bluuebunny/crossref_metadata_2025 huggingface dataset:
Deep states and misfit dislocations in indium-doped GaAs layers grown by molecular beam epitaxy
Authors: Ioannou D. E. , Huang Y. J. , Iliadis A. A.
Published: July 2002

We have studied the effects of adding small amounts of indium (0.6 at. %) in GaAs layers grown by molecular beam epitaxy on GaAs substrates. Photoluminescence spectra showed that epilayers of high crystal quality were obtained, and electron beam induced current microscopy revealed a number of misfit dislocations near the interface and a dislocation-free zone near the top surface. We observed several well known deep traps in the dislocation-free zone, albeit at much lower concentrations in comparison to indium-free, otherwise identical epilayers. We failed to observe any traps in the misfit dislocation zone. If, as is widely held, we assume that these traps are due to point defects and their complexes, a possible explanation of the absence of traps in the dislocation zone is that the traps provide the point defects necessary for the creation and climb of the misfit dislocations.
Extreme nonequilibrium electron transport in heterojunction bipolar transistors
Authors: Berthold K. , Levi A. F. J. , Walker J. , Malik R. J.
Published: July 2002

We use hot-electron spectroscopy to demonstrate the existence of extreme nonequilibrium electron transport in the base of n-p-n heterojunction bipolar transistors. In the device, electrons are tunnel injected into a thin (∼300 Å wide), degenerately doped, p-type GaAs base.
Three-dimensional integration of resonant tunneling structures for signal processing and three-state logic
Authors: Potter Robert C. , Lakhani Amir A. , Beyea Dana , Hier Harry , Hempfling Erica , Fathimulla Ayub
Published: July 2002

We have developed structures with two well-defined negative differential resistance (NDR) regions by sequentially growing two resonant tunneling devices separated by an n+ connecting layer. Devices fabricated from these structures exhibited three stable operating points for multilevel logic circuits and were used in circuits which multiplied the input signal frequency by 3 or 5. This approach can be extended to obtain more than two NDR regions by vertical integration of additional resonant tunneling structures.
Line shape of the optical dielectric function
Authors: Garland J. W. , Abad H. , Viccaro M. , Raccah P. M.
Published: July 2002

A systematic study of the optical dielectric function by spectroscopic ellipsometry and electroreflectance has shown that the proper functional form for the Green’s function for an electron-hole pair in GaAs or CdTe is primarily Gaussian, not Lorentzian as is commonly assumed, although it is primarily Lorentzian for Hg1−xCdxTe. The Lorentzian part of the broadening is shown to measure the alloy, impurity, and defect scattering.
Temperature dependence of threshold current in GaAs/AlGaAs quantum well lasers
Authors: Blood P. , Colak S. , Kucharska A. I.
Published: July 2002

We have calculated the threshold current and its temperature (T) dependence in the range 200–400 K for AlGaAs quantum well lasers with 25-Å-wide GaAs wells using a model which includes lifetime broadening of the transitions and broadening of the density of states function by fluctuations in the well width. The threshold current varies approximately linearly with T and the principal effect of broadening is to increase the threshold current causing a reduction in the fractional change of current with temperature. The apparent value of the parameter T0 is increased to ≊400 K, compared with ≊320 K without broadening. The calculations are compared with experimental data.
Metal-oxide-semiconductor characteristics of rapid thermal nitrided thin oxides
Authors: Shih D. K. , Chang W. T. , Lee S. K. , Ku Y. H. , Kwong D. L. , Lee S.
Published: July 2002

The electrical properties of thin nitrided oxide (∼100 Å) formed by rapid thermal nitridation (RTN) in pure NH3 have been studied. It is found that the current-voltage characteristic of RTN oxides follows a Fowler–Nordheim tunneling behavior with modifications caused by electron trapping processes at the oxide surface and interface. The trapping density is dependent on the RTN conditions. At the interface, both fixed charge (Nf) and interface state (Dit) densities exhibit turnaround phenomena when the RTN process proceeds. The maximum values of Nf and Dit at the turnaround points are lower for the higher temperature RTN, suggesting a viscous flow related strain relieving mechanism associated with RTN of thin oxides. Films with superior endurance behavior (QBD=20.4 C/cm2 compared with QBD=5.1 C/cm2 of thermal oxide under 10 mA/cm2 constant current stress) have been obtained by RTN at 1000 °C, 10 s.
Radiation effects in low-pressure reoxidized nitrided oxide gate dielectrics
Authors: Dunn G. J. , Jayaraman R. , Yang W. , Sodini C. G.
Published: July 2002

Nitridation of the gate oxide is known to improve the radiation resistance of metal-oxide-semiconductor devices on silicon. However, it has generally been found that high-temperature, long-time anneals are required to produce the hardest dielectrics. We report results of 1–5 Mrad (Si) total dose radiation testing of thin gate oxides which have received relatively light nitridations, followed by a reoxidation step. These devices exhibit substantially reduced fixed charge and interface state buildup compared with oxide, and appear to be harder than the more heavily nitrided oxides previously reported.
Selective amplification of self-resistively heated laser-direct-written tungsten lines
Authors: Gottsleben O. , Stuke M.
Published: July 2002

Laser-direct-write pyrolytic deposition of tungsten at 514.5 nm, out of the WF6 /H2 reduction system, was carried out on Al2O3 substrates between sputtered-on gold contact pads. Heat, generated by pulsed current sent through the laser-induced microdeposit, is used to amplify the microstructure by a conventional, however well localized, chemical vapor deposition process. In addition to tungsten, other more volatile materials can be deposited for amplification.
Chemically controlled deep level formation and band bending at metal-CdTe interfaces
Authors: Shaw J. L. , Viturro R. E. , Brillson L. J. , LaGraffe D.
Published: July 2002

We have used reactive metal interlayers to suppress anion outdiffusion at Au-CdTe junctions and thereby to alter the formation of deep interfacial states. Using soft x-ray photoemission and luminescence spectroscopies, we report a dramatically reduced p-type band bending and demonstrate that deep levels observed directly at the interface are responsible for the chemically induced electrical behavior.
New technique and analysis of accelerated electromigration life testing in multilevel metallizations
Authors: Muray L. P. , Rathbun L. C. , Wolf E. D.
Published: July 2002

Electromigration failure of a series-parallel configuration of aluminum interconnects overlayed on tungsten contacts was measured using a novel multiple lognormal analysis. The analysis examined early failure mechanisms and allowed rapid determination of electromigration parameters on a statistically large number of junctions. The primary failure mode of these stuctures was complete migration of Al off of the W pads. This work suggested that Al/W metallizations, with a large number of series contacts, are prone to short mean time to failures.
Low threshold buried-heterostructure quantum well lasers by excimer laser assisted disordering
Authors: Epler J. E. , Thornton R. L. , Mosby W. J. , Paoli T. L.
Published: July 2002

Laser assisted disordering based upon a direct-write Ar+ laser beam has been established as a fabrication technique for high quality optoelectronic devices. In this letter, we report a new form of laser assisted disordering in which an excimer laser beam, photolithographically patterned, is used to define the incorporation of Si impurity into GaAs-AlGaAs heterostructure crystals. During a subsequent thermal anneal the diffusing Si induces layer disordering to a depth of ∼1 μm. The excimer laser assisted disordering process is characterized as a function of the energy density of the laser beam. Also, this technique is used to fabricate high quality buried-heterostructure lasers. With a reflective rear facet, the typical cw threshold current is 4 mA and the maximum power output is 27 mW. The devices exhibit single fundamental mode operation with subsidiary longitudinal side modes suppressed by 34 dB.
A
Bacillus subtilis
operon containing genes of unknown function senses tRNA
Trp
charging and regulates expression of the genes of tryptophan biosynthesis
Authors: Sarsero Joseph P. , Merino Enrique , Yanofsky Charles
Published: July 2002

Strains of
Bacillus subtilis
containing a
temperature-sensitive tryptophanyl-tRNA synthetase produce elevated
levels of the tryptophan pathway enzymes, when grown at high
temperatures in the presence of excess tryptophan. This increase is
because of reduced availability of the tryptophan-activated
trp
RNA-binding attenuation protein (TRAP). To test the
hypothesis that this elevated
trp
gene expression was
caused by the overproduction of a transcript capable of binding and
sequestering TRAP, a computer program was designed to search the
B. subtilis
genome sequence for additional potential
TRAP binding sites. A region containing a stretch of (G/A)AG
trinucleotide repeats, characteristic of a TRAP binding site, was
identified in the
yczA-ycbK
operon. We show that
transcriptional regulation of the
yczA-ycbK
operon is
controlled by the T-box antitermination mechanism in response to the
level of uncharged tRNA
Trp
, and that the presence of a
trpS1
mutant allele increases production of the
yczA
-
ycbK
transcript. Elevated
yczA-ycbK
expression was shown to activate transcription
of the
trp
operon. Deletion of the
yczA-ycbK
operon abolishes the
trpS1
effect on
trp
gene expression. The purpose of increasing
expression of the genes of tryptophan biosynthesis in the
trpS
mutant would be to provide additional tryptophan to
overcome the charged tRNA
Trp
deficiency. Therefore, in
B. subtilis
, as in
Escherichia coli
,
transcription of the tryptophan biosynthetic genes is regulated in
response to changes in the extent of charging of tRNA
Trp
as
well as the availability of tryptophan.
Gain of imprinting at chromosome 11p15: A pathogenetic mechanism identified in human hepatocarcinomas
Authors: Schwienbacher Christine , Gramantieri Laura , Scelfo Rosaria , Veronese Angelo , Calin George A. , Bolondi Luigi , Croce Carlo M. , Barbanti-Brodano Giuseppe , Negrini Massimo
Published: July 2002

Genomic imprinting is a reversible condition that causes parental-specific silencing of maternally or paternally inherited genes. Analysis of DNA and RNA from 52 human hepatocarcinoma samples revealed abnormal imprinting of genes located at chromosome 11p15 in 51% of 37 informative samples. The most frequently detected abnormality was gain of imprinting, which led to loss of expression of genes present on the maternal chromosome. As compared with matched normal liver tissue, hepatocellular carcinomas showed extinction or significant reduction of expression of one of the alleles of the
CDKN1C
,
SLC22A1L
, and
IGF2
genes. Loss of maternal-specific methylation at the
KvDMR1
locus in hepatocarcinoma correlated with abnormal expression of
CDKN1C
and
IGF2
, suggesting a function for
KvDMR1
as a long-range imprinting center active in adult tissues. These results point to the role of epigenetic mechanisms leading to loss of expression of imprinted genes at chromosome region 11p15 in human tumors.
Host microarray analysis reveals a role for the
Salmonella
response regulator
phoP
in human macrophage cell death
Authors: Detweiler Corrella S. , Cunanan Dolores B. , Falkow Stanley
Published: July 2002

Bacterial pathogens manipulate host cells to promote pathogen
survival and dissemination. We used a 22,571 human cDNA microarray to
identify host pathways that are affected by the
Salmonella
enterica
subspecies
typhimurium phoP
gene, a
transcription factor required for virulence, by comparing the
expression profiles of human monocytic tissue culture cells infected
with either the wild-type bacteria or a
phoP∷Tn10
mutant strain. Both wild-type and
phoP∷Tn10
bacteria
induced a common set of genes, many of which are proinflammatory.
Differentially expressed genes included those that affect host cell
death, suggesting that the
phoP
regulatory system
controls bacterial genes that alter macrophage survival. Subsequent
experiments showed that the
phoP∷Tn10
mutant strain
is defective for killing both cultured and primary human macrophages
but is able to replicate intracellularly. These experiments indicate
that
phoP
plays a role in
Salmonella
-induced human macrophage cell death.
Function of the bacterial TATAAT −10 element as single-stranded DNA during RNA polymerase isomerization
Authors: Fenton Mike S. , Gralla Jay D.
Published: July 2002

The bacterial TATAAT −10 region sequence was the first promoter element to be identified, but how it functions is still not clear. Because the duplex element is melted during initiation, the effects of substitutions were studied in both single-and double-strand contexts. Band-shift results were particularly unexpected in the context of melted DNA. The effect of the lac UV5-melted −10 region on polymerase binding was found to include a large sequence nonspecific contribution. Instead the dominant role of single-stranded −10 region nucleotides was in directing the isomerization of the RNA polymerase to its heparin resistant form. This role becomes minimal when the melting is extended beyond the −10 region to encompass the transcription start site, as in the final open complex. The duplex binding results are in agreement with previous reports that showed positions −12T and −11A are of primary importance for promoter recognition. Thus the consensus −10 region sequences function in two ways, both before full promoter melting. They stabilize initial polymerase binding via duplex interactions and subsequently as single-stranded DNA they promote enzyme isomerization to the functional form.
SMAD4 mutations in colorectal cancer probably occur before chromosomal instability, but after divergence of the microsatellite instability pathway
Authors: Woodford-Richens K. L. , Rowan A. J. , Gorman P. , Halford S. , Bicknell D. C. , Wasan H. S. , Roylance R. R. , Bodmer W. F. , Tomlinson I. P. M.
Published: July 2002

Loss of chromosome 18q21 is well documented in colorectal cancer,
and it has been suggested that this loss targets the
DCC
,
DPC4
/
SMAD4
, and
SMAD2
genes. Recently, the importance of
SMAD4
, a downstream regulator in the TGF-β signaling
pathway, in colorectal cancer has been highlighted, although the
frequency of
SMAD4
mutations appears much lower than
that of 18q21 loss. We set out to investigate allele loss, mutations,
protein expression, and cytogenetics of chromosome 18 copy number in a
collection of 44 colorectal cancer cell lines of known status with
respect to microsatellite instability (MSI). Fourteen of thirty-two
MSI
−
lines showed loss of SMAD4 protein expression;
usually, one allele was lost and the other was mutated in one of a
number of ways, including deletions of various sizes, splice site
changes, and missense and nonsense point mutations (although no
frameshifts). Of the 18 MSI
−
cancers with retained SMAD4
expression, four harbored missense mutations in the 3′ part of the gene
and showed allele loss. The remaining 14 MSI
−
lines had no
detectable
SMAD4
mutation, but all showed allele loss at
SMAD4
and/or
DCC
.
SMAD4
mutations can therefore account for about 50–60% of the 18q21 allele
loss in colorectal cancer. No MSI
+
cancer showed loss of
SMAD4 protein or
SMAD4
mutation, and very few had
allelic loss at
SMAD4
or
DCC
, although
many of these MSI
+
lines did carry
TGFBIIR
changes. Although
SMAD4
mutations have been associated
with late-stage or metastatic disease, our combined molecular and
cytogenetic data best fit a model in which
SMAD4
mutations occur before colorectal cancers become aneuploid/polyploid,
but after the MSI
+
and MSI
−
pathways diverge.
Thus, MSI
+
cancers may diverge first, followed by
CIN
+
(chromosomal instability) cancers, leaving other
cancers to follow a CIN
−
MSI
−
pathway.
Origin and significance of a founding settlement in Polynesia
Authors: Burley David V. , Dickinson William R.
Published: July 2002

Selected prehistoric potsherds from the deepest cultural level of
the oldest known archaeological site in the Kingdom of Tonga, within
the Eastern Lapita province of western Polynesia, display decorative
motifs characteristic of the Western Lapita province of modern-day
Island Melanesia, to the west. Most of the stylistically anomalous
sherds contain temper sands exotic to Tonga but, in one case,
petrographically indistinguishable from temper in a Lapita sherd
recovered from the Santa Cruz Islands of Melanesia, and are inferred to
record maritime transport of Lapita ceramics into Tonga from Melanesia
far to the west. The non-Tongan sherds found on Tongatapu provide
direct physical evidence for interisland transfer of earthenware
ceramics between Western and Eastern Lapita provinces, and the Nukuleka
site, where they occur, is interpreted as one of the founding
settlements of Polynesia.
Posttranscriptional regulation of gene expression in learning by the neuronal ELAV-like mRNA-stabilizing proteins
Authors: Quattrone Alessandro , Pascale Alessia , Nogues Xavier , Zhao Weiqin , Gusev Pavel , Pacini Alessandra , Alkon Daniel L.
Published: July 2002

The view that memory is encoded by variations in the strength of
synapses implies that long-term biochemical changes take place within
subcellular microdomains of neurons. These changes are thought
ultimately to be an effect of transcriptional regulation of specific
genes. Localized changes, however, cannot be fully explained by a
purely transcriptional control of gene expression. The neuron-specific
ELAV-like HuB, HuC, and HuD RNA-binding proteins act
posttranscriptionally by binding to adenine- and uridine-rich elements
(AREs) in the 3′ untranslated region of a set of target mRNAs, and by
increasing mRNA cytoplasmic stability and/or rate of
translation. Here we show that neuronal ELAV-like genes undergo a
sustained up-regulation in hippocampal pyramidal cells only of mice and
rats that have learned a spatial discrimination paradigm. This
learning-specific increase of ELAV-like proteins was localized within
cytoplasmic compartments of the somata and proximal dendrites and was
associated with the cytoskeleton. This increase was also accompanied by
enhanced expression of the GAP-43 gene, known to be regulated mainly
posttranscriptionally and whose mRNA is demonstrated here to be an
in vivo
ELAV-like target. Antisense-mediated knockdown
of HuC impaired spatial learning performance in mice and induced a
concomitant down-regulation of GAP-43 expression. Neuronal ELAV-like
proteins could exert learning-induced posttranscriptional control of an
array of target genes uniquely suited to subserve substrates of memory
storage.
Delivery of a stringent dimerizer-regulated gene expression system in a single retroviral vector
Authors: Pollock Roy , Issner Robbyn , Zoller Karen , Natesan Sridaran , Rivera Victor M. , Clackson Tim
Published: July 2002

Small molecule-regulated transcription has broad utility and would
benefit from an easily delivered self-contained regulatory cassette
capable of robust, tightly controlled target gene expression. We
describe the delivery of a modified dimerizer-regulated gene expression
system to cells on a single retrovirus. A transcription factor cassette
responsive to the natural product dimerizer rapamycin was optimized for
retroviral delivery by fusing a highly potent chimeric activation
domain to the rapamycin-binding domain of FKBP-rapamycin-associated
protein (FRAP). This improvement led to an increase in both the potency
and maximal levels of gene expression induced by rapamycin, or
nonimmunosuppressive rapamycin analogs. The modified transcription
factor cassette was incorporated along with a target gene into a single
rapamycin-responsive retrovirus. Cell pools stably transduced with the
single virus system displayed negligible basal expression and gave
induction ratios of at least three orders of magnitude in the presence
of rapamycin or a nonimmunosuppressive rapamycin analog. Levels of
induced gene expression were comparable to those obtained with the
constitutive retroviral long terminal repeat and the single virus
system performed well in four different mammalian cell lines.
Regulation with the dimerizer-responsive retrovirus was tight enough to
allow the generation of cell lines displaying inducible expression of
the highly toxic diphtheria toxin A chain gene. The ability to deliver
the tightly inducible rapamycin system in a single retrovirus should
facilitate its use in the study of gene function in a broad range of
cell types.
Singlet exciton binding energy in poly(phenylene vinylene)
Authors: Moses D. , Wang J. , Heeger A. J. , Kirova N. , Brazovski S.
Published: July 2002

The exciton binding energy (
E

b

) and the band gap energy (
E

g

) of poly(phenylene vinylene) are determined by high-resolution measurements of the photoconductivity excitation profile as a function of light polarization, applied electric field, and temperature. At high applied electric fields, a peak in the photoconductivity is observed when the sample is pumped at a photon energy just below the onset of the band-to-band π
-
π* absorption. This peak is interpreted as resulting from field ionization of a weakly bound exciton with
E

b

≈ 60 meV. The binding energy is obtained from the energy of the exciton peak relative to the band edge and independently from analysis of the dependence of the exciton dissociation on field and temperature.
Leaf-to-shoot apex movement of symplastic tracer is restricted coincident with flowering in
Arabidopsis
Authors: Gisel Andreas , Hempel Frederick D. , Barella Sandra , Zambryski Patricia
Published: July 2002

Classical experiments in plant physiology showed that leaves are the source of signals that control the development of flowers from shoot meristems. Additional physiological and genetic experiments have indicated some of the molecules (e.g., gibberellins, cytokinins, and sucrose) that promote flowering in mustards including
Arabidopsis
. These small hydrophilic molecules are likely to move to the shoot apex symplastically via the phloem and/or via cell-to-cell movement through plasmodesmata. To analyze potential changes in the symplastic trafficking of small molecules during the induction of flowering in
Arabidopsis
, we measured changes in the flow of symplastic tracers from the leaf to the shoot apex. We previously found that the onset of flowering is coincident with an evident decrease in the leaf-to-shoot trafficking of symplastic tracer molecules; this decrease in trafficking is transitory and resumes when floral development is established. Here we provide detailed analyses of symplastic connectivity during floral induction by monitoring tracer movement under different photoperiodic induction conditions and in a number of genetic backgrounds with altered flowering times. In all cases, the correlation between flowering and the reduction of symplastic tracer movement holds true. The lack of tracer movement during the induction of flowering may represent a change in plasmodesmal selectivity at this time or that a period of reduced symplastic communication is associated with floral induction.
The transcriptional transactivator of simian foamy virus 1 binds to a DNA target element in the viral internal promoter.
Authors: Zou J X , Luciw P A
Published: July 2002

The transcriptional transactivator (Tas) of simian foamy virus type 1 strongly augments gene expression directed by both the promoter in the viral long terminal repeat and the newly discovered internal promoter located within the env gene. A region of 121 bp, located immediately 5' to the TATA box in the internal promoter, is required for transactivation by Tas. The present study aimed to identify the precise Tas-responsive target(s) in this region and to determine the role of Tas in transcriptional regulation. By analysis of both clustered-site mutations and hybrid promoters in transient expression assays in murine and simian cells, two separate sequence elements within this 121-bp region were shown to be Tas-dependent transcriptional enhancers. These targets, each < 30 bp in length and displaying no apparent sequence homology one to the other, are designated the promoter-proximal and promoter-distal elements. By means of the gel electrophoresis mobility-shift assays, using purified glutathione S-transferase-Tas fusion protein expressed in Escherichia coli, the target proximal to the TATA box exhibited strong binding to glutathione S-transferase-Tas, whereas the distal element appears not to bind. In addition, footprint analysis revealed that 26 bp in the promoter proximal element was protected by glutathione S-transferase-Tas from DNase I. We propose a model for transactivation of the simian foamy virus type 1 internal promoter in which Tas interacts directly with the proximal target element positioned immediately 5' to the TATA box. In this model, Tas attached to this element is presumed to interact with a component(s) of the cellular RNA polymerase II initiation complex and thereby enhance transcription directed by the viral internal promoter.
The kinesin walk: a dynamic model with elastically coupled heads.
Authors: Derényi I , Vicsek T
Published: July 2002

Recently individual two-headed kinesin molecules have been studied in in vitro motility assays revealing a number of their peculiar transport properties. In this paper we propose a simple and robust model for the kinesin stepping process with elastically coupled Brownian heads that show all of these properties. The analytic and numerical treatment of our model results in a very good fit to the experimental data and practically has no free parameters. Changing the values of the parameters in the restricted range allowed by the related experimental estimates has almost no effect on the shape of the curves and results mainly in a variation of the zero load velocity that can be directly fitted to the measured data. In addition, the model is consistent with the measured pathway of the kinesin ATPase.
Enzyme-mediated spatial segregation on individual polymeric support beads: application to generation and screening of encoded combinatorial libraries.
Authors: Vágner J , Barany G , Lam K S , Krchnák V , Sepetov N F , Ostrem J A , Strop P , Lebl M
Published: July 2002

Proteolysis of short N alpha-protected peptide substrates bound to polyoxyethylene-polystyrene beads releases selectively free amino sites in the enzyme-accessible "surface" area. The substantial majority of functional sites in the "interior" of the polymeric support are not reached by the enzyme and remain uncleaved (protected). Subsequent synthesis with two classes of orthogonal protecting groups-N alpha-tert-butyloxycarbonyl (Boc) and N alpha-9-fluorenylmethyloxy-carbonyl (Fmoc)-allows generation of two structures on the same bead. The surface structure is available for receptor interactions, whereas the corresponding interior structure is used for coding. Coding structures are usually readily sequenceable peptides. This "shaving" methodology was illustrated by the preparation of a peptide-encoded model peptide combinatorial library containing 1.0 x 10(5) members at approximately 6-fold degeneracy. From this single library, good ligands were selected for three different receptors: anti-beta-endorphin anti-body, streptavidin, and thrombin, and the binding structures were deduced correctly by sequencing the coding peptides present on the same beads.
p21Cip1/Waf1 disrupts the recruitment of human Fen1 by proliferating-cell nuclear antigen into the DNA replication complex.
Authors: Chen U , Chen S , Saha P , Dutta A
Published: July 2002

Fen1 or maturation factor 1 is a 5'-3' exonuclease essential for the degradation of the RNA primer-DNA junctions at the 5' ends of immature Okazaki fragments prior to their ligation into a continuous DNA strand. The gene is also necessary for repair of damaged DNA in yeast. We report that human proliferating-cell nuclear antigen (PCNA) associates with human Fen1 with a Kd of 60 nM and an apparent stoichiometry of three Fen1 molecules per PCNA trimer. The Fen1-PCNA association is seen in cell extracts without overexpression of either partner and is mediated by a basic region at the C terminus of Fen1. Therefore, the polymerase delta-PCNA-Fen1 complex has all the activities associated with prokaryotic DNA polymerases involved in replication: 5'-3' polymerase, 3'-5' exonuclease, and 5'-3' exonuclease. Although p21, a regulatory protein induced by p53 in response to DNA damage, interacts with PCNA with a comparable Kd (10 nM) and a stoichiometry of three molecules of p21 per PCNA trimer, a p21-PCNA-Fen1 complex is not formed. This mutually exclusive interaction suggests that the conformation of a PCNA trimer switches such that it can either bind p21 or Fen1. Furthermore, overexpression of p21 can disrupt Fen1-PCNA interaction in vivo. Therefore, besides interfering with the processivity of polymerase delta-PCNA, p21 also uncouples Fen1 from the PCNA scaffold.
Peptide-specific cytotoxic T lymphocytes restricted by nonself major histocompatibility complex class I molecules: Reagents for tumor immunotherapy
Authors: Sadovnikova Elena , Stauss Hans J.
Published: July 2002

Studies in melanoma patients have revealed that self proteins can function as targets for tumor-reactive cytotoxic T lymphocytes (CTL). One group of self proteins MAGE, BAGE, and GAGE are normally only expressed in testis and placenta, whilst another group of CTL recognized proteins are melanocyte-specific differentiation antigens. In this study we have investigated whether CTL can be raised against a ubiquitously expressed self protein, mdm-2, which is frequently overexpressed in tumors. The observation that T-cell tolerance is self major histocompatibility complex-restricted was exploited to generate CTL specific for an mdm-2 derived peptide presented by nonself major histocompatibility complex class I molecules. Thus, the allo-restricted T-cell repertoire of H-2dmice was used to isolate CTL specific for the mdm100 peptide presented by allogeneic H-2Kbclass I molecules.In vitro, these CTL discriminated between transformed and normal cells, killing specifically Kb-positive melanoma and lymphoma tumors but not Kb-expressing dendritic cells.In vivo, the CTL showed antitumor activity and delayed the growth of melanoma as well as lymphoma tumors in H-2brecipient mice. These experiments show that it is possible to circumvent T-cell tolerance to ubiquitously expressed self antigens, and to target CTL responses against tumors expressing elevated levels of structurally unaltered proteins.
Steel mutant mice are deficient in hippocampal learning but not long-term potentiation.
Authors: Motro B , Wojtowicz J M , Bernstein A , van der Kooy D
Published: July 2002

Mice carrying mutations in either the dominant white-spotting (W) or Steel (Sl) loci exhibit deficits in melanogenesis, gametogenesis, and hematopoiesis. W encodes the Kit receptor tyrosine kinase, while Sl encodes the Kit ligand, Steel factor, and the receptor-ligand pair are contiguously expressed at anatomical sites expected from the phenotypes of W and Sl mice. The c-kit and Steel genes are also both highly expressed in the adult murine hippocampus: Steel is expressed in dentate gyrus neurons whose mossy fiber axons synapse with the c-kit expressing CA3 pyramidal neurons. We report here that Sl/Sld mutant mice have a specific deficit in spatial learning. These mutant mice are also deficient in baseline synaptic transmission between the dentate gyrus and CA3 but show normal long-term potentiation in this pathway. These observations demonstrate a role for Steel factor/Kit signaling in the adult nervous system and suggest that a severe deficit in hippocampal-dependent learning need not be associated with reduced hippocampal long-term potentiation.
Transcription of the lymphocyte-specific terminal deoxynucleotidyltransferase gene requires a specific core promoter structure.
Authors: Garraway I P , Semple K , Smale S T
Published: July 2002

The terminal deoxynucleotidyltransferase (TdT) gene encodes a template-independent DNA polymerase that is expressed exclusively in immature lymphocytes. The TdT promoter lacks a TATA box, but an initiator element (Inr) overlaps the transcription start site. The Inr directs basal transcription and also mediates activated transcription in conjunction with an upstream element called D'. We have begun to address the fundamental question of why the TdT promoter contains an Inr rather than a TATA box. First, we tested the possibility that the TdT promoter lacks a TATA box because the -30 region is needed for the binding of an essential regulator. Mutations were introduced into the -30 region, and the mutants were tested in transient transfection and in vitro transcription assays. The mutations had only minor effects on promoter strength, suggesting that this first hypothesis is incorrect. Next, the effect of inserting a TATA box within the -30 region was tested. Although the TATA box enhanced promoter strength, appropriate regulation appeared to be maintained, as transcription in lymphocytes remained dependent on the D' element. Finally, a promoter variant containing a TATA box at -30, but a mutant Inr, was tested. Surprisingly, transcription from this variant, both in vitro and in vivo, was dramatically reduced. These results suggest that the TdT promoter, and possibly other natural promoters, contain an Inr element because one or more activator proteins that interact with surrounding control elements preferentially function in its presence.
Mutator tRNAs are encoded by the Escherichia coli mutator genes mutA and mutC: a novel pathway for mutagenesis.
Authors: Slupska M M , Baikalov C , Lloyd R , Miller J H
Published: July 2002

We have previously described the mutator alleles mutA and mutC, which map at 95 minutes and 42 minutes, respectively, on the Escherichia coli genetic map and which stimulate transversions; the A.T-->T.A and G.C-->T.A substitutions are the most prominent. In this study we show that both mutA and mutC result from changes in the anticodon in one of four copies of the same glycine tRNA, at either the glyV or the glyW locus. This change results in a tRNA that inserts glycine at aspartic acid codons. In view of previous studies of missense suppressor tRNAs, the mistranslation of aspartic acid codons is assumed to occur at approximately 1-2%. We postulate that the mutator tRNA effect is exerted by generating a mutator polymerase and suggest that the epsilon subunit of DNA polymerase, which provides a proofreading function, is the most likely target. The implications of these findings for the contribution of mistranslation to observed spontaneous mutation rates in wild-type strains, as well as other cellular phenomena such as aging, are discussed.
A demographic approach to selection
Authors: Anderson Wyatt W. , Watanabe Takao K.
Published: July 2002

The concepts of demography provide a means of combining the
ecological approach to population growth with the genetical approach to
natural selection. We have utilized the demographic theory of natural
selection developed by Norton and Charlesworth to analyze life history
schedules of births and deaths for populations of genotypes in
Drosophila pseudoobscura
. Our populations illustrate a
stable genetic equilibrium, an unstable genetic equilibrium, and a case
of no equilibrium. We have estimated population growth rates and
Darwinian fitnesses of the genotypes and have explored the role of
population growth in determining natural selection. The age-specific
rates of births and deaths provide insights into components of
selection. Both viability and fertility are important components in our
populations.
Direct involvement of the ubiquitin-conjugating enzyme Ubc9/Hus5 in the degradation of IκBα
Authors: Tashiro Kei , Pando Matthew P. , Kanegae Yumi , Wamsley Penny M. , Inoue Satoshi , Verma Inder M.
Published: July 2002

The NF-κB/Rel proteins are sequestered in the cytoplasm in association with IκBα. In response to external signals, IκBα is phosphorylated, multi-ubiquitinated, and degraded by proteasomes, thereby releasing NF-κB/Rel proteins to migrate to the nucleus. We have cloned a mouse ubiquitin-conjugating enzyme (mE2), which associates with IκBα. mE2 is homologous to the yeast Ubc9/Hus5 ubiquitin-conjugating enzyme. A transdominant-negative mutant of mE2 had no effect on phosphorylation of IκBα, but delayed its degradation. Correspondingly, tumor necrosis factor-α-inducible NF-κB activity was diminished. We propose that mE2 is directly involved in the ubiquitin conjugation of IκBα, a pivotal step in its degradation pathway.
Cell cycle-dependent colocalization of BARD1 and BRCA1 proteins in discrete nuclear domains
Authors: Jin Ying , Xu Xie L. , Yang Meng-Chun W. , Wei Fanglin , Ayi Teck-Choon , Bowcock Anne M. , Baer Richard
Published: July 2002

Germ-line mutations of the
BRCA1
gene predispose women to early-onset breast and ovarian cancer by compromising the gene’s presumptive function as a tumor suppressor. Although the biochemical properties of BRCA1 polypeptides are not understood, their expression pattern and subcellular localization suggest a role in cell-cycle regulation. When resting cells are induced to proliferate, the steady-state levels of BRCA1 increase in late G
1
and reach a maximum during S phase. Moreover, in S phase cells, BRCA1 polypeptides are hyperphosphorylated and accumulate into discrete subnuclear foci termed “BRCA1 nuclear dots.” BRCA1 associates
in vivo
with a structurally related protein termed BARD1. Here we show that the steady-state levels of BARD1, unlike those of BRCA1, remain relatively constant during cell cycle progression. However, immunostaining revealed that BARD1 resides within BRCA1 nuclear dots during S phase of the cell cycle, but not during the G
1
phase. Nevertheless, BARD1 polypeptides are found exclusively in the nuclear fractions of both G
1
- and S-phase cells. Therefore, progression to S phase is accompanied by the aggregation of nuclear BARD1 polypeptides into BRCA1 nuclear dots. This cell cycle-dependent colocalization of BARD1 and BRCA1 indicates a role for BARD1 in BRCA1-mediated tumor suppression.
The analysis of ontogenetic trajectories: When a change in size or shape is not heterochrony
Authors: Rice Sean H.
Published: July 2002

Heterochrony has become a central organizing concept relating development and evolution. Unfortunately, the standard definition of heterochrony—evolutionary change in the rate or timing of developmental processes—is so broad as to apply to any case of phenotypic evolution. Conversely, the standard classes of heterochrony only accurately describe a small subset of the possible ways that ontogeny can change. I demonstrate here that the nomenclature of heterochrony is meaningful only when there is a uniform change in the rate or timing of some ontogenetic process, with no change in the internal structure of that process. Given two ontogenetic trajectories, we can test for this restricted definition of heterochrony by asking if a uniform stretching or translation of one trajectory along the time axis superimposes it on the other trajectory. If so, then the trajectories are related by a uniform change in the rate or timing of development. If not, then there has been change within the ontogenetic process under study. I apply this technique to published data on fossil Echinoids and to the comparison of human and chimpanzee growth curves. For the Echinoids, some characters do show heterochrony (hypermorphosis), while others, which had previously been seen as examples of heterochrony, fail the test—implying that their evolution involved changes in the process of development, not just the rate at which it proceeded. Analysis of human and chimpanzee growth curves indicates a combination of neoteny and sequential hypermorphosis, two processes previously seen as alternate explanations for the differences between these species.
GTP-bound forms of rab6 induce the redistribution of Golgi proteins into the endoplasmic reticulum
Authors: Martinez Olivier , Antony Claude , Pehau-Arnaudet Gérard , Berger Eric G. , Salamero Jean , Goud Bruno
Published: July 2002

rab6 is a ubiquitous ras-like GTPase involved in intra-Golgi transport. We have studied at both morphological and biochemical levels the behavior of Golgi resident proteins in HeLa cells overexpressing wild-type rab6 and GTP- and GDP-bound mutants of rab6 (rab6 Q72L and rab6 T27N, respectively). We show that wild-type rab6 and rab6 Q72L overexpression induces the redistribution of the trans-Golgi protein β-1,4-galactosyltransferase into the endoplasmic reticulum (ER) and allows the addition of sialylated O-glycans on an ER-retained protein, the major histocompatibility complex class II-associated invariant chain. Remarkably, rab6 Q72L effects, which require the integrity of microtubules, were almost indistinguishable from those induced by brefeldin A, a fungic metabolite that causes a mixing of Golgi and ER membranes. In contrast, overexpression of rab6 T27N does not cause the redistribution of Golgi proteins, but inhibits basal O-glycosylation of the major histocompatibility complex class II-associated invariant chain.
Transgenic expression of CD95 ligand on islet β cells induces a granulocytic infiltration but does not confer immune privilege upon islet allografts
Authors: Allison Janette , Georgiou Harry M. , Strasser Andreas , Vaux David L.
Published: July 2002

Binding of CD95 (Fas/APO-1) by its ligand (CD95L) commonly induces apoptosis. Apoptosis of activated T cells, induced by CD95L expressed in the rodent testis, has been proposed to be the mechanism of immune privilege [Bellgrau, D., Gold, D., Selawry, H., Moore, J., Franzusoff, A. & Duke, R. C. (1995)Nature (London)377, 630–632]. To test whether CD95L could protect pancreatic islet grafts from rejection, we made transgenic mice expressing murine CD95L on their islet β cells and transplanted fetal pancreata under the kidney capsules of allogeneic animals. Expression of CD95L failed to protect the grafts from rejection. However, transgenic mice developed a granulocytic infiltration in their pancreata. These results demonstrate a pro-inflammatory function of CD95L and suggest that expression of CD95L may not be sufficient to protect organ allografts.
A steroid-triggered switch in E74 transcription factor isoforms regulates the timing of secondary-response gene expression
Authors: Fletcher Jennifer C. , D’Avino Pier Paolo , Thummel Carl S.
Published: July 2002

The steroid hormone 20-hydroxyecdysone (referred to here as ecdysone) directs
Drosophila
metamorphosis by activating a series of genetic regulatory hierarchies. ETS domain transcription factors encoded by the ecdysone-inducible
E74
early gene, E74A and E74B, act at the top of these hierarchies to coordinate the induction of target genes. We have ectopically expressed these E74 isoforms to understand their regulatory functions during the onset of metamorphosis. We show that
E74
can regulate its own transcription, most likely through binding sites within its gene. Ectopic expression of
E74B
can partially repress the
E78B
and
DHR3
orphan receptor genes, suggesting a role for
E74
in the appropriate timing of early-late gene expression. Furthermore,
E74A
is both necessary and sufficient for
E78B
induction, implicating
E74A
as a key regulator of
E78B
expression. We also show, consistent with our studies of
E74
loss-of-function mutations, that E74B is a potent repressor of late gene transcription and E74A is sufficient to prematurely induce the
L71–1
late gene. However, ectopic expression of both Broad-Complex and E74A activators in an
E74B
mutant background is not sufficient to prematurely induce all late genes, indicating that other factors contribute to this regulatory circuit. These observations demonstrate that the steroid-triggered switch in E74 transcription factor isoforms plays a central role in the proper timing of secondary-response gene expression.
p53-dependent and -independent responses to cisplatin in mouse testicular teratocarcinoma cells
Authors: Zamble Deborah B. , Jacks Tyler , Lippard Stephen J.
Published: July 2002

Testicular cancers respond favorably to chemotherapy with the platinum-containing drug
cis
-diamminedichloroplatinum(II) (cisplatin). One factor that could explain the efficacy of cisplatin is the low frequency of p53 mutations observed in this tumor type. The present study examines the p53-mediated responses in murine testicular teratocarcinoma cells exposed to the drug. Cisplatin treatment of teratocarcinoma cells with a wild-type p53 gene resulted in accumulation of the p53 protein through posttranscriptional mechanisms; induction of p53-target genes was also observed. Drug treatment resulted in rapid apoptosis in p53-wild-type cells but not in p53
−/−
teratocarcinoma cells. In the latter cells, cisplatin exposure caused prolonged cell cycle arrest accompanied by induction of the p21 gene. Clonogenic assays demonstrated that the p53 mutation did not confer resistance to cisplatin. These experiments suggest that cisplatin inhibits cellular proliferation of testicular teratocarcinoma cells by two possible mechanisms, p53-dependent apoptosis and p53-independent cell cycle arrest.
Aberrant intracellular localization of Varicella-Zoster virus regulatory proteins during latency
Authors: Lungu Octavian , Panagiotidis Christos A. , Annunziato Paula W. , Gershon Anne A. , Silverstein Saul J.
Published: July 2002

Varicella-Zoster virus (VZV) is a herpesvirus that becomes latent in sensory neurons after primary infection (chickenpox) and subsequently may reactivate to cause zoster. The mechanism by which this virus maintains latency, and the factors involved, are poorly understood. Here we demonstrate, by immunohistochemical analysis of ganglia obtained at autopsy from seropositive patients without clinical symptoms of VZV infection that viral regulatory proteins are present in latently infected neurons. These proteins, which localize to the nucleus of cells during lytic infection, predominantly are detected in the cytoplasm of latently infected neurons. The restriction of regulatory proteins from the nucleus of latently infected neurons might interrupt the cascade of virus gene expression that leads to a productive infection. Our findings raise the possibility that VZV has developed a novel mechanism for maintenance of latency that contrasts with the transcriptional repression that is associated with latency of herpes simplex virus, the prototypic alpha herpesvirus.