bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification in a pseudoknot of a U⋅G motif essential for the regulation of the expression of ribosomal protein S15
Authors: Bénard Lionel , Mathy Nathalie , Grunberg-Manago Marianne , Ehresmann Bernard , Ehresmann Chantal , Portier Claude
Published: July 2002

The ribosomal protein S15 from
Escherichia coli
binds to a pseudoknot in its own messenger. This interaction is an essential step in the mechanism of S15 translational autoregulation. In a previous study, a recognition determinant for S15 autoregulation, involving a U⋅G wobble pair, was located in the center of stem I of the pseudoknot. In this study, an extensive mutagenesis analysis has been conducted in and around this U⋅G pair by comparing the effects of these mutations on the expression level of S15. The results show that the U⋅G wobble pair cannot be substituted by A⋅G, C⋅A, A⋅C, G⋅U, or C⋅G without loss of the autocontrol. In addition, the base pair C⋅G, adjacent to the 5′ side of U, cannot be flipped or changed to another complementary base pair without also inducing derepression of translation. A unique motif, made of only two adjacent base pairs, U⋅G/C⋅G, is essential for S15 autoregulation and is presumably involved in direct recognition by the S15 protein.
Recombinant viruses expressing a human malaria antigen can elicit potentially protective immune CD8+responses in mice
Authors: Miyahira Y. , García-Sastre A. , Rodriguez D. , Rodriguez J. R. , Murata K. , Tsuji M. , Palese P. , Esteban M. , Zavala F. , Nussenzweig R. S.
Published: July 2002

Extensive studies on protective immunity to rodent malaria provided the basis for the current experiments in which mice were immunized with recombinant (re) influenza and vaccinia viruses expressing selected sequences of the circumsporozoite (CS) protein of the human malaria parasitePlasmodiumfalciparum. Mice of different H-2 haplotypes immunized with re influenza viruses expressing the immunodominant B cell epitope of this CS protein produced high titers of antibodies to the parasite. A cytotoxic T lymphocyte epitope of the CS protein ofP. falciparum, PF3, recognized by CD8+T cells of H-2kmice, was expressed in a re vaccinia virus (VacPf) and a re influenza virus (FluPf). Immunization of mice with either FluPf or VacPf elicited a modest CS-specific CD8+T cell response detected by interferon γ secretion of individual immune cells. Priming of mice with FluPf, followed by a booster with VacPf, resulted in a striking enhancement of this T cell response. The reverse protocol, i.e., priming with VacPf followed by a booster with FluPf, failed to enhance the primary response. VacPf also greatly enhanced the primary response of mice injected withP. falciparumsporozoites or with a lipopeptide containing PF3. A booster with FluPf also amplified the response of lipopeptide- or sporozoite-primed mice but less than a VacPf booster did. Although mice are not susceptible to infection byP. falciparumsporozoites, we demonstrated that administration of two distinct immunogens expressing PF3 elicited activated, extravasating CS-specific T cells that protected against an intracerebral VacPf challenge.
DNA strand separation during activation of a developmental promoter by the
Bacillus subtilis
response regulator Spo0A
Authors: Rowe-Magnus Dean A. , Spiegelman George B.
Published: July 2002

Spo0A is the central regulator of commitment to sporulation in
Bacillus subtilis
. Spo0A is a member of the response regulator family of proteins and both represses and stimulates transcription from promoters when activated.
In vivo
Spo0A activation takes place by phosphorylation and
in vitro
activation can be accomplished by phosphorylation or removal of the N-terminal domain of the protein. We have examined the mechanism of Spo0A stimulation of transcription from the promoter of the
spoIIG
operon. This operon encodes one of the first compartment specific sigma factors whose appearance regulates sporulation development. When activated Spo0A was incubated with RNA polymerase and a DNA fragment containing the
spoIIG
promoter, bases between −13 and −3, relative to the start site of transcription, were denatured. Addition of activated Spo0A or RNA polymerase alone did not induce denaturation. Heteroduplex templates that contained the nontemplate sequence of the wild-type promoter on both strands between positions −3 and −13 were efficiently transcribed without activated Spo0A. These data suggest that DNA strand separation is a two-step process and that the activation of Spo0A creates a form that interacts with the polymerase to induce the first of the two steps.
A normal β-globin allele as a modifier gene ameliorating the severity of α-thalassemia in mice
Authors: Leder Aya , Wiener Edith , Lee Matthew J. , Wickramasinghe Sunitha N. , Leder Philip
Published: July 2002

Thalassemia is a heritable human anemia caused by a variety of mutations that affect expression of the α- or the β-chain of hemoglobin. The expressivity of the phenotype is likely to be influenced by unlinked modifying genes. Indeed, by using a mouse model of α-thalassemia, we find that its phenotype is strongly influenced by the genetic background in which the α-thalassemia mutation resides [129
sv/ev
/129
sv/ev
(severe) or 129
sv/ev
/C57BL/6 (mild)]. Linkage mapping indicates that the modifying gene is very tightly linked to the β-globin locus (Lod score = 13.3). Furthermore, the severity of the phenotype correlates with the size of β-chain-containing inclusion bodies that accumulate in red blood cells and likely accelerate their destruction. The β-major globin chains encoded by the two strains differ by three amino acids, one of which is a glycine-to-cysteine substitution at position 13. The Cys-13 should be available for interchain disulfide bridging and consequent aggregation between excess β-chains. This normal polymorphic variation between murine β-globin chains could account for the modifying action of the unlinked β-globin locus. Here, the variation in severity of the phenotype would not depend on a change in the ratio between α- and β-chains but on the chemical nature of the normal β-chain, which is in excess. This work also indicates that modifying genes can be normal variants that—absent an apparent physiologic rationale—may be difficult to identify on the basis of structure alone.
Site-directed hydroxyl radical probing of 30S ribosomal subunits by using Fe(II) tethered to an interruption in the 16S rRNA chain
Authors: Samaha Raymond R. , Joseph Simpson , O’Brien Bonnie , O’Brien Thomas W. , Noller Harry F.
Published: July 2002

Two
in vitro
transcripts, one corresponding to the 5′ and central domains (residues 1–920) of 16S rRNA and the other corresponding to its 3′ domain (residues 922–1542), assemble efficiently in
trans
with 30S ribosomal proteins to form a compact ribonucleoprotein particle that cosediments with natural 30S subunits. Isolated particles are similar in appearance to natural 30S subunits with electron microscopy and contain a full complement of the small subunit ribosomal proteins. The particles have a reduced ability to bind tRNA (attributable to the location of the discontinuity in a conserved region of the rRNA) near features that have been implicated in tRNA binding. Association of these two halves of 16S rRNA in trans must be stabilized by either previously unidentified RNA–RNA contacts or interactions mediated by ribosomal proteins because there are no known direct interactions between them. The
trans
construct was used to probe the three-dimensional RNA neighborhood around position 922 of 16S rRNA by generating hydroxyl radicals from Fe(II) tethered to the 5′ end of the 3′ transcript. Hydroxyl radical-induced cuts in the 16S rRNA chain were localized by primer extension to nucleotides 923–929 and 1192–1198, providing evidence for the mutual proximity of the 920 and 1192 regions.
Permeant ion regulation of
N
-methyl-
d
-aspartate receptor channel block by Mg
2+
Authors: Antonov Sergei M. , Johnson Jon W.
Published: July 2002

Block of the channel of
N
-methyl-
d
-aspartate (NMDA) receptors by external Mg
2+
(Mg
o
2+
) has broad implications for the many physiological and pathological processes that depend on NMDA receptor activation. An essential property of channel block by Mg
o
2+
is its powerful voltage dependence. A widely cited explanation for the strength of the voltage dependence of block is that the Mg
o
2+
-binding site is located deep in the channel of NMDA receptors; Mg
o
2+
then would sense most of the membrane potential field during block. However, recent electrophysiological and mutagenesis studies suggest that the blocking site cannot be deep enough to account for the voltage dependence of Mg
o
2+
block. Here we describe the basis for this discrepancy: the magnitude and voltage dependence of channel block by Mg
o
2+
are strongly regulated by external and internal permeant monovalent cations. Our data support a model in which access to the channel by Mg
o
2+
is prevented when permeant ion-binding sites at the external entrance to the channel are occupied. Mg
o
2+
can block the channel only when the permeant ion-binding sites are unoccupied and then can either unblock back to the external solution or permeate the channel. Unblock to the external solution is prevented if external permeant ions bind while Mg
2+
blocks the channel, although permeation is still permitted. The model provides an explanation for the strength of the voltage dependence of Mg
o
2+
block and quantifies the interdependence of permanent and blocking ion binding to NMDA receptors.
Volatile buffers can override the “pH memory” of subtilisin catalysis in organic media
Authors: Zacharis Evagelos , Halling Peter J. , Rees D. Gareth
Published: July 2002

The protonation state and activity of enzymes in low-water media
are affected by the aqueous pH before drying (“pH memory”).
However, both protonation and activity will change if buffer ions can
be removed as volatile or organic-extractable weak acids or bases. With
NH
4
OOCH buffers, in which both ions can be removed, pH
memory disappears completely for subtilisin-catalyzed
transesterification in hexane. Only weak pH memory is found with
buffers having one volatile component, NH
4
-phosphate and
NaOOCH. The changes in ionization state result from proton exchanges
like Protein-COO
−
NH
4
+
→ Protein-COOH +
NH
3
(g) and Protein-NH
3
+
HCOO
−
→ Protein-NH
2
+ HOOCH (g). An equivalent, complementary
picture is that net charges on the protein and buffer ions must remain
equal and opposite. With NaOOCH buffers, loss of some
HCOO
−
ions gives a more negative net charge on the
protein, balanced by the excess Na
+
. With
NH
4
-phosphate buffers, loss of NH
3
gives
protein with a more positive net charge. The resulting catalytic
activities were high and low, respectively, similar to those after
drying from Na-phosphate buffers of optimal (8.5) and acid pH. All of
the above effects have been demonstrated for both covalently
immobilized subtilisin and the lyophilized free enzyme. Subtilisin
lyophilized from NH
4
OOCH buffers gave pH ≈4 after
redissolution in water, probably because removal of HCOO
−
counterions remains incomplete. The resulting catalytic activity
was low. The effects are discussed in relation to the possible
locations, in low-dielectric media, of the positive charge that
balances the net negative catalytic triad in active subtilisin.
Folding pathway of a lattice model for proteins
Authors: Pande Vijay S. , Rokhsar Daniel S.
Published: July 2002

The folding of a protein-like heteropolymer is studied by using
direct simulation of a lattice model that folds rapidly to a
well-defined “native” structure. The details of each molecular
folding event depend on the random initial conformation as well as the
random thermal fluctuations of the polymer. By analyzing the
statistical properties of hundreds of folding events, a classical
folding “pathway” for such a polymer is found that includes
partially folded, on-pathway intermediates that are shown to be
metastable equilibrium states of the polymer. These results are
discussed in the context of the “classical” and “new” views of
folding.
Dynamics of ATP-induced Calcium Signaling in Single Mouse Thymocytes
Authors: Ross Paul E. , Ehring George R. , Cahalan Michael D.
Published: July 2002

Extracellular ATP (ATPo) elicits a robust change in the concentration of intracellular Ca2+ ([Ca2+]i) in fura-2–loaded mouse thymocytes. Most thymocytes (60%) exposed to ATPo exhibited a biphasic rise in [Ca2+]i; [Ca2+]i rose slowly at first to a mean value of 260 nM after 163 s and then increased rapidly to a peak level of 735 nM. In many cells, a declining plateau, which lasted for more than 10 min, followed the crest in [Ca2+]i. Experiments performed in the absence of extracellular [Ca2+]o abolished the rise in thymocyte [Ca2+]i, indicating that Ca2+ influx, rather than the release of stored Ca2+, is stimulated by ATPo. ATPo- mediated Ca2+ influx was potentiated as the [Mg2+]o was reduced, confirming that ATP4− is the active agonist form. In the absence of Mg2+o, 3′-O-(4-benzoyl)benzoyl-ATP (BzATP) proved to be the most effective agonist of those tested. The rank order of potency for adenine nucleotides was BzATP4−>ATP4−>MgATP2−>ADP3−, suggesting purinoreceptors of the P2X7/P2Z class mediate the ATPo response. Phenotyping experiments illustrate that both immature (CD4−CD8−, CD4+CD8+) and mature (CD4+CD8−, CD4−CD8+) thymocyte populations respond to ATP. Further separation of the double-positive population by size revealed that the ATPo-mediated [Ca2+]i response was much more pronounced in large (actively dividing) than in small (terminally differentiated) CD4+CD8+ thymocytes. We conclude that thymocytes vary in sensitivity to ATPo depending upon the degree of maturation and suggest that ATPo may be involved in processes that control cellular differentiation within the thymus.
Cytosolic Phosphorylation of Calnexin Controls Intracellular Ca2+ Oscillations via an Interaction with Serca2b
Authors: Roderick H. Llewelyn , Lechleiter James D. , Camacho Patricia
Published: July 2002

Calreticulin (CRT) and calnexin (CLNX) are lectin chaperones that participate in protein folding in the endoplasmic reticulum (ER). CRT is a soluble ER lumenal protein, whereas CLNX is a transmembrane protein with a cytosolic domain that contains two consensus motifs for protein kinase (PK) C/proline- directed kinase (PDK) phosphorylation. Using confocal Ca2+ imaging in Xenopus oocytes, we report here that coexpression of CLNX with sarco endoplasmic reticulum calcium ATPase (SERCA) 2b results in inhibition of intracellular Ca2+ oscillations, suggesting a functional inhibition of the pump. By site-directed mutagenesis, we demonstrate that this interaction is regulated by a COOH-terminal serine residue (S562) in CLNX. Furthermore, inositol 1,4,5-trisphosphate– mediated Ca2+ release results in a dephosphorylation of this residue. We also demonstrate by coimmunoprecipitation that CLNX physically interacts with the COOH terminus of SERCA2b and that after dephosphorylation treatment, this interaction is significantly reduced. Together, our results suggest that CRT is uniquely regulated by ER lumenal conditions, whereas CLNX is, in addition, regulated by the phosphorylation status of its cytosolic domain. The S562 residue in CLNX acts as a molecular switch that regulates the interaction of the chaperone with SERCA2b, thereby affecting Ca2+ signaling and controlling Ca2+-sensitive chaperone functions in the ER.
Reduced Incidence and Severity of Antigen-induced Autoimmune Diseases in Mice Lacking Interferon Regulatory Factor-1
Authors: Tada Yoshifumi , Ho Alexandra , Matsuyama Toshifumi , Mak Tak W.
Published: July 2002

Interferon regulatory factor-1 (IRF-1) is a transcription factor that regulates interferon-induced genes and type I interferons. Recently, studies of IRF-1-deficient mice have revealed that IRF-1 regulates the induction of molecules that play important roles in inflammation, such as inducible nitric oxide synthase (iNOS) and interleukin-1β-converting enzyme (ICE). To study the role of IRF-1 in autoimmunity, we investigated type II collagen-induced arthritis (CIA), and experimental allergic encephalomyelitis (EAE), in mice lacking IRF-1. The incidence and severity of CIA were significantly decreased in IRF-1−/− mice compared with IRF-1+/− mice, as was the production of interferon (IFN)-γ in lymph node cells. Both IRF-1+/− and IRF-1−/− mice exhibited mild and transient disease after adoptive transfer of a type II collagen (CII)-specific T cell line together with sera from arthritic mice, but the IRF-1−/− mice were less severely affected than the IRF-1+/− mice. In addition, the incidence of EAE in IRF-1−/− mice was decreased as compared with IRF-1+/− mice. Reverse transcription polymerase chain reaction showed that IRF-1 mRNA was constitutively expressed in the spinal cords of IRF-1+/− mice, and was upregulated in mice with clinical EAE. Expression of iNOS was also detected in inflamed spinal cords. These results suggest that IRF-1 plays a key role in promoting inflammation and autoimmunity in CIA and EAE animal models.
Disruption of Lymphocyte Function and Signaling in CD45–associated Protein–null Mice
Authors: Matsuda Akio , Motoya Satoshi , Kimura Shioko , McInnis Renee , Maizel Abby L. , Takeda Akiko
Published: July 2002

CD45-AP specifically associates with CD45, a protein tyrosine phosphatase essential for lymphocyte differentiation and antigen receptor–mediated signal transduction. CD45 is thought to mediate antigen receptor signaling by dephosphorylating regulatory tyrosine residues on Src family protein tyrosine kinases such as Lck. However, the mechanism for regulating CD45 protein tyrosine phosphatase activity remains unclear. CD45-AP–null mice were created to examine the role of CD45-AP in CD45-mediated signal transduction. T and B lymphocytes showed reduced proliferation in response to antigen receptor stimulation. Both mixed leukocyte reaction and cytotoxic T lymphocyte functions of T cells were also markedly decreased in CD45-AP–null mice. Interestingly, the interaction between CD45 and Lck was significantly reduced in CD45-AP–null T cells, indicating that CD45-AP directly or indirectly mediates the interaction of CD45 with Lck. Our data indicate that CD45-AP is required for normal antigen receptor signaling and function in lymphocytes.
Self-Inhibition in Amiloride-sensitive Sodium Channels in Taste Receptor Cells
Authors: Gilbertson Timothy A. , Zhang Huai
Published: July 2002

Electrophysiological recording techniques were used to study the Na+ dependence of currents through amiloride-sensitive sodium channels (ASSCs) in rat taste cells from the fungiform and vallate papillae. Perforated patch voltage clamp recordings were made from isolated fungiform and vallate taste receptor cells (TRCs) and Na+ transport was measured across lingual epithelia containing fungiform or vallate taste buds in a modified Ussing chamber. In isolated fungiform TRCs that contain Na+ currents sensitive to the diuretic amiloride, Na+ ions inhibit their own influx through ASSCs, a process known as sodium self-inhibition. Due to the interaction between self-inhibition and the driving force for Na+ entry, self-inhibition is most evident in whole-cell recordings at Na+ concentrations from 50 to 75 mM. In amiloride-sensitive cells, the Na permeability is significantly higher in extracellular solutions containing 35 mM Na+ than in 70 or 140 mM Na+. Compared with the block by amiloride, the development of self-inhibition is slow, taking up to 15 s to become maximally inhibited. Approximately one third of fungiform TRCs and all vallate TRCs lack functional ASSCs. These amiloride-insensitive TRCs show no signs of self-inhibition, tying this phenomenon to the presence of ASSCs. The sulfhydryl reagent, p-hydroxymercuribenzoate (p-HMB; 200 μM), reversibly removed self-inhibition from amiloride-sensitive Na+ currents, apparently by modifying cysteine residues in the ASSC. Na+ currents in amiloride-insensitive TRCs were unaffected by p-HMB. In sodium transport studies in fungiform taste bud–containing lingual epithelia, ∼40% of the change in short-circuit current (Isc) after addition of 500 mM NaCl to the mucosal chamber is amiloride sensitive (0.5 mM). p-HMB significantly enhanced mucosal NaCl-induced changes in these epithelia at mucosal Na+ concentrations of 50 mM and above. In contrast, the vallate-containing epithelia, which are insensitive to amiloride, showed no enhancement of Isc during p-HMB treatment. These findings suggest that sodium self-inhibition is present in ASSCs in taste receptor cells where it may play a crucial role in performance of salt-sensitive pathways in taste tissue during sodium stimulation. This phenomenon may be important in the process of TRC adaptation, in the conservation of cellular resources during chronic sodium exposure, or in the gustatory response to water.
Non-reflecting boundary conditions for waveguides
Authors: Bendali A. , Guillaume Ph.
Published: July 2002

New non-reflecting boundary conditions are introduced for the solution of the Helmholtz equation in a waveguide. These boundary conditions are perfectly transparent for all propagating modes. They do not require the determination of these propagating modes but only their propagation constants. A quasi-local form of these boundary conditions is well suited as terminating boundary condition beyond finite element meshes. Related convergence properties to the exact solution and optimal error estimates are established.
SEGMENT: identifying compositional domains in DNA sequences
Authors: L.Oliver José , Román-Roldán Ramón , Pérez Javier , Bernaola-Galván Pedro
Published: July 2002

Motivation: DNA sequences are formed by patches or domains of different nucleotide composition. In a few simple sequences, domains can simply be identified by eye; however, most DNA sequences show a complex compositional heterogeneity (fractal structure), which cannot be properly detected by current methods. Recently, a computationally efficient segmentation method to analyse such nonstationary sequence structures, based on the Jensen–Shannon entropic divergence, has been described. Specific algorithms implementing this method are now needed.

Results: Here we describe a heuristic segmentation algorithm for DNA sequences, which was implemented on a Windows program (SEGMENT). The program divides a DNA sequence into compositionally homogeneous domains by iterating a local optimization procedure at a given statistical significance. Once a sequence is partitioned into domains, a global measure of sequence compositional complexity (SCC), accounting for both the sizes and compositional biases of all the domains in the sequence, is derived. SEGMENT computes SCC as a function of the significance level, which provides a multiscale view of sequence complexity.

Availability: SEGMENT is available on the Web at http://www.ugr.es/local/oliver/segment/

Contact: oliver@ugr.es
Long-term consequences for offspring of diabetes during pregnancy
Authors: Van Assche Frans A , Holemans Kathleen , Aerts Leona
Published: July 2002

There is evidence that the diabetic intra-uterine environment has consequences for later life. Maternal diabetes mainly results in asymmetric macrosomia. This macrosomia is associated with an increased insulin secretion and overstimulation of the insulin producing B-cells during fetal life. In later life, a reduced insulin secretion is found. Intra-uterine growth restriction is present in severe maternal diabetes associated with vasculopathy. Intra-uterine growth restriction is associated with low insulin secretion and reduced development of the insulin receptors. In later life, these alterations can induce insulin resistance. The long-term consequences of an abnormal intra-uterine environment are of primary importance world-wide. Concentrated efforts are needed to explore how these long-term effects can be prevented.
Masked antisense: a molecular configuration for discriminating similar RNA targets
Authors: Stocks Martin R , Rabbitts Terence H
Published: July 2002

Antisense technology has great potential for the control of RNA expression, but there remain few successful applications of the technology. Expressed antisense RNA can effectively down‐regulate expression of a gene over long periods, but cannot differentiate partly identical sequences, such as the mRNA of fusion genes or those with point mutants. We have designed a structured form of expressed antisense, which can discriminate between highly similar mRNA molecules. These 'masked' antisense RNAs have most of the antisense sequence sequestered within duplex elements, leaving a short single‐stranded region to initiate binding to target RNA. After contacting the correct target, the structured RNA can unravel, releasing the masked antisense region to form a stable duplex with the mRNA. We demonstrate that suitable masked antisense RNA can discriminate between the two forms of BCR–ABL mRNA that result from the Philadelphia chromosomal translocations, as well as discriminating the normal BCR and ABL mRNA.
Uncoupling yeast intron recognition from transcription with recursive splicing
Authors: Lopez Pascal J , Séraphin Bertrand
Published: July 2002

Pre‐mRNA splicing has to be coordinated with other processes occurring in the nucleus including transcription, mRNA 3′ end formation and mRNA export. To analyze the relationship between transcription and splicing, we constructed a network of nested introns. Introns were inserted in the 5′ splice site and/or branchpoint of a synthetic yeast intron interrupting a reporter gene. The inserted introns mask the recipient intron from the cellular machinery until they are removed by splicing. Production of functional mRNA from these constructs therefore requires recognition of a spliced RNA as a splicing substrate. We show that recurrent splicing occurs in a sequential and ordered fashion in vivo. Thus, in Saccharomyces cerevisiae, intron recognition and pre‐spliceosome assembly is not tightly coupled to transcription.
Chromosomal DNA demethylation specified by protein binding
Authors: Lin Iping G , Hsieh Chih‐Lin
Published: July 2002

In the present study, we utilize the well‐characterized Escherichia coli lac repressor/operator system to demonstrate that protein binding can lead to demethylation at the binding sites in the chromosome. Similar to the findings using the episome, we found that the presence of LacI in the cells can lead to demethylation of methylated lacO in the chromosome and the LacI inhibitor, isopropyl‐β‐D‐thiogalactopyranoside (IPTG), can prevent demethylation of the methylated lacO. The lacO sites become progressively more demethylated over time with the presence of LacI, supporting the role of protein occupancy in demethylation targeting. These results validate our earlier conclusions using a stable episomal system, and establish for the first time that protein binding can specify sites of demethylation in the chromosome.
RNA and sex determination in Caenorhabditis elegans
Authors: Puoti Alessandro , Pugnale Paolo , Belfiore Marco , Schläppi Anne‐Catherine , Saudan Zarifja
Published: July 2002

The Caenorhabditis elegans hermaphrodite sequentially produces sperm and oocytes from a single pool of precursors. Therefore, the hermaphrodite's germ line is the site of two major cell fate decisions: a germ cell precursor first undergoes a mitosis/meiosis decision and then a sperm/oocyte decision. While the mitosis/meiosis decision is governed by Notch/GLP‐1 signalling, the sperm/oocyte decision relies on post‐transcriptional regulation of two key mRNAs, tra‐2 and fem‐3. This review focuses on factors that are required for the silencing of these mRNAs, which results in the sequential production of sperm and oocytes. Most factors that regulate the expression of tra‐2 and fem‐3 are homologous to proteins involved in RNA regulation in yeast, mammals or Drosophila, suggesting that at least some of the molecular mechanisms regulating the two worm mRNAs have been conserved throughout evolution.
Position‐effect variegation in Drosophila: the modifier Su(var)3‐7 is a modular DNA‐binding protein
Authors: Cléard Fabienne , Spierer Pierre
Published: July 2002

An increase in the dose of the Su(var)3‐7 locus of Drosophila augments heterochromatin‐promoted variegated silencing. The deduced protein sequence of Su(var)3‐7 reveals seven widely spaced zinc fingers. We found that Su(var)3‐7 has affinity for DNA in vitro and that the minimal protein sequence requirement for DNA binding is any module containing two zinc fingers and the interval between them. As Su(var)3‐7 is a heterochromatin‐associated protein, we tested its affinity for various satellite DNA sequences in vitro. The AATAT and 353‐bp elements have the highest affinity. If affinity for satellite DNAs contributes to the presence of Su(var)3‐7 in heterochromatin, a general affinity for DNA, or sequences yet to be determined, suggests a function in the genomic silencing of position‐effect variegation: expansion of heterochromatin, whether continuous by spreading or discontinuous by pairing with sequence elements scattered through euchromatin, could use the affinity of Su(var)3‐7 for DNA.
Selection of targets and the most efficient hairpin ribozymes for inactivation of mRNAs using a self‐cleaving RNA library
Authors: Barroso‐delJesus Alicia , Berzal‐Herranz Alfredo
Published: July 2002

The identification of proficient target sites within long RNA molecules, as well as the most efficient ribozymes for each, is a major concern for the use of ribozymes as gene suppressers. In vitro selection methods using combinatorial libraries are powerful tools for the rapid elucidation of interactions between macromolecules, and have been successfully used for different types of ribozyme study. This paper describes a new method for selecting effective target sites within long RNAs using a combinatorial library of self‐cleaving hairpin ribozymes that includes all possible specificities. The method also allows the identification of the most appropriate ribozyme for each identified site. Searching for targets within the lacZ gene with this strategy yielded a clearly accessible site. Sequence analysis of ribozymes identified two variants as the most appropriate for this site. Both selected ribozymes showed significant inhibitory activity in the cell milieu.
When repair meets chromatin
Authors: Green Catherine M , Almouzni Geneviève
Published: July 2002

In eukaryotic cells, the inheritance of both the DNA sequence and its organization into chromatin is critical to maintain genome stability. This maintenance is challenged by DNA damage. To fully understand how the cell can tolerate genotoxic stress, it is necessary to integrate knowledge of the nature of DNA damage, its detection and its repair within the chromatin environment of a eukaryotic nucleus. The multiplicity of the DNA damage and repair processes, as well as the complex nature of chromatin, have made this issue difficult to tackle. Recent progress in each of these areas enables us to address, both at a molecular and a cellular level, the importance of inter‐relationships between them. In this review we revisit the ‘access, repair, restore’ model, which was proposed to explain how the conserved process of nucleotide excision repair operates within chromatin. Recent studies have identified factors potentially involved in this process and permit refinement of the basic model. Drawing on this model, the chromatin alterations likely to be required during other processes of DNA damage repair, particularly double‐strand break repair, are discussed and recently identified candidates that might perform such alterations are highlighted.
A brief history of human autosomes
Authors: Haig David
Published: July 2002

Comparative gene mapping and chromosome painting permit the tentative reconstruction of ancestral karyotypes. The modern human karyotype is proposed to differ from that of the most recent common ancestor of catarrhine primates by two major rearrangements. The first was the fission of an ancestral chromosome to produce the homologues of human chromosomes 14 and 15. This fission occurred before the divergence of gibbons from humans and other apes. The second was the fusion of two ancestral chromosomes to form human chromosome 2. This fusion occurred after the divergence of humans and chimpanzees. Moving further back in time, homologues of human chromosomes 3 and 21 were formed by the fission of an ancestral linkage group that combined loci of both human chromosomes, whereas homologues of human chromosomes 12 and 22 were formed by a reciprocal translocation between two ancestral chromosomes. Both events occurred at some time after our most recent common ancestor with lemurs. Less direct evidence suggests that the short and long arms of human chromosomes 8, 16 and 19 were unlinked in this ancestor. Finally, the most recent common ancestor of primates and artiodactyls is proposed to have possessed a chromosome that combined loci from human chromosomes 4 and 8p, a chromosome that combined loci from human chromosomes 16q and 19q, and a chromosome that combined loci from human chromosomes 2p and 20.
Searching for kinesin's mechanical amplifier
Authors: Vale Ronald D. , Case Ryan , Sablin Elena , Hart Cindy , Fletterick Robert
Published: July 2002

Kinesin, a microtubule–based motor, and myosin, an actin–based motor, share a similar core structure, indicating that they arose from a common ancestor. However, kinesin lacks the long lever–arm domain that is believed to drive the myosin power stroke. Here, we present evidence that a much smaller region ofca. 10–40 amino acids serves as a mechanical element for kinesin motor proteins. These ‘neck regions’ are class conserved and have distinct structures in plus–end and minus–end–directed kinesin motors. Mutagenesis studies also indicate that the neck regions are involved in coupling ATP hydrolysis and energy into directional motion along the microtubule. We suggest that the kinesin necks drive motion by undergoing a conformational change in which they detach and re–dock onto the catalytic core during the ATPase cycle. Thus, kinesin and myosin have evolved unique mechanical elements that amplify small, nucleotide–dependent conformational changes that occur in their similar catalytic cores.
The roles of turn formation and cross‐strand interactions in fibrillization of peptides derived from the OspA single‐layer β‐sheet
Authors: Ohnishi Satoshi , Koide Akiko , Koide Shohei
Published: July 2002

We previously demonstrated that a β‐hairpin peptide, termed BH9–10, derived from a single‐layer β‐sheet of Borrelia OspA protein, formed a native‐like β‐turn in trifluoroethanol (TFE) solution, and it assembled into amyloid‐like fibrils at higher TFE concentrations. This peptide is highly charged, and fibrillization of such a hydrophilic peptide is quite unusual. In this study, we designed a circularly permutated peptide of BH9–10, termed BH10–9. When folded into their respective β‐hairpin structures found in OspA, these peptides would have identical cross‐strand interactions but different turns connecting the strands. NMR study revealed that BH10–9 had little propensity to form a turn structure both in aqueous and TFE solutions. At higher TFE concentration, BH10–9 precipitated with a concomitant α‐to‐β conformational conversion, in a similar manner to the BH9–10 fibrillization. However, the BH10–9 precipitates were nonfibrillar aggregation. The precipitation kinetics of BH10–9 was exponential, consistent with a first‐order molecular assembly reaction, while the fibrillization of BH9–10 showed sigmoidal kinetics, indicative of a two‐step reaction consisting of nucleation and molecular assembly. The correlation between native‐like turn formation and fibrillization of our peptide system strongly suggests that BH9–10 adopts a native‐like β‐hairpin conformation in the fibrils. Remarkably, seeding with the preformed BH10–9 precipitates changed the two‐step BH9–10 fibrillization to a one‐step molecular assembly reaction, and disrupted the BH9–10 fibril structure, indicating interactions between the BH10–9 aggregates and the BH9–10 peptide. Our results suggest that, in these peptides, cross‐strand interactions are the driving force for molecular assembly, and turn formation limits modes of peptide assembly.
Molecular dynamics simulation of Escherichia coli dihydrofolate reductase and its protein fragments: Relative stabilities in experiment and simulations
Authors: Sham Yuk Yin , Ma Buyong , Tsai Chung‐Jung , Nussinov Ruth
Published: July 2002

We have carried out molecular dynamics simulations of the native dihydrofolate reductase from Escherichia coli and several of its folded protein fragments at standard temperature. The simulations have shown fragments 1–36, 37–88, and 89–159 to be unstable, with a CαRMSD (Cα root mean squared deviation) >5 Å after 3.0 nsec of simulation. The unfolding of fragment 1–36 was immediate, whereas fragments 37–88 and 89–159 gradually unfolded because of the presence of the β‐sheet core structure. In the absence of residues 1–36, the two distinct domains comprising fragment 39–159 associated with each other, resulting in a stable conformation. This conformation retained most of its native structural elements. We have further simulated fragments derived from computational protein cutting. These were also found to be unstable, with the exception of fragment 104–159. In the absence of α4, the loose loop region of residues 120–127 exhibited a β‐strand‐like behavior, associating itself with the β‐sheet core of the protein fragment. The current study suggests that the folding of dihydrofolate reductase involves cooperative folding of distinct domains which otherwise would have been unstable as independent folded units in solution. Finally, the critical role of residues 1–36 in allowing the two distinct domains of fragment 104–159 to fold into the final native conformation is discussed.
The active site of hydroxynitrile lyase from Prunus amygdalus: Modeling studies provide new insights into the mechanism of cyanogenesis
Authors: Dreveny Ingrid , Kratky Christoph , Gruber Karl
Published: July 2002

The FAD‐dependent hydroxynitrile lyase from almond (Prunus amygdalus, PaHNL) catalyzes the cleavage of R‐mandelonitrile into benzaldehyde and hydrocyanic acid. Catalysis of the reverse reaction—the enantiospecific formation of α‐hydroxynitriles—is now widely utilized in organic syntheses as one of the few industrially relevant examples of enzyme‐mediated C–C bond formation. Starting from the recently determined X‐ray crystal structure, systematic docking calculations with the natural substrate were used to locate the active site of the enzyme and to identify amino acid residues involved in substrate binding and catalysis. Analysis of the modeled substrate complexes supports an enzymatic mechanism that includes the flavin cofactor as a mere “spectator” of the reaction and relies on general acid/base catalysis by the conserved His‐497. Stabilization of the negative charge of the cyanide ion is accomplished by a pronounced positive electrostatic potential at the binding site. PaHNL activity requires the FAD cofactor to be bound in its oxidized form, and calculations of the pKa of enzyme‐bound HCN showed that the observed inactivation upon cofactor reduction is largely caused by the reversal of the electrostatic potential within the active site. The suggested mechanism closely resembles the one proposed for the FAD‐independent, and structurally unrelated HNL from Hevea brasiliensis. Although the actual amino acid residues involved in the catalytic cycle are completely different in the two enzymes, a common motif for the mechanism of cyanogenesis (general acid/base catalysis plus electrostatic stabilization of the cyanide ion) becomes evident.
Tissue mimicking materials for a multi‐imaging modality prostate phantom
Authors: D'Souza Warren D. , Madsen Ernest L. , Unal Orhan , Vigen Karl K. , Frank Gary R. , Thomadsen Bruce R.
Published: July 2002

Materials that simultaneously mimic soft tissue in vivo for magnetic resonance imaging (MRI), ultrasound (US), and computed tomography (CT) for use in a prostate phantom have been developed. Prostate and muscle mimicking materials contain water, agarose, lipid particles, protein, EDTA, glass beads, and thimerosal (preservative). Fat was mimicked with safflower oil suffusing a random mesh (network) of polyurethane. Phantom material properties were measured at 22 °C. (22 °C is a typical room temperature at which phantoms are used.) The values of material properties should match, as well as possible, the values for tissues at body temperature, 37 °C. For MRI, the primary properties of interest are T1 and T2 relaxations times, for US they are the attenuation coefficient, propagation speed, and backscatter, and for CT, the x‐ray attenuation. Considering the large number of parameters to be mimicked, rather good agreement was found with actual tissue values obtained from the literature. Using published values for prostate parenchyma, T1 and T2 at 37 °C and 40 MHz are estimated to be about 1100 and 98 ms, respectively. The CT number for in vivo prostate is estimated to be 45 HU (Hounsfield units). The prostate mimicking material has a T1 of 937 ms and a T2 of 88 ms at 22 °C and 40 MHz; the propagation speed and attenuation coefficient slope are 1540 m/s and 0.36 dB/cm/MHz, respectively, and the CT number of tissue mimicking prostate is 43 HU. Tissue mimicking (TM) muscle differs from TM prostate in the amount of dry weight agarose, EDTA, and the quality and quantity of glass beads. The 18 μm glass beads used in TM muscle increase US backscatter and US attenuation; the presence of the beads also has some effect on T1 but no effect on T2. The composition of tissue‐mimicking materials developed is such that different versions can be placed in direct contact with one another in a phantom with no long term change in US, MRI, or CT properties. Thus, anthropomorphic phantoms can be constructed.
The magic tesseracts and Bell’s theorem
Authors: Aravind P. K.
Published: July 2002

This paper describes a toy, consisting of a pair of magic tesseracts, that can be used to perform an amazing trick that defies explanation in conventional terms. An account of the toy, and the trick possible with it, is given in nontechnical terms that should be accessible to a layman. An explanation is then given, for physicists, of how this trick can be interpreted as a proof of Bell’s theorem. An appendix (again intended for physicists) describes the internal machinery of the toy and how it works its magic.
Simplified quantum oscillator solution
Authors: Jordan Thomas F.
Published: July 2002

To find the eigenvalues and eigenvectors of the quantum oscillator Hamiltonian, it is enough to construct eigenvectors with the raising operator and see that these eigenvectors are complete. The subspace they span must be the entire space of square-integrable functions because it is invariant for the raising and lowering operators and therefore invariant for the position and momentum operators. This shows that the Hamiltonian has no continuous spectrum.
One-dimensional laser cooling of an atomic beam in a sealed vapor cell
Authors: Vale C. J. , Walkiewicz M. R. , Farrell P. M. , Scholten R. E.
Published: July 2002

We describe a simple experiment that demonstrates one-dimensional laser cooling in a sealed vapor cell. A velocity selective optical pumping scheme was first used to define a collimated beam of atoms within the cell. A particular velocity group of atoms was labeled by optical pumping with one laser and detected by absorption with a second laser. Transverse cooling of this velocity group, which formed an atomic beam between the two laser beams, was then observed by applying a third laser beam, in analogy with transverse cooling of a conventional atomic beam.
Thermodynamics of mixtures: Functions of mixing and excess functions
Authors: Nieto R. , González M. C. , Herrero F.
Published: July 2002

Applying thermodynamics to realistic systems requires a knowledge of the thermodynamic properties of mixtures. Functions of mixing and excess functions provide a useful approach. The concepts are simple and their application straightforward, but students often fail to apply them correctly when they are given only a theoretical explanation. We discuss some typical mistakes and some problems we have found useful for overcoming them.
Thermodynamics at work: The pressure derivative of the specific heat
Authors: Güémez J. , Fiolhais C. , Fiolhais M.
Published: July 2002

Thermodynamics relates measurable quantities such as thermal coefficients and specific heats. The first law, which implies that the enthalpy is a function of state, yields a relation for the pressure derivative of the specific heat cP. The second law gives a simpler and well-known relation for this pressure derivative. We compare the values of the pressure derivative of cP obtained from the first and second laws to the values obtained from measurements for water at different pressures. The comparison illustrates the scope and methodology of thermodynamics.
A walk in phase space: Solidification into crystalline and amorphous states
Authors: Adler Joan
Published: July 2002

The formation of crystalline and amorphous solids is described using a simple glass bead demonstration and a discussion of annealing and rapid quenching on a computer.
Centripetal acceleration: Another round
Authors: Dietz Eric R.
Published: July 2002

Described in this brief note is a novel approach to the classroom discussion of the direction of the acceleration for uniform circular motion.
Two definitions of the electric polarizability of a bound system in relativistic quantum theory
Authors: Coutinho F. A. B. , Nogami Y. , Tomio Lauro
Published: July 2002

For the electric polarizability of a bound system in relativistic quantum theory, there are two definitions that have appeared in the literature. They differ depending on whether or not the vacuum background is included in the system. A recent confusion in this connection is clarified.
The Boltzmann reservoir: A model constant-temperature environment
Authors: Leff Harvey S.
Published: July 2002

The Boltzmann reservoir (BR) is a model constant-temperature environment that exhibits highly atypical thermodynamic behavior. Its microcanonical ensemble entropy is a linear, nonconcave function of its internal energy U, and its zero-work heat capacity is infinite. Its canonical partition function diverges because all possible energies are equally likely, so the microcanonical and canonical ensembles are not equivalent. If two BRs with the same temperature TB are put in thermal contact, either can have any fraction of the total energy; i.e., there is no unique equilibrium state. If two BRs with different temperatures are in thermal contact, the higher temperature BR gives all its energy to the other. A BR’s temperature cannot be changed by a heat process but, in principle, can be altered by a work process. These and other properties that challenge conventional wisdom provide thought-provoking examples for thermal physics courses.
Crater property in two-particle bound states: When and why
Authors: Chow Chi-Keung
Published: July 2002

Crater has shown that, for two particles (with masses m1 and m2) in a Coulombic bound state, the charge distribution is equal to the sum of the two charge distributions obtained by taking m1→∞ and m2→∞, respectively, while keeping the same Coulombic potential. We provide a simple scaling criterion to determine whether an arbitrary Hamiltonian possesses this property. In particular, we show that, for a Coulombic system, fine structure corrections preserve this Crater property while two-particle relativistic corrections and/or hyperfine corrections may destroy it.
To advance and diffuse the knowledge of physics: An account of the one-hundred-year history of the American Physical Society
Authors: Lustig Harry
Published: July 2002

On May 20, 1899, thirty-six physicists founded the American Physical Society. A hundred years later, 11,239 scientists—the Society had by then 41,786 members—gathered to mark both a century of physics and the centennial of the Society. How the APS came about and developed, how it pursued its mission—the advancement and diffusion of the knowledge of physics—through its meetings, journals, and public activities, who its leaders were, and what it looks like today, is the subject of this article.