joint inflammation. Surprisingly, although GPI was expressed systemically, the anti-GPI B cell response was focused to the lymph nodes (LN) draining the distal joints where arthritis was evident. In lymphotoxin-β receptor-Ig-treated mice, which lack LNs, the development of arthritis was completely inhibited up to 5–6 weeks. At later times, some arthritis did develop, but at a significantly reduced level. Thus, in this spontaneous model of autoimmunity, the LNs draining the distal joints are essential for both the inhibition and amplification of the arthritogenic B cell response. These findings imply that the immune physiology of a joint is unique, resulting in a local immune response to a systemic autoantigen.
An approach to membrane protein structure without crystals
Authors: Sorgen Paul L. , Hu Yonglin , Guan Lan , Kaback H. Ronald , Girvin Mark E.
Published: October 2002

The lactose permease of
Escherichia coli
catalyzes coupled translocation of galactosides and H
+
across the cell membrane. It is the best-characterized member of the Major Facilitator Superfamily, a related group of membrane proteins with 12 transmembrane domains that mediate transport of various substrates across cell membranes. Despite decades of effort and their functional importance in all kingdoms of life, no high-resolution structures have been solved for any member of this family. However, extensive biochemical, genetic, and biophysical studies on lactose permease have established its transmembrane topology, secondary structure, and numerous interhelical contacts. Here we demonstrate that this information is sufficient to calculate a structural model at the level of helix packing or better.
Carbon emissions from tropical deforestation and regrowth based on satellite observations for the 1980s and 1990s
Authors: DeFries Ruth S. , Houghton Richard A. , Hansen Matthew C. , Field Christopher B. , Skole David , Townshend John
Published: October 2002

Carbon fluxes from tropical deforestation and regrowth are highly uncertain components of the contemporary carbon budget, due in part to the lack of spatially explicit and consistent information on changes in forest area. We estimate fluxes for the 1980s and 1990s using subpixel estimates of percent tree cover derived from coarse (National Oceanic and Atmospheric Administration's Advanced Very High Resolution Radiometer) satellite data in combination with a terrestrial carbon model. The satellite-derived estimates of change in forest area are lower than national reports and remote-sensing surveys from the United Nations Food and Agriculture Organization Forest Resource Assessment (FRA) in all tropical regions, especially for the 1980s. However, our results indicate that the net rate of tropical forest clearing increased ≈10% from the 1980s to 1990s, most notably in southeast Asia, in contrast to an 11% reduction reported by the FRA. We estimate net mean annual carbon fluxes from tropical deforestation and regrowth to average 0.6 (0.3–0.8) and 0.9 (0.5–1.4) petagrams (Pg)⋅yr
−1
for the 1980s and 1990s, respectively. Compared with previous estimates of 1.9 (0.6–2.5) Pg⋅yr
−1
based on FRA national statistics of changes in forest area, this alternative estimate suggests less “missing” carbon from the global carbon budget but increasing emissions from tropical land-use change.
Tissue-specific RNA interference in postimplantation mouse embryos with endoribonuclease-prepared short interfering RNA
Authors: Calegari Federico , Haubensak Wulf , Yang Dun , Huttner Wieland B. , Buchholz Frank
Published: October 2002

RNA interference (RNAi) using double-stranded RNA has been used for the systematic analysis of gene function in invertebrate organisms. Here we have explored the use of short interfering RNA (siRNA) to knock down gene expression during the development of mammalian postimplantation embryos. The developing CNS system of embryonic day 10 mouse embryos was used as a model tissue. siRNA prepared by endoribonuclease digestion (esiRNA) was injected into the lumen of the neural tube at specific regions and delivered into neuroepithelial cells by directed electroporation. Injected and electroporated embryos were grown for 1 day in whole-embryo culture and the effects of RNAi were examined. esiRNA directed against β-galactosidase (β-gal), coelectroporated into neuroepithelial cells together with reporter plasmids expressing GFP and β-gal, abolished expression of β-gal but not GFP, showing the specificity of the esiRNA-mediated RNAi. To demonstrate RNAi of endogenous gene expression, we used heterozygous embryos of a knock-in mouse line expressing GFP from the
Tis21
locus, a gene turned on in neuroepithelial cells that switch from proliferation to neurogenesis. GFP-directed esiRNA electroporated into neuroepithelial cells of such embryos blocked the GFP expression normally occurring on the onset of neurogenesis. Taken together, our data indicate that esiRNA delivered in a tissue-specific manner by topical injection followed by directed electroporation can efficiently silence endogenous gene expression in mammalian postimplantation embryos.
The osteopetrotic mutation
toothless
(
tl
) is a loss-of-function frameshift mutation in the rat
Csf1
gene: Evidence of a crucial role for CSF-1 in osteoclastogenesis and endochondral ossification
Authors: Van Wesenbeeck Liesbeth , Odgren Paul R. , MacKay Carole A. , D'Angelo Marina , Safadi Fayez F. , Popoff Steven N. , Van Hul Wim , Marks Sandy C.
Published: October 2002

The
toothless
(
tl
) mutation in the rat is a naturally occurring, autosomal recessive mutation resulting in a profound deficiency of bone-resorbing osteoclasts and peritoneal macrophages. The failure to resorb bone produces severe, unrelenting osteopetrosis, with a highly sclerotic skeleton, lack of marrow spaces, failure of tooth eruption, and other pathologies. Injections of CSF-1 improve some, but not all, of these. In this report we have used polymorphism mapping, sequencing, and expression studies to identify the genetic lesion in the
tl
rat. We found a 10-base insertion near the beginning of the open reading of the
Csf1
gene that yields a truncated, nonfunctional protein and an early stop codon, thus rendering the
tl
rat CSF-1

null

. All mutants were homozygous for the mutation and all carriers were heterozygous. No CSF-1 transcripts were identified in rat mRNA that would avoid the mutation via alternative splicing. The biology and actions of CSF-1 have been elucidated by many studies that use another naturally occurring mutation, the
op
mouse, in which a single base insertion also disrupts the reading frame. The
op
mouse has milder osteoclastopenia and osteopetrosis than the
tl
rat and recovers spontaneously over the first few months of life. Thus, the
tl
rat provides a second model in which the functions of CSF-1 can be studied. Understanding the similarities and differences in the phenotypes of these two models will be important to advancing our knowledge of the many actions of CSF-1.
Molecular sequelae of proteasome inhibition in human multiple myeloma cells
Authors: Mitsiades Nicholas , Mitsiades Constantine S. , Poulaki Vassiliki , Chauhan Dharminder , Fanourakis Galinos , Gu Xuesong , Bailey Charles , Joseph Marie , Libermann Towia A. , Treon Steven P. , Munshi Nikhil C. , Richardson Paul G. , Hideshima Teru , Anderson Kenneth C.
Published: October 2002

The proteasome inhibitor PS-341 inhibits IκB degradation, prevents NF-κB activation, and induces apoptosis in several types of cancer cells, including chemoresistant multiple myeloma (MM) cells. PS-341 has marked clinical activity even in the setting of relapsed refractory MM. However, PS-341-induced apoptotic cascade(s) are not yet fully defined. By using gene expression profiling, we characterized the molecular sequelae of PS-341 treatment in MM cells and further focused on molecular pathways responsible for the anticancer actions of this promising agent. The transcriptional profile of PS-341-treated cells involved down-regulation of growth/survival signaling pathways, and up-regulation of molecules implicated in proapoptotic cascades (which are both consistent with the proapoptotic effect of proteasome inhibition), as well as up-regulation of heat-shock proteins and ubiquitin/proteasome pathway members (which can correspond to stress responses against proteasome inhibition). Further studies on these pathways showed that PS-341 decreases the levels of several antiapoptotic proteins and triggers a dual apoptotic pathway of mitochondrial cytochrome
c
release and caspase-9 activation, as well as activation of Jun kinase and a Fas/caspase-8-dependent apoptotic pathway [which is inhibited by a dominant negative (decoy) Fas construct]. Stimulation with IGF-1, as well as overexpression of Bcl-2 or constitutively active Akt in MM cells also modestly attenuates PS-341-induced cell death, whereas inhibitors of the BH3 domain of Bcl-2 family members or the heat-shock protein 90 enhance tumor cell sensitivity to proteasome inhibition. These data provide both insight into the molecular mechanisms of antitumor activity of PS-341 and the rationale for future clinical trials of PS-341, in combination with conventional and novel therapies, to improve patient outcome in MM.