bluuebunny/crossref_metadata_2025 huggingface dataset:
Self-assembly of a heteroduplex helicate from two different ligand strands and Cu(II) cations.
Authors: Hasenknopf B , Lehn J M , Baum G , Fenske D
Published: July 2002

Cu(II) ions have been reacted with a 1/1 mixture of two linear ligands, one containing three 2,2'- bipyridine groups and the other three 2,2':6',2"-terpyridine groups. Absorption spectroscopy and fast atom bombardment mass spectrometry indicate the formation of a trinuclear complex containing one ligand of each kind. Determination of the crystal structure of this compound has confirmed that it is indeed a linear trinuclear complex in which two different ligands are wrapped in a helical fashion around the pentacoordinated metal ions. The central coordination geometry is trigonal bipyramidal; the two lateral Cu(II) ions are in a square pyramidal environment. Thus, a heteroduplex helicate is formed by the self-assembly of two different ligand strands and three specific metal ions induced by the coordination number and geometry of the latter. The self-assembly process may be considered to result from the reading of the steric and binding information present in the two ligands by Cu(II) ions through a pentacoordination algorithm. The same ligands have been shown earlier to yield homoduplex helicates from ions of tetrahedral and octahedral coordination geometry and strands of bidentate bipyridines and tridentate terpyridines, respectively. These two types of artificial double helical species may be related on one hand to the natural homoduplex nucleic acids and on the other hand to the DNA:RNA heteroduplex.
Trinucleotide repeats and long homopeptides in genes and proteins associated with nervous system disease and development.
Authors: Karlin S , Burge C
Published: July 2002

Several human neurological disorders are associated with proteins containing abnormally long runs of glutamine residues. Strikingly, most of these proteins contain two or more additional long runs of amino acids other than glutamine. We screened the current human, mouse, Drosophila, yeast, and Escherichia coli protein sequence data bases and identified all proteins containing multiple long homopeptides. This search found multiple long homopeptides in about 12% of Drosophila proteins but in only about 1.7% of human, mouse, and yeast proteins and none among E. coli proteins. Most of these sequences show other unusual sequence features, including multiple charge clusters and excessive counts of homopeptides of length > or = two amino acid residues. Intriguingly, a large majority of the identified Drosophila proteins are essential developmental proteins and, in particular, most play a role in central nervous system development. Almost half of the human and mouse proteins identified are homeotic homologs. The role of long homopeptides in fine-tuning protein conformation for multiple functional activities is discussed. The relative contributions of strand slippage and of dynamic mutation are also addressed. Several new experiments are proposed.
Oxygen supply and oxygen-dependent gene expression in differentiating embryonic stem cells.
Authors: Gassmann M , Fandrey J , Bichet S , Wartenberg M , Marti H H , Bauer C , Wenger R H , Acker H
Published: July 2002

Blastocyst-derived pluripotent mouse embryonic stem cells can differentiate in vitro to form so-called embryoid bodies (EBs), which recapitulate several aspects of murine embryogenesis. We used this in vitro model to study oxygen supply and consumption as well as the response to reduced oxygenation during the earliest stages of development. EBs were found to grow equally well when cultured at 20% (normoxia) or 1% (hypoxia) oxygen during the first 5 days of differentiation. Microelectrode measurements of pericellular oxygen tension within 13- to 14-day-old EBs (diameter 510-890 micron) done at 20% oxygen revealed efficient oxygenation of the EBs' core region. Confocal laser scanning microscopy analysis of EBs incubated with fluorescent dyes that specifically stain living cells confirmed that the cells within an EB were viable. To determine the EBs' capability to sense low oxygen tension and to specifically respond to low ambient oxygen by modulating gene expression we quantified aldolase A and vascular endothelial growth factor (VEGF) mRNAs, since expression of these genes is upregulated by hypoxia in a variety of cells. Compared with the normoxic controls, we found increased aldolase A and VEGF mRNA levels after exposing 8- to 9-day-old EBs to 1% oxygen. We propose that EBs represent a powerful tool to study oxygen-regulated gene expression during the early steps of embryogenesis, where the preimplantation conceptus resides in a fluid environment with low oxygen tension until implantation and vascularization allow efficient oxygenation.
Fumonisins and Alternaria alternata lycopersici toxins: sphinganine analog mycotoxins induce apoptosis in monkey kidney cells.
Authors: Wang W , Jones C , Ciacci-Zanella J , Holt T , Gilchrist D G , Dickman M B
Published: July 2002

Fusarium moniliforme toxins (fumonisins) and Alternaria alternata lycopersici (AAL) toxins are members of a new class of sphinganine analog mycotoxins that occur widely in the food chain. These mycotoxins represent a serious threat to human and animal health, inducing both cell death and neoplastic events in mammals. The mechanisms by which this family of chemical congeners induce changes in cell homeostasis were investigated in African green monkey kidney cells (CV-1) by assessing the appearance of apoptosis, cell cycle regulation, and putative components of signal transduction pathways involved in apoptosis. Structurally, these mycotoxins resemble the sphingoid bases, sphingosine and sphinganine, that are reported to play critical roles in cell communication and signal transduction. The addition of fumonisin B1 or AAL toxin, TA, to CV-1 cells induced the stereotypical hallmarks of apoptosis, including the formation of DNA ladders, compaction of nuclear DNA, and the subsequent appearance of apoptotic bodies. Neither mycotoxin induced cell death, DNA ladders, or apoptotic bodies in CV-1 cells expressing simian virus 40 large T antigen (COS-7) at toxin concentrations that readily killed CV-1 cells. Fumonisin B1 induced cell cycle arrest in the G1 phase in CV-1 cells but not in COS-7 cells. AAL toxin TA did not arrest cell cycle progression in either cell line. The induction of apoptosis combined with the widespread presence of these compounds in food crops and animal feed identifies a previously unrecognized health risk to humans and livestock. These molecules also represent a new class of natural toxicants that can be used as model compounds to further characterize the molecular and biochemical pathways leading to apoptosis.
Population dynamics, demographic stochasticity, and the evolution of cooperation
Authors: Doebeli Michael , Blarer Albert , Ackermann Martin
Published: July 2002

A basic evolutionary problem posed by the Iterated Prisoner’s Dilemma game is to understand when the paradigmatic cooperative strategy Tit-for-Tat can invade a population of pure defectors. Deterministically, this is impossible. We consider the role of demographic stochasticity by embedding the Iterated Prisoner’s Dilemma into a population dynamic framework. Tit-for-Tat can invade a population of defectors when their dynamics exhibit short episodes of high population densities with subsequent crashes and long low density periods with strong genetic drift. Such dynamics tend to have reddened power spectra and temporal distributions of population size that are asymmetric and skewed toward low densities. The results indicate that ecological dynamics are important for evolutionary shifts between adaptive peaks.
The leukocyte response to fluid stress
Authors: Moazzam Fariborz , DeLano Frank A. , Zweifach Benjamin W. , Schmid-Schönbein Geert W.
Published: July 2002

Leukocyte migration from a hemopoietic pool across marrow endothelium requires active pseudopod formation and adhesion. Leukocytes rarely show pseudopod formation while in circulation. At question then is the mechanism that serves to minimize leukocyte pseudopod formation in the circulation. We tested the hypothesis that fluid shear stress acts to prevent pseudopod formation. When individual human leukocytes (neutrophils, monocytes) spreading on glass surfaces
in vitro
were subjected to fluid shear stress (≈1 dyn/cm
2
), an instantaneous retraction of pseudopods was observed. Removal of the fluid shear stress in turn led to the return of pseudopod projection and cell spreading. When steady shear stress was prolonged over several minutes, leukocyte swelling occurs together with an enhanced random motion of cytoplasmic granules and a reduction of cytoplasmic stiffness. The response to shear stress could be suppressed by K
+
channel blockers and chelation of external Ca
2+
. In rat mesentery microvessels after occlusion, circulating leukocytes project pseudopods in free suspension or when attached to the endothelium, even though immediately after occlusion only few pseudopods were present. When flow was restored, pseudopods on adhering leukocytes were retracted and then the cells began to roll and detach from the endothelium. In conclusion, plasma shear stress in the circulation serves to reduce pseudopod projection and adhesion of circulating leukocytes and vice versa reduction of shear stress leads to pseudopod projection and spreading of leukocytes on the endothelium.
Reaction of aflatoxin B
1
exo
-8,9-epoxide with DNA: Kinetic analysis of covalent binding and DNA-induced hydrolysis
Authors: Johnson William W. , Guengerich F. Peter
Published: July 2002

The
exo
isomer of aflatoxin B
1
(AFB
1
) 8,9-epoxide appears to be the only product of AFB
1
involved in reaction with DNA and reacts with the
N
7
atom of guanine via an S
N
2 reaction from an intercalated state. Although the epoxide hydrolyzes rapidly in H
2
O (0.6 s
−1
at 25°C), very high yields of DNA adduct result. Experimental binding data were fit to a model in which the epoxide forms a reversible complex with calf thymus DNA (
K
d
= 0.43 mg ml
−1
, or 1.4 mM monomer equivalents) and reacts with guanine with a rate of 35 s
−1
. Stopped-flow kinetic analysis revealed attenuation of fluorescence in the presence of DNA that was dependent on DNA concentration. Kinetic spectral analysis revealed that this process represents conjugation of epoxide with DNA, with an extrapolated rate maximum of 42 s
−1
and half-maximal velocity at a DNA concentration of 1.8 mg ml
−1
(5.8 mM monomer equivalents). The rate of hydrolysis of the epoxide was accelerated by calf thymus DNA in the range of pH 6–8, with a larger enhancement at the lower pH (increase of 0.23 s
−1
at pH 6.2 with 0.17 mg DNA ml
−1
). The same rate enhancement effect was observed with poly[dA-dT]⋅poly[dA-dT], in which the epoxide can intercalate but not form significant levels of
N
7
purine adducts, and with single-stranded DNA. The increased rate of hydrolysis by DNA resembles that reported earlier for epoxides of polycyclic hydrocarbons and is postulated to involve a previously suggested localized proton field on the periphery of DNA. The epoxide preferentially intercalates between base pairs, and the proton field is postulated to provide acid catalysis to the conjugation reaction.
Deficiency of a protein-repair enzyme results in the accumulation of altered proteins, retardation of growth, and fatal seizures in mice
Authors: Kim Edward , Lowenson Jonathan D. , MacLaren Duncan C. , Clarke Steven , Young Stephen G.
Published: July 2002

l
-Asparaginyl and
l
-aspartyl residues in proteins are subject to spontaneous degradation reactions that generate isomerized and racemized aspartyl derivatives. Proteins containing
l
-isoaspartyl and
d
-aspartyl residues can have altered structures and diminished biological activity. These residues are recognized by a highly conserved cytosolic enzyme, the protein
l
-isoaspartate(
d
-aspartate)
O
-methyltransferase (EC
2.1.1.77
). The enzymatic methyl esterification of these abnormal residues
in vitro
can lead to their conversion (i.e., repair) to normal
l
-aspartyl residues and should therefore prevent the accumulation of potentially dysfunctional proteins
in vivo
as cells and tissues age. Particularly high levels of the repair methyltransferase are present in the brain, although enyzme activity is present in all vertebrate tissues. To define the physiological relevance of this protein-repair pathway and to determine whether deficient protein repair would cause central nervous system dysfunction, we used gene targeting in mouse embryonic stem cells to generate protein
l
-isoaspartate(
d
-aspartate)
O
-methyltransferase-deficient mice. Analyses of tissues from methyltransferase knockout mice revealed a striking accumulation of protein substrates for this enzyme in the cytosolic fraction of brain, heart, liver, and erythrocytes. The knockout mice showed significant growth retardation and succumbed to fatal seizures at an average of 42 days after birth. These results suggest that the ability of mice to repair
l
-isoaspartyl- and
d
-aspartyl-containing proteins is essential for normal growth and for normal central nervous system function.
A mouse model for X-linked adrenoleukodystrophy
Authors: Lu Jyh-Feng , Lawler Ann M. , Watkins Paul A. , Powers James M. , Moser Ann B. , Moser Hugo W. , Smith Kirby D.
Published: July 2002

X-linked adrenoleukodystrophy (X-ALD) is a peroxisomal disorder with impaired β-oxidation of very long chain fatty acids (VLCFAs) and reduced function of peroxisomal very long chain fatty acyl-CoA synthetase (VLCS) that leads to severe and progressive neurological disability. The X-ALD gene, identified by positional cloning, encodes a peroxisomal membrane protein (adrenoleukodystrophy protein; ALDP) that belongs to the ATP binding cassette transporter protein superfamily. Mutational analyses and functional studies of the X-ALD gene confirm that it and not VLCS is the gene responsible for X-ALD. Its role in the β-oxidation of VLCFAs and its effect on the function of VLCS are unclear. The complex pathology of X-ALD and the extreme variability of its clinical phenotypes are also unexplained. To facilitate understanding of X-ALD pathophysiology, we developed an X-ALD mouse model by gene targeting. The X-ALD mouse exhibits reduced β-oxidation of VLCFAs, resulting in significantly elevated levels of saturated VLCFAs in total lipids from all tissues measured and in cholesterol esters from adrenal glands. Lipid cleft inclusions were observed in adrenocortical cells of X-ALD mice under the electron microscope. No neurological involvement has been detected in X-ALD mice up to 6 months. We conclude that X-ALD mice exhibit biochemical defects equivalent to those found in human X-ALD and thus provide an experimental system for testing therapeutic intervention.
CP12 provides a new mode of light regulation of Calvin cycle activity in higher plants
Authors: Wedel Norbert , Soll Jürgen , Paap Brigitte K.
Published: July 2002

CP12 is a small nuclear encoded chloroplast protein of higher plants, which was recently shown to interact with NAD(P)H–glyceraldehyde-3-phosphate dehydrogenase (GAPDH; EC
1.2.1.13
), one of the key enzymes of the reductive pentosephosphate cycle (Calvin cycle). Screening of a pea cDNA library in the yeast two-hybrid system for proteins that interact with CP12, led to the identification of a second member of the Calvin cycle, phosphoribulokinase (PRK; EC
2.7.1.19
), as a further specific binding partner for CP12. The exchange of cysteines for serines in CP12 demonstrate that interaction with PRK occurs at the N-terminal peptide loop of CP12. Size exclusion chromatography and immunoprecipitation assays reveal the existence of a stable 600-kDa PRK/CP12/GAPDH complex in the stroma of higher plant chloroplasts. Its stoichiometry is proposed to be of two N-terminally dimerized CP12 molecules, each carrying one PRK dimer on its N terminus and one A2B2 complex of GAPDH subunits on the C-terminal peptide loop. Incubation of the complex with NADP or NADPH, in contrast to NAD or NADH, causes its dissociation. Assays with the stromal 600-kDa fractions in the presence of the four different nicotinamide-adenine dinucleotides indicate that PRK activity depends on complex dissociation and might be further regulated by the accessible ratio of NADP/NADPH. From these results, we conclude that light regulation of the Calvin cycle in higher plants is not only via reductive activation of different proteins by the well-established ferredoxin/thioredoxin system, but in addition, by reversible dissociation of the PRK/CP12/GAPDH complex, mediated by NADP(H).
Absence of granulocyte colony-stimulating factor signaling and neutrophil development in CCAAT enhancer binding protein α-deficient mice
Authors: Zhang Dong-Er , Zhang Pu , Wang Nai-dy , Hetherington Christopher J. , Darlington Gretchen J. , Tenen Daniel G.
Published: July 2002

Transcription factors are master regulatory switches of differentiation, including the development of specific hematopoietic lineages from stem cells. Here we show that mice with targeted disruption of the CCAAT enhancer binding protein α gene (C/EBPα) demonstrate a selective block in differentiation of neutrophils. Mature neutrophils and eosinophils are not observed in the blood or fetal liver of mutant animals, while other hematopoietic lineages, including monocytes, are not affected. Instead, most of the white cells in the peripheral blood of mutant mice had the appearance of myeloid blasts. We also observed a selective loss of expression of a critical gene target of CCAAT enhancer binding protein α, the granulocyte colony-stimulating factor receptor. As a result, multipotential myeloid progenitors from the mutant fetal liver are unable to respond to granulocyte colony-stimulating factor signaling, although they are capable of forming granulocyte–macrophage and macrophage colonies in methylcellulose in response to other growth factors. Finally, we demonstrate that the lack of granulocyte development results from a defect intrinsic to the hematopoietic system; transplanted fetal liver from mutant mice can reconstitute lymphoid but not neutrophilic cells in irradiated recipients. These studies suggest a model by which transcription factors can direct the differentiation of multipotential precursors through activation of expression of a specific growth factor receptor, allowing proliferation and differentiation in response to a specific extracellular signal. In addition, thec/ebpα−/−mice may be useful in understanding the mechanisms involved in acute myelogenous leukemia, in which a block in differentiation of myeloid precursors is a key feature of the disease.
The efficiency of propulsion by a rotating flagellum
Authors: Purcell Edward M.
Published: July 2002

[At very low Reynolds number, the regime in which fluid dynamics
is governed by Stokes equations, a helix that translates along its axis
under an external force but without an external torque will necessarily
rotate. By the linearity of the Stokes equations, the same helix that
is caused to rotate due to an external torque will necessarily
translate. This is the physics that underlies the mechanism of
flagellar propulsion employed by many microorganisms. Here, I examine
the linear relationships between forces and torques and translational
and angular velocities of helical objects to understand the nature of
flagellar propulsion.]
Phylogenetic relationships between the Acantharea and the Polycystinea: A molecular perspective on Haeckel’s Radiolaria
Authors: Zettler Linda Amaral , Sogin Mitchell L. , Caron David A.
Published: July 2002

Polycystine radiolaria are among few protistan groups that possess a comprehensive fossil record available for study by micropaleontologists. The Polycystinea and the Acantharea, whose skeletons do not become fossilized, were once members of the class “Radiolaria” (“Radiolaria”
sensu lato
: Polycystinea, Phaeodarea, and Acantharea) originally proposed by Haeckel but are now included in the superclass Actinopoda. Phylogenetic relationships within this superclass remain largely enigmatic. We investigated the evolutionary relationship of the Acantharea and the Polycystinea to other protists using phylogenetic analyses of 16S-like ribosomal RNA (rRNA) coding regions. We circumvented the need to culture these organisms by collecting and maintaining reproductive stages that contain many copies of their genomic DNA. This strategy facilitated extraction of genomic DNA and its purification from symbiont and prey DNA. Phylogenetic trees inferred from comparisons of 16S-like coding regions do not support a shared history between the Acantharea and the Polycystinea. However, the monophyly of the Acantharea and the separate monophyly of the Polycystinea (Spumellarida) are well supported by our molecular-based trees. The acantharian lineage branches among crown organisms whereas the polycystine lineage diverges before the radiation of the crown groups. We conclude that the Actinopoda does not represent a monophyletic evolutionary assemblage and recommend that this taxonomic designation be discarded.
Isolation and characterization of neutral-lipid-containing organelles and globuli-filled plastids from
Brassica napus
tapetum
Authors: Wu Sherry S. H. , Platt Kathryn A. , Ratnayake Chandra , Wang Tzann-Wei , Ting Julie T. L. , Huang Anthony H. C.
Published: July 2002

The monolayer tapetum cells of the maturing flowers of
Brassica napus
contain abundant subcellular globuli-filled plastids and special lipid particles, both enriched with lipids that are supposed to be discharged and deposited onto the surface of adjacent maturing pollen. We separated the two organelles by flotation density gradient centrifugation and identified them by electron microscopy. The globuli-filled plastids had a morphology similar to those described in other plant species and tissues. They had an equilibrium density of 1.02 g/cm
3
and contained neutral esters and unique polypeptides. The lipid particles contained patches of osmiophilic materials situated among densely packed vesicles and did not have an enclosing membrane. They exhibited osmotic properties, presumably exerted by the individual vesicles. They had an equilibrium density of 1.05 g/cm
3
and possessed triacylglycerols and unique polypeptides. Several of these polypeptides were identified, by their N-terminal sequences or antibody cross-reactivity, as oleosins, proteins known to be associated with seed storage oil bodies. The morphological and biochemical characteristics of the lipid particles indicate that they are novel organelles in eukaryotes that have not been previously isolated and studied. After lysis of the tapetum cells at a late stage of floral development, only the major plastid neutral ester was recovered, whereas the other abundant lipids and proteins of the two tapetum organelles were present in fragmented forms or absent on the pollen surface.
α
+
-Thalassemia protects children against disease caused by other infections as well as malaria
Authors: Allen S. J. , O’Donnell A. , Alexander N. D. E. , Alpers M. P. , Peto T. E. A. , Clegg J. B. , Weatherall D. J.
Published: July 2002

In the South West Pacific region, the striking geographical correlation between the frequency of α
+
-thalassemia and the endemicity of
Plasmodium falciparum
suggests that this hemoglobinopathy provides a selective advantage against malaria. In Vanuatu, paradoxically, α
+
-thalassemia increases the incidence of contracting mild malaria in the first 2 years of life, but severe disease was too uncommon to assess adequately. Therefore, we undertook a prospective case-control study of children with severe malaria on the north coast of Papua New Guinea, where malaria transmission is intense and α
+
-thalassemia affects more than 90% of the population. Compared with normal children, the risk of having severe malaria was 0.40 (95% confidence interval 0.22–0.74) in α
+
-thalassemia homozygotes and 0.66 (0.37–1.20) in heterozygotes. Unexpectedly, the risk of hospital admission with infections other than malaria also was reduced to a similar degree in homozygous (0.36; 95% confidence interval 0.22–0.60) and heterozygous (0.63; 0.38–1.07) children. This clinical study demonstrates that a malaria resistance gene protects against disease caused by infections other than malaria. The mechanism of the remarkable protective effect of α
+
-thalassemia against severe childhood disease remains unclear but must encompass the clear interaction between this hemoglobinopathy and both malarial and nonmalarial infections.
egl-17
encodes an invertebrate fibroblast growth factor family member required specifically for sex myoblast migration in
Caenorhabditis elegans
Authors: Burdine Rebecca D. , Chen Estella B. , Kwok Shing F. , Stern Michael J.
Published: July 2002

The proper guidance of the
Caenorhabditis elegans
hermaphrodite sex myoblasts (SMs) requires the genes
egl-15
and
egl-17. egl-15
has been shown to encode the
C. elegans
orthologue of the fibroblast growth factor receptor (FGFR). Here we clone
egl-17
and show it to be a member of the fibroblast growth factor (FGF) family, one of the first functional invertebrate FGFs known.
egl-17
shares homology with other FGF members, conserving the key residues required to form the distinctive tertiary structure common to FGFs. Genetic and molecular evidence demonstrates that the SM migration defect seen in
egl-17
mutant animals represents complete loss of
egl-17
function. While mutations in
egl-17
affect only SM migration, mutations in
egl-15
can result in larval arrest, scrawny body morphology, and the ability to suppress mutations in
clr-1
. We propose that EGL-17 (FGF) acts as a ligand for EGL-15 (FGFR) specifically during SM migration and that another ligand(s) activates EGL-15 for its other functions.
soc-2
encodes a leucine-rich repeat protein implicated in fibroblast growth factor receptor signaling
Authors: Selfors Laura M. , Schutzman Jennifer L. , Borland Christina Z. , Stern Michael J.
Published: July 2002

Activation of fibroblast growth factor (FGF) receptors elicits diverse cellular responses including growth, mitogenesis, migration, and differentiation. The intracellular signaling pathways that mediate these important processes are not well understood. In
Caenorhabditis elegans
, suppressors of
clr-1
identify genes, termed
soc
genes, that potentially mediate or activate signaling through the EGL-15 FGF receptor. We demonstrate that three
soc
genes,
soc-1
,
soc-2
, and
sem-5
, suppress the activity of an activated form of the EGL-15 FGF receptor, consistent with the
soc
genes functioning downstream of EGL-15. We show that
soc-2
encodes a protein composed almost entirely of leucine-rich repeats, a domain implicated in protein–protein interactions. We identified a putative human homolog, SHOC-2, which is 54% identical to SOC-2. We find that
shoc-2
maps to 10q25,
shoc-2
mRNA is expressed in all tissues assayed, and SHOC-2 protein is cytoplasmically localized. Within the leucine-rich repeats of both SOC-2 and SHOC-2 are two YXNX motifs that are potential tyrosine-phosphorylated docking sites for the SEM-5/GRB2 Src homology 2 domain. However, phosphorylation of these residues is not required for SOC-2 function
in vivo
, and SHOC-2 is not observed to be tyrosine phosphorylated in response to FGF stimulation. We conclude that this genetic system has allowed for the identification of a conserved gene implicated in mediating FGF receptor signaling in
C. elegans
.
Genetic evidence for a Paleolithic human population expansion in Africa
Authors: Reich David E. , Goldstein David B.
Published: July 2002

Human populations have undergone dramatic expansions in size, but other than the growth associated with agriculture, the dates and magnitudes of those expansions have never been resolved. Here, we introduce two new statistical tests for population expansion, which use variation at a number of unlinked genetic markers to study the demographic histories of natural populations. By analyzing genetic variation in various aboriginal populations from throughout the world, we show highly significant evidence for a major human population expansion in Africa, but no evidence of expansion outside of Africa. The inferred African expansion is estimated to have occurred between 49,000 and 640,000 years ago, certainly before the Neolithic expansions, and probably before the splitting of African and non-African populations. In showing a significant difference between African and non-African populations, our analysis supports the unique role of Africa in human evolutionary history, as has been suggested by most other genetic work. In addition, the missing signal in non-African populations may be the result of a population bottleneck associated with the emergence of these populations from Africa, as postulated in the “Out of Africa” model of modern human origins.
The structural protein ODV-EC27 of
Autographa californica
nucleopolyhedrovirus is a multifunctional viral cyclin
Authors: Belyavskyi Michail , Braunagel Sharon C. , Summers Max D.
Published: July 2002

Two major characteristics of baculovirus infection are arrest of the host cell at G
2
/M phase of the cell cycle with continuing viral DNA replication. We show that
Autographa californica
nucleopolyhedrovirus (AcMNPV) encodes for a multifunctional cyclin that may partially explain the molecular basis of these important characteristics of AcMNPV (baculovirus) infection. Amino acids 80–110 of the viral structural protein ODV-EC27 (−EC27) demonstrate 25–30% similarity with cellular cyclins within the cyclin box. Immunoprecipitation results using antibodies to −EC27 show that −EC27 can associate with either cdc2 or cdk6 resulting in active kinase complexes that can phosphorylate histone H1 and retinoblastoma protein
in vitro
. The cdk6-EC27 complex also associates with proliferating cell nuclear antigen (PCNA) and we demonstrate that PCNA is a structural protein of both the budded virus and the occlusion-derived virus. These results suggest that −EC27 can function as a multifunctional cyclin: when associated with cdc2, it exhibits cyclin B-like activity; when associated with cdk6, the complex possesses cyclin D-like activity and binds PCNA. The possible roles of such a multifunctional cyclin during the life cycle of baculovirus are discussed, along with potential implications relative to the expression of functionally authentic recombinant proteins by using baculovirus-infected cells.
Tracking the motion of chemisorbed molecules on their adsorption sites
Authors: Yates John T. , Ahner Joachim , Mocuta Dan
Published: July 2002

A new method, the time of flight–electron-stimulated desorption ion angular distribution method (TOF-ESDIAD), is described. The method permits the mapping of the lateral momentum distribution of adsorbed species on their adsorption sites. Examples of the study of the rotation of chemisorbed PF
3
molecules and the frustrated lateral translation of chemisorbed CO molecules, both on single-crystal metal surfaces, are given. The observed anisotropy of the frustrated translation of CO is postulated to occur as a result of the 2π* orbital alignment in chemisorbed CO along the close packed direction of the Cu(110) substrate.
DNA-dependent protein kinase: DNA binding and activation in the absence of Ku
Authors: Hammarsten Ola , Chu Gilbert
Published: July 2002

In mammalian cells, double-strand break repair and V(D)J recombination require DNA-dependent protein kinase (DNA-PK), a serine/threonine kinase that is activated by DNA. DNA-PK consists of a 460-kDa subunit (p460) that contains a putative kinase domain and a heterodimeric subunit (Ku) that binds to double-stranded DNA ends. Previous reports suggested that the activation of DNA-PK requires the binding of Ku to DNA. To investigate this further, p460 and Ku were purified separately to homogeneity. Surprisingly, p460 was capable of binding to DNA in the absence of Ku. The binding of p460 to double-stranded DNA ends was salt-labile and could be disrupted by single-stranded or supercoiled DNA, properties distinct from the binding of Ku to DNA. Under low salt conditions, which permitted the binding of p460 to DNA ends, the kinase was activated. Under higher salt conditions, which inhibited the binding of p460, activation of the kinase required the addition of Ku. Significantly, when the length of DNA decreased to 22 bp, Ku competed with p460 for DNA binding and inhibited kinase activity. These data demonstrate that p460 is a self-contained kinase that is activated by direct interaction with double-stranded DNA and that the role of Ku is to stabilize the binding of p460 to DNA ends.
pVHL
19
is a biologically active product of the von Hippel–Lindau gene arising from internal translation initiation
Authors: Iliopoulos Othon , Ohh Michael , Kaelin William G.
Published: July 2002

The von Hippel–Lindau (VHL) gene encodes a protein consisting of 213 amino acid residues with an apparent molecular mass of 30 kDa (pVHL
30
). Here we show that cells also produce a VHL protein (pVHL
19
) that appears to arise as a result of internal translation from the second methionine within the VHL ORF. pVHL
30
resides primarily in the cytosol, with less amounts found in the nucleus or associated with cell membranes. In contrast pVHL
19
, in biochemical fractionation experiments, is equally distributed between the nucleus and cytosol and is not found in association with membranes. pVHL
19
, like pVHL
30
, can bind to elongin B, elongin C, and Hs-Cul2 in coimmunoprecipitation assays and can inhibit the production of hypoxia-inducible proteins such as vascular endothelial growth factor (VEGF) and GLUT1 when reintroduced into renal carcinoma cells that lack a wild-type VHL allele. Thus, cells contain two biologically active VHL gene products.
A viral suppressor of gene silencing in plants
Authors: Anandalakshmi Radhamani , Pruss Gail J. , Ge Xin , Marathe Rajendra , Mallory Allison C. , Smith Trenton H. , Vance Vicki B.
Published: July 2002

Gene silencing is an important but little understood regulatory mechanism in plants. Here we report that a viral sequence, initially identified as a mediator of synergistic viral disease, acts to suppress the establishment of both transgene-induced and virus-induced posttranscriptional gene silencing. The viral suppressor of silencing comprises the 5′-proximal region of the tobacco etch potyviral genomic RNA encoding P1, helper component-proteinase (HC-Pro) and a small part of P3, and is termed the P1/HC-Pro sequence. A reversal of silencing assay was used to assess the effect of the P1/HC-Pro sequence on transgenic tobacco plants (line T4) that are posttranscriptionally silenced for theuidAreporter gene. Silencing was lifted in offspring of T4 crosses with four independent transgenic lines expressing P1/HC-Pro, but not in offspring of control crosses. Viral vectors were used to assess the effect of P1/HC-Pro expression on virus-induced gene silencing (VIGS). The ability of a potato virus X vector expressing green fluorescent protein to induce silencing of a green fluorescent protein transgene was eliminated or greatly reduced when P1/HC-Pro was expressed from the same vector or from coinfecting potato virus X vectors. Expression of the HC-Pro coding sequence alone was sufficient to suppress virus-induced gene silencing, and the HC-Pro protein product was required for the suppression. This discovery points to the role of gene silencing as a natural antiviral defense system in plants and offers different approaches to elucidate the molecular basis of gene silencing.
Hypsographic demography: The distribution of human population by altitude
Authors: Cohen Joel E. , Small Christopher
Published: July 2002

The global distribution of the human population by elevation is quantified here. As of 1994, an estimated 1.88 × 10
9
people, or 33.5% of the world’s population, lived within 100 vertical meters of sea level, but only 15.6% of all inhabited land lies below 100 m elevation. The median person lived at an elevation of 194 m above sea level. Numbers of people decreased faster than exponentially with increasing elevation. The integrated population density (IPD, the number of people divided by the land area) within 100 vertical meters of sea level was significantly larger than that of any other range of elevations and represented far more people. A significant percentage of the low-elevation population lived at moderate population densities rather than at the highest densities of central large cities. Assessments of coastal hazards that focus only on large cities may substantially underestimate the number of people who could be affected.
Abundant microsatellite polymorphism in
Saccharomyces cerevisiae
, and the different distributions of microsatellites in eight prokaryotes and
S. cerevisiae
, result from strong mutation pressures and a variety of selective forces
Authors: Field Dawn , Wills Christopher
Published: July 2002

We examined the distributions of short tandemly repeated DNAs (microsatellites) in nine complete microbial genomes (
Saccharomyces cerevisiae
,
Archaeoglobus fulgidus
,
Escherichia coli
,
Haemophilus influenzae
,
Helicobacter pylori
,
Methanococcus jannaschii
,
Mycoplasma pneumoniae
,
M. genitalium
, and
Synechocystis
PCC6803.) These repeats contribute differently to the global features of these genomes, and we explore the evolutionary implications of these differences by empirical examination of length polymorphisms at 20 long triplet-repeats repeats in
S. cerevisiae
, and by comparison of observed and expected repeat distributions. All of a sample of 20 microsatellites found in
S. cerevisiae
are highly polymorphic in length, suggesting that mutation pressure overcomes overall selection for small genome size that will tend to shorten or eliminate unnecessary DNA. By comparison, prokaryotes have fewer long repeats than expected, except for a few statistically improbable repeats that appear to function in gene regulation. Finally, we find that in all these genomes there is an excess of repeats shorter than those traditionally considered to be microsatellites. This finding suggests that even in prokaryotes these repeats are being generated by mutational pressures. These results have important potential implications for understanding genome stability and evolution in these microbial species.
The T cell leukemia LIM protein Lmo2 is necessary for adult mouse hematopoiesis
Authors: Yamada Yoshihiro , Warren Alan J. , Dobson Claire , Forster Alan , Pannell Richard , Rabbitts Terence H.
Published: July 2002

The LIM-finger protein Lmo2, which is activated in T cell leukemias by chromosomal translocations, is required for yolk sac erythropoiesis. Because
Lmo2
null mutant mice die at embryonic day 9–10, it prevents an assessment of a role in other stages of hematopoiesis. We have now studied the hematopoietic contribution of homozygous mutant
Lmo2
−/− mouse embryonic stem cells and found that
Lmo2
−/− cells do not contribute to any hematopoietic lineage in adult chimeric mice, but reintroduction of an Lmo2-expression vector rescues the ability of
Lmo2
null embryonic stem cells to contribute to all lineages tested. This disruption of hematopoiesis probably occurs because interaction of Lmo2 protein with factors such as Tal1/Scl is precluded. Thus,
Lmo2
is necessary for early stages of hematopoiesis, and the
Lmo2
master gene encodes a protein that has a central and crucial role in the hematopoietic development.
Attempted hydrogen–deuterium exchange of the
protio
-trimethyloxonium dication (CH
3
)
3
OH
2+
, study of methylating ability of (CH
3
)
3
O
+
in superacids and theoretical investigations
Authors: Olah George A. , Rasul Golam , Burrichter Arwed , Prakash G. K. Surya
Published: July 2002

Attempted hydrogen–deuterium exchange of trimethyloxonium ion, (CH
3
)
3
O
+
with excess of 1:1
2
HF/SbF
5
superacid at −30°C over a period of 30 days showed no exchange. Theoretical calculations at the MP2/6–31G** level are in accord with the lack of hydrogen–deuterium exchange in the methyl group of the (CH
3
)
3
O
+
cation as protonation (protosolvation) prefers the oxygen lone pair of electrons, instead of a C—H bond. Methylation of aromatics with the (CH
3
)
3
O
+
CF
3
SO
3
−
in CF
3
SO
3
H and 2CF
3
SO
3
H:B(O
3
SCF
3
)
3
was also studied. Whereas in triflic acid no alkylation was observed, in triflatoboric acid, a powerful superacid, alkylation takes place, indicating protolytic activation of the trimethyloxonium ion.
Forebrain mechanisms of nociception and pain: Analysis through imaging
Authors: Casey Kenneth L.
Published: July 2002

Pain is a unified experience composed of interacting
discriminative, affective-motivational, and cognitive components, each
of which is mediated and modulated through forebrain mechanisms acting
at spinal, brainstem, and cerebral levels. The size of the human
forebrain in relation to the spinal cord gives anatomical emphasis to
forebrain control over nociceptive processing. Human forebrain
pathology can cause pain without the activation of nociceptors.
Functional imaging of the normal human brain with positron emission
tomography (PET) shows synaptically induced increases in regional
cerebral blood flow (rCBF) in several regions specifically during pain.
We have examined the variables of gender, type of noxious stimulus, and
the origin of nociceptive input as potential determinants of the
pattern and intensity of rCBF responses. The structures most
consistently activated across genders and during contact heat pain,
cold pain, cutaneous laser pain or intramuscular pain were the
contralateral insula and anterior cingulate cortex, the bilateral
thalamus and premotor cortex, and the cerebellar vermis. These regions
are commonly activated in PET studies of pain conducted by other
investigators, and the intensity of the brain rCBF response correlates
parametrically with perceived pain intensity. To complement the human
studies, we developed an animal model for investigating
stimulus-induced rCBF responses in the rat. In accord with behavioral
measures and the results of human PET, there is a progressive and
selective activation of somatosensory and limbic system structures in
the brain and brainstem following the subcutaneous injection of
formalin. The animal model and human PET studies should be mutually
reinforcing and thus facilitate progress in understanding forebrain
mechanisms of normal and pathological pain.
A role for coenzyme M (2-mercaptoethanesulfonic acid) in a bacterial pathway of aliphatic epoxide carboxylation
Authors: Allen Jeffrey R. , Clark Daniel D. , Krum Jonathan G. , Ensign Scott A.
Published: July 2002

The bacterial metabolism of short-chain aliphatic alkenes occurs via oxidation to epoxyalkanes followed by carboxylation to β-ketoacids. Epoxyalkane carboxylation requires four enzymes (components I–IV), NADPH, NAD
+
, and a previously unidentified nucleophilic thiol. In the present work, coenzyme M (2-mercaptoethanesulfonic acid), a compound previously found only in the methanogenic Archaea where it serves as a methyl group carrier and activator, has been identified as the thiol and central cofactor of aliphatic epoxide carboxylation in the Gram-negative bacterium
Xanthobacter
strain Py2. Component I catalyzed the addition of coenzyme M to epoxypropane to form a β-hydroxythioether, 2-(2-hydroxypropylthio)ethanesulfonate. Components III and IV catalyzed the NAD
+
-dependent stereoselective dehydrogenation of
R-
and
S-
enantiomers of 2-(2-hydroxypropylthio)ethanesulfonate to form 2-(2-ketopropylthio)ethanesulfonate. Component II catalyzed the NADPH-dependent cleavage and carboxylation of the β-ketothioether to form acetoacetate and coenzyme M. These findings evince a newfound versatility for coenzyme M as a carrier and activator of alkyl groups longer in chain-length than methane, a function for coenzyme M in a catabolic pathway of hydrocarbon oxidation, and the presence of coenzyme M in the bacterial domain of the phylogenetic tree. These results serve to unify bacterial and Archaeal metabolism further and showcase diverse biological functions for an elegantly simple organic molecule.
Protein synthesis by native chemical ligation: Expanded scope by using straightforward methodology
Authors: Hackeng Tilman M. , Griffin John H. , Dawson Philip E.
Published: July 2002

The total chemical synthesis of proteins has great potential for increasing our understanding of the molecular basis of protein function. The introduction of native chemical ligation techniques to join unprotected peptides next to a cysteine residue has greatly facilitated the synthesis of proteins of moderate size. Here, we describe a straightforward methodology that has enabled us to rapidly analyze the compatibility of the native chemical ligation strategy for X–Cys ligation sites, where X is any of the 20 naturally occurring amino acids. The simplified methodology avoids the necessity of specific amino acid thioester linkers or alkylation of C-terminal thioacid peptides. Experiments using matrix-assisted laser-desorption ionization MS analysis of combinatorial ligations of LYRAX-C-terminal thioester peptides to the peptide CRANK show that all 20 amino acids are suitable for ligation, with Val, Ile, and Pro representing less favorable choices because of slow ligation rates. To illustrate the method’s utility, two 124-aa proteins were manually synthesized by using a three-step, four-piece ligation to yield a fully active human secretory phospholipase A2and a catalytically inactive analog. The combination of flexibility in design with general access because of simplified methodology broadens the applicability and versatility of chemical protein synthesis.
Arterivirus discontinuous mRNA transcription is guided by base pairing between sense and antisense transcription-regulating sequences
Authors: van Marle Guido , Dobbe Jessika C. , Gultyaev Alexander P. , Luytjes Willem , Spaan Willy J. M. , Snijder Eric J.
Published: July 2002

To generate an extensive set of subgenomic (sg) mRNAs, nidoviruses (arteriviruses and coronaviruses) use a mechanism of discontinuous transcription. During this process, mRNAs are generated that represent the genomic 5′ sequence, the so-called leader RNA, fused at specific positions to different 3′ regions of the genome. The fusion of the leader to the mRNA bodies occurs at a short, conserved sequence element, the transcription-regulating sequence (TRS), which precedes every transcription unit in the genome and is also present at the 3′ end of the leader sequence. Here, we have used site-directed mutagenesis of the infectious cDNA clone of the arterivirus equine arteritis virus to show that sg mRNA synthesis requires a base-pairing interaction between the leader TRS and the complement of a body TRS in the viral negative strand. Mutagenesis of the body TRS of equine arteritis virus RNA7 reduced sg RNA7 transcription severely or abolished it completely. Mutations in the leader TRS dramatically influenced the synthesis of all sg mRNAs. The construction of double mutants in which a mutant leader TRS was combined with the corresponding mutant RNA7 body TRS resulted in the specific restoration of mRNA7 synthesis. The analysis of the mRNA leader–body junctions of a number of mutants with partial transcriptional activity provided support for a mechanism of discontinuous minus-strand transcription that resembles similarity-assisted, copy-choice RNA recombination.
Odor space and olfactory processing: Collective algorithms and neural implementation
Authors: Hopfield J. J.
Published: July 2002

Several basic olfactory tasks must be solved by highly olfactory animals, including background suppression, multiple object separation, mixture separation, and source identification. The large number N of classes of olfactory receptor cells—hundreds or thousands—permits the use of computational strategies and algorithms that would not be effective in a stimulus space of low dimension. A model of the patterns of olfactory receptor responses, based on the broad distribution of olfactory thresholds, is constructed. Representing one odor from the viewpoint of another then allows a common description of the most important basic problems and shows how to solve them when N is large. One possible biological implementation of these algorithms uses action potential timing and adaptation as the “hardware” features that are responsible for effective neural computation.