bluuebunny/crossref_metadata_2025 huggingface dataset:
Efficient technique for calculating the bound and virtual energy spectrum of semiconductor structures
Authors: Schwartz Carey
Published: July 2002

A novel technique for determining the energies of bound and resonant states of semiconductor structures is presented. The new method is based upon R-matrix propagation techniques (known to be numerically stable) and avoids direct integration of the Schrödinger equation. The new method is recursive in nature, numerically stable, and can be applied easily to any one-dimensional potential. We show that the new technique yields the well-known analytic results describing the energy spectrum of a single quantum well. Further examples illustrate application of the R-matrix technique to determine the bound state and resonant energy spectrum of multiple quantum well structures in the presence or absence of an applied bias.
Surface-emitting laser diode with vertical GaAs/GaAlAs quarter-wavelength multilayers and lateral buried heterostructure
Authors: Ogura Mutsuo , Hsin Wei , Wu Ming-Chiang , Wang Shyh , Whinnery John R. , Wang S. C. , Yang Jane J.
Published: July 2002

Threshold current of 2 mA at room temperature cw operation is realized in a vertical distributed feedback surface-emitting laser diode with lateral buried heterostructure (LBH). In this LBH structure, the vertical distributed feedback active region (AlGaAs/GaAs multilayer) is entirely surrounded with n- and p-type AlGaAs cladding layers for minority-carrier confinement. The far-field angle is 7°. The beam shape is nearly circular. However, the lasing spectrum is broad (2–3 nm) compared with the conventional edge-emitting laser. Major differences between the surface-emitting laser diode presented here and the conventional edge-emitting laser diode are discussed.
High-energy elastic backscattering of helium ions for compositional analysis of high-temperature superconductor thin films
Authors: Martin J. A. , Nastasi M. , Tesmer J. R. , Maggiore C. J.
Published: July 2002

A thin-film technique for measuring the chemical composition of Y-Ba-Cu-O thin films to a few percent accuracy is described. This technique utilizes non-Rutherford backscattering of 8.8 MeV helium ions, which has an increased sensitivity 16O by a factor of 25 over Rutherford backscattering spectrometry. The ratios of the cross sections for He++ scattered from oxygen, copper, and yttrium relative to barium are easily determined using thin-film standards that can be fabricated in any deposition system capable of producing thin-film superconductors. The technique does not require the constant use of standards or accurate charge determination.
Polarons in DNA
Authors: Conwell Esther M. , Rakhmanova Svetlana V.
Published: July 2002

Many experiments have been done to determine how far and how freely holes can move along the stack of base pairs in DNA. The results of these experiments are usually described in terms of a parameter β under the assumption that it describes an exponential decay with distance. The reported values range from β < 0.2/Å to β > 1.4/Å. For the larger values of β, the transport can be accounted for as single step superexchange-mediated hole transfer. To account for the smaller values, hopping models have been proposed, the simplest being nearest-neighbor hopping. This model assumes that, between hops, the hole is localized on a single base with no overlap to neighbors. Noting that an electron or hole added to a DNA stack, as to other essentially one-dimensional entities, should distort its structure to form a polaron, Schuster and coworkers [Henderson, P. T., Jones, D., Hampikian, G., Kan, Y. & Schuster, G. B. (1999)
Proc. Natl. Acad. Sci. USA
96, 8353–8358 and Ly, D., Sanii, L. & Schuster, G. B. (1999)
J. Am. Chem. Soc.
121, 9400–9410] proposed that transport occurs by polaron hopping between sites having approximately equal energies as a result of overlap. A recent experimental determination by Wan
et al.
[Wan, C., Fiebig, T., Kelley, S. O., Treadway, C. R., Barton, J. K. & Zewail, A. H. (1999)
Proc. Natl. Acad. Sci. USA
96, 6014–6019] of the time required for an injected hole on DNA to travel a known distance leads to a large value of the diffusion constant. From this constant, a mobility of 0.2 cm
2
/V⋅s was deduced, orders of magnitude larger than typical hopping mobilities. We suggest that this ultrafast transport is due to polaron drift, which has been shown to lead to similar mobilities in chains of conjugated polymers. Using a simple model for the polaron, similar to that used for conjugated polymers such as polyacetylene, we show that, for reasonable values of the parameters, an injected electron or hole can form a polaron on a DNA stack.
Ribozymes that cleave reovirus genome segment S1 also protect cells from pathogenesis caused by reovirus infection
Authors: Shahi Shweta , Shanmugasundaram Ganapathy K. , Banerjea Akhil C.
Published: July 2002

Reovirus genome segment S1 encodes protein σ1, which is the
receptor binding protein, modulates tissue tropism, and specifies the
nature of the antiviral immune response. It makes up less than 2% of
reovirus particles and is synthesized in very small amounts in infected
cells. Any antiviral strategy aimed at reducing specifically the
expression of this genome segment should, in principle, reduce the
infectivity of the virus. To test this hypothesis, we have assembled
two hammer-head motif-containing ribozymes (Rzs) targeted to cleave at
the conserved B and C domains of the reovirus s1 RNA.
Protein-independent but Mg
2+
-dependent
sequence-specific cleavage of s1 RNA was achieved by both the Rzs
in trans. Cells that transiently express these Rzs, when challenged
with reovirus, were protected against the cytopathic effects caused by
the virus. This protection correlated with the specific intracellular
reduction of s1 transcripts that was due to their cleavage by the Rzs.
Rz-treated cells that were challenged with reovirus showed almost
complete disappearance of protein σ1 without significantly altering
the levels of the other reovirus structural proteins. Thus, Rzs,
besides acting as antiviral agents, could be exploited as biological
tools to delineate specific functions of target genes.
Logic gates using high Rydberg states
Authors: Remacle F. , Schlag E. W. , Selzle H. , Kompa K. L. , Even U. , Levine R. D.
Published: July 2002

Connected logic gates can be operated on the levels of one molecule
by making use of the special properties of high Rydberg states.
Explicit experimental results for the NO molecule are provided as an
example. A number of other options, including that of several gates
concatenated so as to operate as a full adder, are discussed. Specific
properties of high Rydberg states that are used are: their
autoionization is delayed so that they can be distinguished from direct
multiphoton ionization, during their long life such states also can
decay by energy transfer to the molecular core in a way that can be
controlled by the judicious application of very weak external
electrical fields, and the Rydberg states can be detected by the
application of an ionizing electrical field. The combination of two (or
three) color photons with and without external weak fields allows the
construction of quite elaborate logic circuit diagrams and shows that
taking advantage of the different intramolecular dynamics of levels
that differ by their excitation enables the compounding of logic
operations on one molecular frame.
The embryonic stem cell transcription factors Oct-4 and FoxD3 interact to regulate endodermal-specific promoter expression
Authors: Guo Ying , Costa Robert , Ramsey Heather , Starnes Trevor , Vance Gail , Robertson Kent , Kelley Mark , Reinbold Rolland , Scholer Hans , Hromas Robert
Published: July 2002

The POU homeodomain protein Oct-4 and the Forkhead Box protein FoxD3 (previously Genesis) are transcriptional regulators expressed in embryonic stem cells. Down-regulation of Oct-4 during gastrulation is essential for proper endoderm development. After gastrulation, FoxD3 is generally down-regulated during early endoderm formation, although it specifically remains expressed in the embryonic neural crest. In these studies, we have found that Oct-4 and FoxD3 can bind to identical regulatory DNA sequences. In addition, Oct-4 physically interacted with the FoxD3 DNA-binding domain. Cotransfection of Oct-4 and FoxD3 expression vectors activated the osteopontin enhancer, which is expressed in totipotent embryonic stem cells. FoxA1 and FoxA2 (previously HNF-3α and HNF-3β) are Forkhead Box transcription factors that participate in liver and lung formation from foregut endoderm. Although FoxD3 activated the FoxA1 and FoxA2 promoters, Oct-4 inhibited FoxD3 activation of the FoxA1 and FoxA2 endodermal promoters. These data indicate that Oct-4 functions as a corepressor of FoxD3 to provide embryonic lineage-specific transcriptional regulatory activity to maintain appropriate developmental timing.
Self-assembly directed by NH⋅⋅⋅O hydrogen bonding: New layered molecular arrays derived from 4-
tert
- butylbenzoic acid and aliphatic diamines
Authors: Armstrong Robert S. , Atkinson Ian M. , Carter Elizabeth , Mahinay Myrna S. , Skelton Brian W. , Turner Peter , Wei Gang , White Allan H. , Lindoy Leonard F.
Published: July 2002

1
H and
13
C NMR titrations in both CDCl
3
and CD
3
OD demonstrate that 4-
tert
-butylbenzoic acid interacts with both propane-1,2-diamine and propane-1,3-diamine to yield 1:2 host–guest complexes in these solvents. Based on this observation, the isolation of new three-dimensional molecular arrays through cocrystallization of the above diamines and 4-
tert
-butylbenzoic acid (in a 1:2 molar ratio) has been achieved. X-ray studies of these self-assembled structures show that they incorporate [propane-1,2-diamine⋅(4-
tert
-butylbenzoic acid)
2
] or [propane-1,3-diamine⋅(4-
tert
-butylbenzoic acid)
2
] hydrogen-bonded motifs. Three structural derivatives of the latter type (two monohydrate forms and one anhydrous form) have been characterized. The structures are compared with a previously described three-dimensional array based on the “parent” [ethane-1,2-diamine⋅(benzoic acid)
2
] motif. Similarities occur between each of the structures. In each, a two-dimensional “ionic” layer consisting of an extensive network of hydrogen bonds is sandwiched between two “less polar” aromatic ring-containing layers. In the respective ionic layers, the carboxylic acid protons have been transferred onto the amines to yield diammonium cations, with all ammonium protons being involved in hydrogen bonding. In part, the adoption of these unusual layered structures seems to reflect a tendency toward maximization of both the number and strengths of the hydrogen bond interactions in the respective ionic layers.
Selecting rRNA binding sites for the ribosomal proteins L4 and L6 from randomly fragmented rRNA: Application of a method called SERF
Authors: Stelzl Ulrich , Spahn Christian M. T. , Nierhaus Knud H.
Published: July 2002

Two-thirds of the 54 proteins of the
Escherichia coli
ribosome interact directly with the rRNAs, but the rRNA binding sites of only a very few proteins are known. We present a method (selection of random RNA fragments; SERF) that can identify the minimal binding region for proteins within ribonucleo-protein complexes such as the ribosome. The power of the method is exemplified with the ribosomal proteins L4 and L6. Binding sequences are identified for both proteins and characterized by phosphorothioate footprinting. Surprisingly, the binding region of L4, a 53-nt rRNA fragment of domain I of 23S rRNA, can simultaneously and independently bind L24, one of the two assembly initiator proteins of the large subunit.
Toward development of a screen to identify randomly encoded, foldable sequences
Authors: Hagihara Yoshihisa , Kim Peter S.
Published: July 2002

The ability to identify sequences in a randomly encoded polypeptide library that are capable of acquiring unique and stably folded structures would be valuable in the examination of protein-folding issues. The quality control system of the yeast secretory pathway prevents the release of incompletely folded polypeptides. Earlier work has shown that this feature can be used in a screen to identify mutations that increase the stability of a protein. We sought to extend this strategy for use with random sequence libraries by combining a quality-control system-based screen with generic tag-based immunodetection that can be applied to any sequence. To test this method, we screened a library encoding random mutations in a bovine pancreatic trypsin inhibitor variant containing a small generic tag. Initial on-plate screening resulted in a large number of false positives: sequences that were secreted but not foldable. These false positives were excluded successfully in additional screening steps that used a liquid-culture secretion screen and a gel electrophoresis assay. Three positive clones were obtained that showed midpoint thermal denaturation temperatures 10–16°C higher than the original bovine pancreatic trypsin inhibitor variant. Thus, this multistep screening method may be useful for finding novel, foldable sequences.
An acyltransferase catalyzing the formation of diacylglucose is a serine carboxypeptidase-like protein
Authors: Li Alice X. , Steffens John C.
Published: July 2002

1-
O
-β-acyl acetals serve as activated donors in group transfer reactions involved in plant natural product biosynthesis and hormone metabolism. However, the acyltransferases that mediate transacylation from 1-
O
-β-acyl acetals have not been identified. We report the identification of a cDNA encoding a 1-
O
-β-acylglucose-dependent acyltransferase functioning in glucose polyester biosynthesis by
Lycopersicon pennellii
. The acyltransferase cDNA encodes a serine carboxypeptidase-like protein, with a conserved Ser-His-Asp catalytic triad. Expression of the acyltransferase cDNA in
Saccharomyces cerevisiae
conferred the ability to disproportionate 1-
O
-β-acylglucose to diacylglucose. The disproportionation reaction is regiospecific, catalyzing the conversion of two equivalents of 1-
O
-β-acylglucose to 1,2-di-
O
-acylglucose and glucose. Diisopropyl fluorophosphate, a transition-state analog inhibitor of serine carboxypeptidases, inhibited acyltransferase activity and covalently labeled the purified acyltransferase, suggesting the involvement of an active serine in the mechanism of the transacylation. The acyltransferase exhibits no carboxypeptidase activity; conversely, the serine carboxypeptidases we have tested show no ability to transacylate using 1-
O
-acyl-β-glucoses. This acyltransferase may represent one member of a broader class of enzymes recruited from proteases that have adapted a common catalytic mechanism of catabolism and modified it to accommodate a wide range of group transfer reactions used in biosynthetic reactions of secondary metabolism. The abundance of serine carboxypeptidase-like proteins in plants suggests that this motif has been used widely for metabolic functions.
Transient expression of the bHLH factor neurogenin-2 marks a subpopulation of neural crest cells biased for a sensory but not a neuronal fate
Authors: Zirlinger Mariela , Lo Liching , McMahon Jill , McMahon Andrew P. , Anderson David J.
Published: July 2002

Lineage-tracing experiments have indicated that some premigratory neural crest cells (NCCs) are pleuripotent, generating sensory and sympathetic neurons and their associated glia. Using an inducible Cre recombinase-based fate mapping system, we have permanently marked a subpopulation of NCCs that expresses
Ngn2
, a bHLH transcription factor required for sensory neurogenesis, and compared its fate to the bulk NCC population marked by expression of
Wnt1
.
Ngn2
+
progenitors were four times more likely than
Wnt1
+
NCCs to contribute to sensory rather than sympathetic ganglia. Within dorsal root ganglia, however, both
Ngn2
- and
Wnt1-
expressing cells were equally likely to generate neurons or glia. These data suggest that
Ngn2
marks an NCC subpopulation with a predictable fate bias, early in migration. Taken together with previous work, these data suggest that NCCs become restricted to sensory or autonomic sublineages before becoming committed to neuronal or glial fates.
The tRNA function of SsrA contributes to controlling repression of bacteriophage Mu prophage
Authors: Ranquet Caroline , Geiselmann Johannes , Toussaint Ariane
Published: July 2002

The small regulatory RNA SsrA has both tRNA and mRNA activities. It charges alanine and interacts with stalled ribosomes, allowing for translation to resume on the SsrA mRNA moiety. Hence, unfinished peptides carry a short amino acid tag, which serves as a signal for degradation by energy-dependent proteases. In SsrA-defective
Escherichia
coli
strains, thermoinducible mutants of the transposable bacteriophage Mu (Mu
cts
) are no longer induced at high temperature. Here we show that truncated forms of the key regulator of Mu lysogeny, the repressor Repc, accumulate in the absence of SsrA. These forms resemble C-terminally truncated dominant Mu repressor mutants previously isolated from Mu
cts
, which are no longer thermoinducible and bind operator DNA with a high affinity even at high temperature. Using various
ssrA
alleles, we demonstrate the importance of SsrA charging on the ribosome for controlling Mu prophage repression. Our results thus substantiate the previous observation that trans-translation is not the only function of the SsrA. The alternative function of SsrA appears to influence the stability of Mu lysogens by controlling the translation of the C-terminal domain of the repressor protein, which modulates the affinity of the protein for DNA and its susceptibility to proteolytic degradation.
Institutional radicalization, the state, and the development process in Africa
Authors: Mabogunje Akin L.
Published: July 2002

The concept of “institutional radicalization” or “the
changing of institutions from their roots” is put forward as
critical for understanding the manner in which civil societies in most
African countries have been grappling with the challenges of
development. Given the well embedded precapitalist social formation in
most African countries, various institutions have been radicalized
around such critical identities as age, gender, land, occupation,
credit, community, and rituals of solidarity to promote development.
However, until the state intervenes to legitimize these initiatives of
civil societies, their full impact on the development process is bound
to be moot. The enormous tasks entailed in such legitimization through
identifying, registering, realigning, providing incentives, directing,
and monitoring these radicalized institutions remain critical and
imperative for African countries if they are to accelerate and deepen
the extent of their integration into the global free market economy and
democratic governance.
Mathematical frontiers in optical solitons
Authors: Bronski Jared C. , Segev Mordechai , Weinstein Michael I.
Published: July 2002

Solitons are localized concentrations of field energy, resulting from a balance of dispersive and nonlinear effects. They are ubiquitous in the natural sciences. In recent years optical solitons have arisen in new and exciting contexts that differ in many ways from the original context of coherent propagation in a uniform medium. We review recent developments in incoherent spatial solitons and in gap solitons in periodic structures.
Expression of transduced genes in mice generated by infecting blastocysts with avian leukosis virus-based retroviral vectors.
Authors: Federspiel M J , Swing D A , Eagleson B , Reid S W , Hughes S H
Published: July 2002

Transgenic mouse lines have been developed that express the tv-a receptor under the control of the chicken beta-actin promoter. These mice express the tv-a receptor in most or all tissues and in the early embryo. An avian leukosis virus (ALV)-based retroviral vector system was used for the efficient delivery of genes into preimplantation mouse embryos from these transgenic lines. Experimental animals could be generated quickly and easily by infecting susceptible blastocysts with ALV-based retroviral vectors. Expression of the delivered genes was controlled by either the constitutive viral promoter contained in the long terminal repeat or an internal nonviral tissue-specific promoter. Mating the infected founder chimeric animals produced animals that carry the ALV provirus as a transgene. A subset of the integrated proviruses expressed the chloramphenicol acetyltransferase reporter gene from either the promoter in the long terminal repeat or an internal promoter, which we believe indicates that many of the sites that are accessible to viral DNA insertion in preimplantation embryos are incompatible with expression in older animals. This approach should prove useful for studies on murine cell lineage and development, providing models for studying oncogenesis, and testing gene therapy strategies.
Subcellular imaging of intramitochondrial Ca2+ with recombinant targeted aequorin: significance for the regulation of pyruvate dehydrogenase activity.
Authors: Rutter G A , Burnett P , Rizzuto R , Brini M , Murgia M , Pozzan T , Tavaré J M , Denton R M
Published: July 2002

Specific targeting of the recombinant, Ca2+ -sensitive photoprotein, aequorin to intracellular organelles has provided new insights into the mechanisms of intracellular Ca2+ homeostasis. When applied to small mammalian cells, a major limitation of this technique has been the need to average the signal over a large number of cells. This prevents the identification of inter- or intracellular heterogeneities. Here we describe the imaging in single mammalian cells (CHO.T) of [Ca2+] with recombinant chimeric aequorin targeted to mitochondria. This was achieved by optimizing expression of the protein through intranuclear injection of cDNA and through the use of a charge-coupled device camera fitted with a dual microchannel plate intensifier. This approach allows accurate quantitation of the kinetics and extent of the large changes in mitochondrial matrix [Ca2+] ([Ca2+](m)) that follow receptor stimulation and reveal different behaviors of mitochondrial populations within individual cells. The technique is compared with measurements of [Ca2+](m) using the fluorescent indicator, rhod2. Comparison of [Ca2+](m) with the activity of the Ca2+ -sensitive matrix enzyme, pyruvate dehydrogenase (PDH), reveals that this enzyme is a target of the matrix [Ca2+] changes. Peak [Ca2+](m) values following receptor stimulation are in excess of those necessary for full activation of PDH in situ, but may be necessary for the activation of other mitochondrial dehydrogenases. Finally, the data suggest that the complex regulation of PDH activity by a phosphorylation-dephosphorylation cycle may provide a means by which changes in the frequency of cytosolic (and hence mitochondrial) [Ca2+] oscillations can be decoded by mitochondria.
Natural abundance solid-state carbon NMR studies of photosynthetic reaction centers with photoinduced polarization.
Authors: Zysmilich M G , McDermott A
Published: July 2002

Solid-state NMR spectra of natural abundance 13C in reaction centers from photosynthetic bacteria Rhodobacter sphaeroides R-26 was measured. When the quinone acceptors were removed and continuous visible illumination of the sample was provided, exceptionally strong nuclear spin polarization was observed in NMR lines with chemical shifts resembling those of the aromatic carbons in bacteriochlorophyll and bacteriopheophytin. The observation of spin polarized 15N nuclei in bacteriochlorophyll and bacteriopheophytin was previously demonstrated with nonspecifically 15N-labeled reaction centers. Both the carbon and the nitrogen NMR studies indicate that the polarization is developed on species that carry unpaired electrons in the early electron transfer steps, including the bacteriochlorophyll dimer donor P860 and probably the bacteriopheophytin acceptor. I. Both enhanced-absorptive and emissive polarization were seen in the carbon spectrum; most lines were absorptive but the methine carbons of the porphyrin ring (alpha, beta, gamma, ) exhibited emissive polarization. The change in the sign of the hyperfine coupling at these sites indicates the existence of nodes in the spin density distribution on the tetrapyrrole cofactors flanking each methine carbon bridge.
Linker histones affect patterns of digestion of supercoiled plasmids by single-strand-specific nucleases.
Authors: Ivanchenko M , Zlatanova J , Varga-Weisz P , Hassan A , van Holde K
Published: July 2002

The effect of histone H1 binding on the cleavage of superhelical plasmids by single-strand-specific nucleases was investigated. Mapping of P1 cleavage sites in pBR322, achieved by EcoRI digestion after the original P1 attack, showed an intriguing phenomenon: preexisting susceptible sites became "protected," whereas some new sites appeared at high levels of H1. Similar results were obtained with another single-strand-specific nuclease, S1. Disappearance of cutting at preexisting sites and appearance of new sites was also observed in a derivative plasmid that contains a 36-bp stretch of alternating d(AT) sequence that is known to adopt an altered P1-sensitive conformation. On the other hand, H1 titration of a dimerized version of the d(AT)18-containing plasmid led to protection of all preexisting sites except the d(AT)18 inserts, which were still cut even at high H1 levels; in this plasmid no new sites appeared. The protection of preexisting sites is best explained by long-range effects of histone H1 binding on the superhelical torsion of the plasmid. The appearance of new sites, on the other hand, probably also involves a local effect of stabilization of specific sequences in Pl-sensitive conformation, due to direct H1 binding to such sequences. That such binding involves linker histone N- and/or C-terminal tails is indicated by the fact that titration with the globular domain of H5, while causing disappearance of preexisting sites, does not lead to the appearance of any new sites.
Enhanced phospholipase C-gamma1 activity produced by association of independently expressed X and Y domain polypeptides.
Authors: Horstman D A , DeStefano K , Carpenter G
Published: July 2002

The X and Y domains of phospholipase C (PLC)-gamma1, which are conserved in all mammalian phosphoinositide-specific PLC isoforms and are proposed to interact to form the catalytic site, have been expressed as individual hexahistidine-tagged fusion proteins in the baculovirus system. Following coinfection of insect cells with recombinant viruses, association of X and Y polypeptides was demonstrated in coprecipitation assays. When enzyme activity was examined, neither domain possessed catalytic activity when expressed alone; however, coexpression of the X and Y polypeptides produced a functional enzyme. This reconstituted phospholipase activity remained completely dependent on the presence of free Ca2+. The specific activity of the X:Y complex was significantly greater (20- to 100-fold) than that of holoPLC-gamma1 and was only moderately influenced by varying the concentration of substrate. The enzyme activities of holoPLC-gamma1 and the X:Y complex exhibited distinct pH optima. For holoPLC-gamma1 maximal activity was detected at pH 5.0, while activity of the X:Y complex was maximal at pH 7.2.
Induction of mucosal immune responses against a heterologous antigen fused to filamentous hemagglutinin after intranasal immunization with recombinant Bordetella pertussis.
Authors: Renauld-Mongénie G , Mielcarek N , Cornette J , Schacht A M , Capron A , Riveau G , Locht C
Published: July 2002

Live vaccine vectors are usually very effective and generally elicit immune responses of higher magnitude and longer duration than nonliving vectors. Consequently, much attention has been turned to the engineering of oral pathogens for the delivery of foreign antigens to the gut-associated lymphoid tissues. However, no bacterial vector has yet been designed to specifically take advantage of the nasal route of mucosal vaccination. Herein we describe a genetic system for the expression of heterologous antigens fused to the filamentous hemagglutinin (FHA) in Bordetella pertussis. The Schistosoma mansoni glutathione S-transferase (Sm28GST) fused to FHA was detected at the cell surface and in the culture supernatants of recombinant B. pertussis. The mouse colonization capacity and autoagglutination of the recombinant microorganism were indistinguishable from those of the wild-type strain. In addition, and in contrast to the wild-type strain, a single intranasal administration of the recombinant strain induced both IgA and IgG antibodies against Sm28GST and against FHA in the bronchoalveolar lavage fluids. No anti-Sm28GST antibodies were detected in the serum, strongly suggesting that the observed immune response was of mucosal origin. This demonstrates, to our knowledge, for the first time that recombinant respiratory pathogens can induce mucosal immune responses against heterologous antigens, and this may constitute a first step toward the development of combined live vaccines administrable via the respiratory route.
Mammalian cells resistant to tumor suppressor genes.
Authors: Pietenpol J A , Lengauer C , Jordan J , Kinzler K W , Vogelstein B
Published: July 2002

Expression of p53 causes growth arrest or apoptosis in many normal and neoplastic cell types, but the relationship between these two effects has remained obscure. To begin to dissect the underlying mechanisms at a genetic level, we have generated mutant cells resistant to the action of wild-type p53. Rat embryo fibroblasts transformed with ras and a temperature-sensitive p53 (tsp53(135val)) gene were chemically mutagenized and selected for growth at a temperature at which p53 adopts a wild-type conformation (31.5 degrees C). Clones that grew exponentially at 31.5 degrees C were selected. Cell fusion experiments demonstrated that the mutations conferring resistance to p53-mediated growth arrest were dominant. The mutagenized clones were resistant not only to p53-mediated growth arrest, but also to the apoptosis induced by E1A in conjunction with p53, and partially resistant to the retinoblastoma tumor suppressor, pRB. The results suggest that a single downstream pathway can control the induction of growth arrest and apoptosis, and that both p53 and RB function through this pathway.
Distortion in the spacer region of Pm during activation of middle transcription of phage Mu.
Authors: Artsimovitch I , Kahmeyer-Gabbe M , Howe M M
Published: July 2002

Transcription from the middle promoter, Pm, of phage Mu is initiated by Escherichia coli RNA polymerase holoenzyme (E sigma 70; RNAP) and the phage-encoded activator, Mor. Point mutations in the spacer region between the -10 hexamer and the Mor binding site result in changes of promoter activity in vivo. These mutations are located at the junction between a rigid T-tract and adjacent, potentially deformable G + C-rich DNA segment, suggesting that deformation of the spacer region may play a role in the transcriptional activation of Pm. This prediction was tested by using dimethyl sulfate and potassium permanganate footprinting analyses. Helical distortion involving strand separation was detected at positions -32 to -34, close to the predicted interface between Mor and RNAP. Promoter mutants in which this distortion was not detected exhibited a lack of melting in the -12 to -1 region and reduced promoter activity in vivo. We propose that complexes containing the distortion represent stressed intermediates rather than stable open complexes and thus can be envisaged as a transition state in the kinetic pathway of Pm activation in which stored torsional energy could be used to facilitate melting around the transcription start point.
Human immunodeficiency virus type 1 and 2 Tat proteins specifically interact with RNA polymerase II.
Authors: Mavankal G , Ignatius Ou S H , Oliver H , Sigman D , Gaynor R B
Published: July 2002

The Tat-responsive region (TAR) element is a critical RNA regulatory element in the human immunodeficiency virus (HIV) long terminal repeat, which is required for activation of gene expression by the transactivator protein Tat. Recently, we demonstrated by gel-retardation analysis that RNA polymerase II binds to TAR RNA and that Tat prevents this binding even when Tat does not bind to TAR RNA. These results suggested that direct interactions between Tat and RNA polymerase II may prevent RNA polymerase II pausing and lead to Tat-mediated increases in transcriptional elongation. To test this possibility, we performed protein interaction studies with RNA polymerase II and both the HIV-1 and the closely related HIV-2 Tat protein. These studies indicated that both the HIV-1 and HIV-2 Tat proteins could specifically interact with RNA polymerase II. Mutagenesis of both HIV-1 and HIV-2 Tat demonstrated that the basic domains of both the HIV-1 and HIV-2 Tat proteins were required for this interaction. Furthermore, "far Western" analysis suggested that the largest subunit of RNA polymerase II was the site for interaction with Tat. The interactions between Tat and RNA polymerase II were of similar magnitude to those detected between RNA polymerase II and the cellular transcription factor RAP30, which stably associates with RNA polymerase II during transcriptional elongation. These studies are consistent with the model that RNA polymerase II is a cellular target for Tat resulting in Tat-mediated increases in transcriptional elongation from the HIV long terminal repeat.
Expression of the C terminus of the amyloid precursor protein alters growth factor responsiveness in stably transfected PC12 cells.
Authors: Sandhu F A , Kim Y , Lapan K A , Salim M , Aliuddin V , Zain S B
Published: July 2002

The amyloid precursor protein (APP) is a molecule centrally involved in Alzheimer disease pathology, but whose normal function is still poorly understood. To investigate the consequences of increased intracellular production of various regions of APP on cellular physiology, we stably transfected PC12 cells with the C-terminal 100 amino acids of the human APP. In eight transfected clones that express the APP(C100) protein, exposure to nerve growth factor (NGF) did not promote differentiation. Transfectants continued to divide and failed to elaborate extensive neurites, whereas control PC12 cells, mock-transfected PC12 cells, and a nonexpressing transfected cell line did develop neurites and stopped dividing after NGF stimulation. Unlike NGF treatment, treatment with basic fibroblast growth factor profoundly accelerated neurite outgrowth in transfected cells. Also, a dramatic increase in a tyrosine phosphatase activity was noted. Expression and accumulation of APP C100 protein in PC12 cells results in an abnormal response to growth factor stimulation.
Bombesin antagonist prevents CO2 laser-induced promotion of oral cancer.
Authors: Kozacko M F , Mang T S , Schally A V , Priore R L , Liebow C
Published: July 2002

We previously reported that CO2 laser incisions in carcinogen-initiated fields promoted cancer development and caused release of growth factors. Here we examined the quantitative and additive properties of this tumor-promoting event and examined whether this promotion could be nullified by treatment with a bombesin antagonist, which down-regulates epidermal growth factor receptors. The model used for cancer promotion was the hamster buccal cheek pouch that had been treated with a carcinogen (9,10-dimethyl-1,2-benzanthracene) for 6 weeks, producing premalignant lesions. These lesions would evolve into a cancer eventually without further treatment. Promotion was measured both by increased fluorescence in response to systemically administered Photofrin, measured noninvasively using an in vivo fluorescence photometer, and by the timing of appearance of clinical tumors. Laser incisions (0-3) were made into the hamster cheek 1 week apart, or three incisions were done 1 day apart. Another group of animals received bombesin antagonist RC-3095 for 4 weeks during the time incisions were made, again measuring promotion. Laser incisions 1 week apart produced additive promotion, whereas three incisions 1 day apart were not statistically different from the group receiving only one incision. RC-3095 treatment completely eliminated the promoting effects of incision and totally stopped promotion for the 4-week period of treatment. After discontinuing treatment with RC-3095, lesion progression resumed at the untreated control rate. This work confirms that the promoting event of a laser incision follows a comparable time course to release of growth factors after such an incision and that it can be eliminated by treatment with bombesin antagonists.
A selective phenomenology of the seismicity of Southern California.
Authors: Knopoff L
Published: July 2002

Predictions of earthquakes that are based on observations of precursory seismicity cannot depend on the average properties of the seismicity, such as the Gutenberg-Richter (G-R) distribution. Instead it must depend on the fluctuations in seismicity. We summarize the observational data of the fluctuations of seismicity in space, in time, and in a coupled space-time regime over the past 60 yr in Southern California, to provide a basis for determining whether these fluctuations are correlated with the times and locations of future strong earthquakes in an appropriate time- and space-scale. The simple extrapolation of the G-R distribution must lead to an overestimate of the risk due to large earthquakes. There may be two classes of earthquakes: the small earthquakes that satisfy the G-R law and the larger and large ones. Most observations of fluctuations of seismicity are of the rate of occurrence of smaller earthquakes. Large earthquakes are observed to be preceded by significant quiescence on the faults on which they occur and by an intensification of activity at distance. It is likely that the fluctuations are due to the nature of fractures on individual faults of the network of faults. There are significant inhomogeneities on these faults, which we assume will have an important influence on the nature of self-organization of seismicity. The principal source of the inhomogeneity on the large scale is the influence of geometry--i.e., of the nonplanarity of faults and the system of faults.
Acridones: a chemically new group of protonophores.
Authors: Horváth G , Droppa M , Fodorpataki L , Istokóvics A , Garab G , Oettmeier W
Published: July 2002

Although the interaction of proton-conducting ionophores (protonophores) with photosynthetic electron transport has been extensively studied during the past decade, the mode of action of protonophores remained uncertain. For a better understanding of the molecular mechanism of the action of protonophores, the introduction of chemically new types of molecules will be required. In this work, we demonstrate that acridones (9-azaanthracene-10-ones) completely fulfill this requirement. At low concentrations of acridones, the thermoluminescence bands at +20 degrees C and +10 degrees C were strongly inhibited, while normal electron transport activity was retained. This indicates that the concentrations of S2 and S3 states involved in the generation of these bands are reduced. At higher concentrations, an increased activity of electron transport was observed, which is attributed to the typical uncoupler effect of protonophores. Indeed, acridones accelerate the decay of the electrochromic absorbance change at 515 nm and also inhibit the generation of the transmembrane proton gradient, measured as an absorbance transient of neutral red. Variable fluorescence induction was quenched even at low concentrations of acridones but was restored by either a long-term illumination or high light intensity. Acridones, similarly to other protonophores, promoted the autooxidation of the high-potential form of cytochrome b559 and partially converted it to lower potential forms. These results suggest that acridones, acting as typical protonophores, uncouple electron transport, accelerate the deactivation of the S2 and S3 states on the donor side, and facilitate the oxidation of cytochrome b559 on the acceptor side of photosystem II.
Is there a Population II analogy to the F star lithium dip?
Authors: Dearborn D. S. P. , Schramm David N.
Published: July 2002

Observers have found a small number of lithium-depleted halo stars in the temperature range of the Spite plateau. The current status of the mass-loss hypothesis for producing the observed lithium dip in Population (Pop) I stars is briefly discussed and extended to Pop II stars as a possible explanation for these halo objects. Based on detections of F-type main-sequence variables, mass loss is assumed to occur in a narrow temperature region corresponding to this “instability strip.” As Pop II main-sequence stars evolve to the blue, they enter this narrow temperature region, then move back through the lower temperature area of the Spite plateau. If 0.05
M
⊙
(solar mass) or more have been lost, they will show lithium depletion. This hypothesis affects the lithium-to- beryllium abundance, the ratio of high- to low-lithium stars, and the luminosity function. Constraints on the mass-loss hypothesis due to these effects are discussed. Finally, mass loss in this temperature range would operate in stars near the turnoff of metal-poor globular clusters, resulting in apparent ages 2 to 3 Gyr (gigayears) older than they actually are.
Amphotropic murine leukemia viruses induce spongiform encephalomyelopathy
Authors: Münk Carsten , Löhler Jürgen , Prassolov Vladimir , Just Ursula , Stockschläder Marcus , Stocking Carol
Published: July 2002

Recombinants of amphotropic murine leukemia virus (A-MuLV) have found widespread use in retroviral vector systems due to their ability to efficiently and stably infect cells of several different species, including human. Previous work has shown that replication-competent recombinants containing the amphotropic
env
gene, encoding the major SU envelope glycoprotein that determines host tropism, induce lymphomas
in vivo
. We show here that these viruses also induce a spongiform encephalomyelopathy in mice inoculated perinatally. This fatal central nervous system disease is characterized by noninflammatory spongiform lesions of nerve and glial cells and their processes, and is associated with moderate astro- and microgliosis. The first clinical symptoms are ataxia, tremor, and spasticity, progressing to complete tetraparesis and incontinence, and finally death of the animal. Sequences within the amphotropic
env
gene are necessary for disease induction. Coinfection of A-MuLV recombinants with nonneuropathogenic ecotropic or polytropic MuLV drastically increases the incidence, degree, and distribution of the neurodegenerative disorder. The consequence of these results in view of the use of A-MuLV recombinants in the clinic is discussed.
Analysis of linear trade models and relation to scale economies
Authors: Gomory Ralph E. , Baumol William J.
Published: July 2002

We discuss linear Ricardo models with a range of parameters. We show that the exact boundary of the region of equilibria of these models is obtained by solving a simple integer programming problem. We show that there is also an exact correspondence between many of the equilibria resulting from families of linear models and the multiple equilibria of economies of scale models.
The α chain of laminin-1 is independently secreted and drives secretion of its β- and γ-chain partners
Authors: Yurchenco Peter D. , Quan Yong , Colognato Holly , Mathus Todd , Harrison David , Yamada Yoshihiko , O’Rear Julian J.
Published: July 2002

A mammalian recombinant strategy was established to dissect rules of basement membrane laminin assembly and secretion. The α-, β-, and γ-chain subunits of laminin-1 were expressed in all combinations, transiently and/or stably, in a near-null background. In the absence of its normal partners, the α chain was secreted as intact protein and protein that had been cleaved in the coiled-coil domain. In contrast, the β and γ chains, expressed separately or together, remained intracellular with formation of ββ or βγ, but not γγ, disulfide-linked dimers. Secretion of the β and γ chains required simultaneous expression of all three chains and their assembly into αβγ heterotrimers. Epitope-tagged recombinant α subunit and recombinant laminin were affinity-purified from the conditioned medium of αγ and αβγ clones. Rotary-shadow electron microscopy revealed that the free α subunit is a linear structure containing N-terminal and included globules with a foreshortened long arm, while the trimeric species has the typical four-arm morphology of native laminin. We conclude that the α chain can be delivered to the extracellular environment as a single subunit, whereas the β and γ chains cannot, and that the α chain drives the secretion of the trimeric molecule. Such an α-chain-dependent mechanism could allow for the regulation of laminin export into a nascent basement membrane, and might serve an important role in controlling basement membrane formation.
In vitro
selection of self-cleaving RNAs with a low pH optimum
Authors: Jayasena Vineetha K. , Gold Larry
Published: July 2002

RNAs that undergo a rapid site-specific cleavage at low pH have been selected by
in vitro
selection (the SELEX process). The cleavage does not require the addition of any divalent metal ions, and is in fact inhibited by divalent metal ions, spermine, or high concentrations of monovalent metal ions. This low pH catalyzed cleavage results in a 2′,3′-cyclic phosphate at the 3′ end and a free hydroxyl at the 5′ end. The reaction proceeds with a calculated rate of 1.1 min
−1
at room temperature in cacodylate buffer at pH 5.0. The rate of cleavage is dependent on the pH and shows an optimum around pH 4.0. The rate constant is independent of RNA concentration, indicating to an intramolecular reaction. Autocatalytic cleavage at low pH, in the absence of a metal ion requirement, adds to the reaction possibilities that may have existed on the prebiotic earth.
Binding of RNA template to a complex of HIV-1 reverse transcriptase/primer/template
Authors: Canard Bruno , Sarfati Robert , Richardson Charles C.
Published: July 2002

HIV-1 reverse transcriptase (RT) catalyzes the synthesis of DNA from DNA or RNA templates. During this process, it must transfer its primer from one template to another RNA or DNA template. Binary complexes made of RT and a primer/template bind an additional single-stranded RNA molecule of the same nucleotide sequence as that of the DNA or RNA template. The additional RNA strand leads to a 10-fold decrease of the off-rate constant,
k
off
, of RT from a primer/DNA template. In a binary complex of RT and a primer/template, the primer can be cross-linked to both the p66 and p51 subunits. Depending on the location of the photoreactive group in the primer, the distribution of the cross-linked primers between subunits is dependent on the nature of the template and of the additional single-stranded molecule. Greater cross-linking of the primer to p51 occurs with DNA templates, whereas cross-linking to p66 predominates with RNA templates. Excess single-stranded DNA shifts the distribution of cross-linking from p66 to p51 with RNA templates, and excess single-stranded RNA shifts the cross-linking from p51 to p66 with DNA templates. RT thus uses two primer/template binding modes depending on the nature of the template.
Hormone-regulatable neoplastic transformation induced by a Jun-estrogen receptor chimera
Authors: Kruse Ulrich , Iacovoni Jason S. , Goller Martin E. , Vogt Peter K.
Published: July 2002

The v-
jun
oncogene encodes a nuclear DNA binding protein that functions as a transcription factor and is part of the activator protein 1 complex. Oncogenic transformation by v-
jun
is thought to be mediated by the aberrant expression of specific target genes. To identify such Jun-regulated genes and to explore the mechanisms by which Jun affects their expression, we have fused the full-length v-Jun and an amino-terminally truncated form of v-Jun to the hormone-binding domain of the human estrogen receptor. The two chimeric proteins function as ligand-inducible transactivators. Expression of the fusion proteins in chicken embryo fibroblasts causes estrogen-dependent transformation.