bluuebunny/crossref_metadata_2025 huggingface dataset:
Leukemic potential of doubly mutant Nf1 andWv hematopoietic cells
Authors: Ingram David A. , Wenning Mary Jo , Shannon Kevin , Clapp D. Wade
Published: February 2003

The development of molecularly targeted treatments of adult leukemias warrants investigation of these targets in similar pediatric leukemias. The NF1 tumor suppressor gene, which encodes a GTPase activating protein for p21ras, is frequently inactivated in juvenile myelomonocytic leukemia (JMML). Other patients with JMML acquire activating RAS gene mutations. Recipient mice reconstituted with Nf1−/− fetal hematopoietic cells develop a myeloproliferative disease (MPD) that models the human disease. JMML arises from clonal expansion of a hematopoietic stem cell, and JMML cells and murineNf1−/− hematopoietic cells are hypersensitive to granulocyte macrophage-colony stimulating factor and KitL, the ligand for c-kit. We generated embryos doubly mutant for theWv allele of c-kit and Nf1 to ask if reduction of c-kit activity would delay or prevent the development of MPD. Despite a reduction in c-kit activity to approximately 10% of wild-type levels,Nf1−/−;Wv/Wvcells induced MPD in recipient mice.
Evi3, a common retroviral integration site in murine B-cell lymphoma, encodes an EBFAZ-related Krüppel-like zinc finger protein
Authors: Warming Søren , Liu Pentao , Suzuki Takeshi , Akagi Keiko , Lindtner Susan , Pavlakis George N. , Jenkins Nancy A. , Copeland Neal G.
Published: February 2003

Retroviral insertional mutagenesis in inbred mouse strains provides a powerful method for cancer gene discovery. Here, we show that a common retroviral integration site (RIS) in AKXD B-cell lymphomas, termed Evi3, encodes a novel zinc finger protein with 30 Krüppel-like zinc finger repeats. Most integrations atEvi3 are located upstream of the first translated exon and result in 3′ long-terminal repeat (LTR)–driven overexpression ofEvi3. Evi3 is highly related to the early B-cell factor–associated zinc finger gene (Ebfaz), and all 30 zinc fingers found in EVI3 are conserved in EBFAZ. EBFAZ binds to and negatively regulates early B-cell factor (EBF) (also known as olfactory-1, OLF1), a basic helix-loop-helix (bHLH) transcription factor required for B-lineage commitment and the development of the olfactory epithelium. EBFAZ also binds to SMA- and MAD-related protein–1 (SMAD1) and SMAD4 in response to bone morphogenetic protein–2 (BMP2) signaling, which in turn activates the homeobox regulator of Xenopus mesoderm and neural development Xvent-2. Surprisingly, while Ebfazand Evi3 are coexpressed in many tissues, and both proteins are nuclear, we could not detect Ebfaz expression in B cells by reverse transcriptase–polymerase chain reaction (RT-PCR), whereas Evi3 expression could be detected at all stages of B-cell development. Our results suggest that EVI3, like EBFAZ, is a multifunctional protein that participates in many signaling pathways via its multiple zinc fingers. Furthermore, our results suggest that EVI3, not EBFAZ, is the member of this protein family that interacts with and regulates EBF in B cells.
Complement receptor 1 inhibitors for prevention of immune-mediated red cell destruction: potential use in transfusion therapy
Authors: Yazdanbakhsh Karina , Kang Stanley , Tamasauskas Daniel , Sung Dorothy , Scaradavou Andromachi
Published: February 2003

Activation of complement cascade via the antibody-mediated classical pathway can initiate red blood cell (RBC) destruction, causing transfusion reactions and hemolytic anemia. In the present study, we have assessed the ability of a human recombinant soluble form of complement receptor 1 (sCR1) to inhibit complement-mediated RBC destruction in vitro and in vivo. Using an in vitro alloimmune incompatibility model, sCR1 inhibited complement activation and prevented hemolysis. Following transfusion of human group O RBCs into mice lacking detectable pre-existing antibodies against the transfused RBCs, systemic coadministration of 10 mg/kg sCR1, a dose well tolerated in human subjects for prevention of tissue injury, completely inhibited the in vivo clearance of the transfused RBCs and surface C3 deposition in the first hour after transfusion, correlating with the half-life of sCR1 in the circulation. Treatment with sCR1 increased the survival of transfused human group A RBCs in the circulation of mice with pre-existing anti-A for 2 hours after transfusion by 50%, reduced intravascular hemolysis, and lowered the levels of complement deposition (C3 and C4), but not immunoglobulin G (IgG) or IgM, on the transfused cells by 100-fold. We further identified potential functional domains in CR1 that can act to limit complement-mediated RBC destruction in vitro and in vivo. Collectively, our data highlight a potential use of CR1-based inhibitors for prevention of complement-dependent immune hemolysis.
Stabilities and Isomeric Equilibria in Aqueous Solution of Monomeric Metal Ion Complexes of Adenosine 5′‐Diphosphate (ADP3−) in Comparison with Those of Adenosine 5′‐Monophosphate (AMP2−)
Authors: Bianchi Emanuela M. , Sajadi S. Ali A. , Song Bin , Sigel Helmut
Published: February 2003

Under experimental conditions in which the self‐association of the adenine phosphates (AP), that is, of adenosine 5′‐monophosphate (AMP2−) and adenosine 5′‐diphosphate (ADP3−), is negligible, potentiometric pH titrations were carried out to determine the stabilities of the M(H;AP) and M(AP) complexes where M2+=Mg2+, Ca2+, Sr2+, Ba2+, Mn2+, Co2+, Ni2+, Cu2+, Zn2+, or Cd2+ (25 °C; I=0.1 M, NaNO3). It is concluded that in the M(H;AMP)+ species M2+ is bound at the adenine moiety and in the M(H;ADP) complexes at the diphosphate unit; however, the proton resides in both types of monoprotonated complexes at the phosphate residue. The stabilities of nearly all the M(AMP) and M(ADP)− complexes are significantly larger than what is expected for a sole coordination of M2+ to the phosphate residue. This increased complex stability is attributed, in agreement with previous 1H NMR shift studies and further information existing in the literature, to the formation of macrochelates of the phosphate‐coordinated metal ions with N7 of the adenine residues. On the basis of recent measurements with simple phosphate monoesters and phosphonate ligands (R‐MP2−) as well as with diphosphate monoesters (R‐DP3−), where R is a noncoordinating and noninhibiting residue, the increased stabilities of the M(AMP) and M(ADP)− complexes due to the M2+–N7 interaction could be evaluated and the extent of macrochelate formation calculated. The results show that the formation degrees of the macrochelates for the complexes of the alkaline earth ions are small (about 15 % at the most), whereas for the 3d metal ions as well as for Zn2+ and Cd2+ the formation degrees vary between about 15 % (Mn2+) and 75 % (Ni2+) with values of about 40 and 50 % for Zn2+ and Cu2+, respectively. It is interesting to note, taking earlier results for M(ATP)2− complexes also into account (ATP4−=adenosine 5′‐triphosphate), that for a given metal ion in nearly all instances the formation degrees of the macrochelates are within the error limits the same for M(AMP), M(ADP)− and M(ATP)2− complexes; except for Co2+ and Ni2+ it holds M(AMP) > M(ADP)− ∼ M(ATP)2−. This result is astonishing if one considers that the absolute stability constants of these complexes, which are determined largely by the affinity of the phosphate residues, can differ by more than two orders of magnitude. The impact and conclusions of these observations for biological systems are shortly lined out.
An Unprecedented [2+3] Cycloadditive Dimerization of a Transient Thiocarbonyl S‐Ylide
Authors: El‐Sayed Ibrahim , Grønbæk Hazell Rita , Øgaard Madsen Jørgen , Norrby Per‐Ola , Senning Alexander
Published: February 2003

4‐Chlorophenyl C‐(phenylsulfonyl)dithioformate reacts with diazo(trimethylsilyl)methane to form 2‐(4‐chlorophenylthio)‐2‐phenylsulfonyl‐4,5‐bis(trimethylsilyl)‐1,3‐dithiolan‐1‐ium (4‐chlorophenylthio)(phenylsulfonyl)methylide. It is assumed that this product is formed by [2+3] cycloadditive dimerization of the transient (trimethylsilyl)methanethial S‐[(4‐chlorophenylthio)(phenylsulfonyl)methylide]. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003)
Asymmetric Total Synthesis of the Gastroprotective Microbial Agent AI‐77‐B
Authors: Ghosh Arun K. , Bischoff Alexander , Cappiello John
Published: February 2003

An enantioselective total synthesis of the pseudopeptide microbial agent AI‐77‐B, which has shown potent antiulcerogenic properties, is described. The synthesis is convergent and involves the assembly of a dihydroisocoumarin fragment and a hydroxy amino acid. The dihydroisocoumarin derivative was synthesised by means of a Diels−Alder reaction between 1‐methoxy‐1,3‐cyclohexadiene and an alkynyl ester derivative as the dienophile. The alkynyl ester was obtained stereoselectively by two different synthetic routes: (1) A stereoselective allylation of leucinal, and (2) a titanium enolate‐mediated anti‐aldol reaction with trichlorobutyraldehyde, a novel homopropargylaldehyde equivalent. The stereocentres of the hydroxy amino acid moiety were generated through a titanium enolate‐mediated syn‐aldol reaction, Curtius rearrangement, and application of Dondoni's aldehyde homologation. Condensation of the dihydroisocoumarin and hydroxy amino acid moieties and subsequent removal of the protecting groups furnished optically active AI‐77‐B. (© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2003)
Nitrogen isotopes in mammalian herbivores: hair δ15N values from a controlled feeding study
Authors: Sponheimer M. , Robinson T. , Ayliffe L. , Roeder B. , Hammer J. , Passey B. , West A. , Cerling T. , Dearing D. , Ehleringer J.
Published: February 2003

Nitrogen isotope analysis is a common technique for investigating dietary behaviour in modern and archaeological populations. One of its primary uses is to provide trophic level information. This application is possible because of a ∼3‰ enrichment in15N along each step in the food chain, resulting in carnivores having higher δ15N values than herbivores, which in turn have higher δ15N values than plants. Much variation has also been observed within a trophic level, although the reasons for this are poorly understood. Here we present the results of a controlled feeding study designed to test the effects of gut anatomy and dietary protein levels on hair δ15N values within a trophic level. The data reveal that mammalian herbivores eating identical diets can have hair δ15N values that differ by as much as 3.6‰. This is particularly striking as it suggests that interspecific physiological differences can lead to larger shifts in δ15N values than a shift in trophic level. We also found that diet‐hair fractionation was 2.3‰ greater when herbivores were fed high‐protein (19%) diets than when they were fed low‐protein (9%) diets. The primary nitrogen losses in mammalian herbivores are15N‐depleted urine and15N‐enriched faeces. We reason that an increase in the ratio of urinary to faecal nitrogen efflux leads to greater diet‐hair fractionation on the high‐protein diet. Copyright © 2003 John Wiley & Sons, Ltd.
Theory of ultrasonically induced birefringence
Authors: Lipeles Russell , Kivelson Daniel
Published: February 2003

An extended hydrodynamic theory of birefringence induced by ultrasonic waves in liquids composed of rigid symmetric top molecules is presented. This theory is closely related to that for HH depolarized light scattering spectra and to that of sound absorption. These phenomena involve the interactions of orientations with longitudinal or compressional modes, and the parameters that enter into the theory are the same as those involved in the corresponding theory of the interaction of orientations with shear modes, a theory which is applicable to flow birefringence and VH depolarized light scattering spectra. The theory predicts a linewidth expression for Brillouin scattering and a sound absorption coefficient for a liquid composed of anisometric molecules that differ from expressions previously derived; these expressions depend upon the frequency dependence of the rotational component of the shear viscosity but not upon the frequency dependence of the nonrotational component of the shear viscosity.
The adiabatic theorem in the complex plane and the semiclassical calculation of nonadiabatic transition amplitudes
Authors: Hwang Jenn-Tai , Pechukas Philip
Published: February 2003

This paper is concerned with the problem of calculating amplitudes for nonadiabatic transitions induced by a time-dependent Hamiltonian, in the semiclassical limit h/→0, with emphasis on questions relevant to semiclassical theories of electronically inelastic scattering. For this problem the semiclassical limit is mathematically equivalent to the adiabatic limit, and the adiabatic theorem says that all these transition amplitudes vanish in the limit; the question is, what is the asymptotic form of the nonadiabatic amplitudes, as they go to zero? We consider Hamiltonia that are analytic matrix functions of time. We prove a generalization of the adiabatic theorem to the complex time plane; paradoxically, the adiabatic theorem in the complex plane gives us directly the nonadiabatic amplitudes along the real time axis. We derive Dykhne’s remarkable formula for the two-state case, which says that the limiting form of the transition amplitude depends only on the energy curves of the two states, not on the nonadiabatic coupling which is responsible for transition between them. We discuss the three-state problem at length and show that the obvious generalization of the Dykhne formula is sometimes true, sometimes false. To indicate the scope of methods based on the adiabatic theorem in the complex plane, we give an elementary proof of the semiclassical formula for above-barrier reflection of a one-dimensional particle.
Electron-impact excitation of the low-lying electronic states of HCN
Authors: Chutjian A. , Tanaka H. , Wicke B. G. , Srivastava S. K.
Published: February 2003

We report the first study of the low-energy electron-impact excitation of low-lying electronic transitions in the HCN molecule. Measurements were made at incident electron energies of 11.6 and 21.6 eV in the energy-loss range of 3–10 eV, and at scattering angles of 20°–130°. Inelastic scattering spectra were placed on the absolute cross-section scale by determining first the ratio of inelastic-to-elastic scattering cross sections, and then separately measuring the absolute elastic scattering cross section. Several new electronic transitions are observed which are intrinsically overlapped in the molecule itself. Assignments of these electronic transitions are suggested. These assignments are based on present spectroscopic and cross-sections measurments, high-energy electron scattering spectra, optical absorption spectra, and ab initio molecular orbital calculations.
Methyl NMR relaxation: The effects of spin rotation and chemical shift anisotropy mechanisms
Authors: Matson Gerald B.
Published: February 2003

The formalism developed by the author in a previous publication for the calculation of methyl NMR relaxation by intramolecular dipolar interactions is applied to the spin rotation and chemical shift anisotropy mechanisms. The spin rotation and chemical shielding tensors are put into spherical form to allow the Hamiltonians for the above mechanisms to be developed in terms of spherical tensors. Because the calculations are directed towards elucidating the behavior of a methyl group attached to a macromolecule in solution, only the methyl internal rotation is assumed to contribute through the spin rotation mechanism to the overall relaxation. The importance of properly taking the spin rotation contribution into account, even when the dipolar mechanism is dominant, is demonstrated. A simple picture for the occurrence of correlations between the dipolar and chemical shift anisotropy mechanisms is presented, and the relaxation matrices, including the cross terms, are given for these two mechanisms. The forms of the relaxation matrices indicate that the effect due to the cross terms is small, and this contention is supported by calculations performed for a trifluoromethyl group.
Near infrared reflectance spectra and analysis of H2S frost as a function of temperature
Authors: Ferraro John R. , Fink Uwe
Published: February 2003

The reflection spectra of H2S frost in the near infrared region (1.6–4 μm), as a function of temperature to 62 °K, were measured. Analyses of these spectra were made, and assignments for the most intense combinations attempted. Dramatic changes in spectra are observed from 62–143 °K, connected with a phase transition between a tetragonal, low-temperature phase and a cubic, high-temperature phase, occurring at 125±5 °K. The data present evidence for only two phases being involved for H2S frost.
Laser photoelectron spectrometry of C5H−5: A determination of the electron affinity and Jahn–Teller coupling in cyclopentadienyl
Authors: Engelking P. C. , Lineberger W. C.
Published: February 2003

The photodetachment electron spectrum of cyclopentadienide (C5H−5) yields a value for the cyclopentadienyl electron affinity of 1.786±0.020 eV. Additionally, the spectrum shows structure corresponding to vibrations in the neutral radical. From this an upper bound is placed on the linear Jahn–Teller coupling constant k2<0.5. The quadratic coupling is shown theoretically to be absent for this molecule, a special case of the absence of quadratic coupling in Dn or Cn groups when n is not divisible by three. The magnitude of this Jahn–Teller distortion is sufficient to induce nontotally symmetric Jahn–Teller modes in the spectrum, but is insufficient to suppress the symmetric modes and, in particular, a strong 0–0 transition is observed. Thus, the geometries of the ion and the neutral, apart from the Jahn–Teller distortion, are not radically different.
Master equations, Langevin equations, and the effect of diffusion on concentration fluctuations
Authors: Keizer Joel
Published: February 2003

An anomalous prediction of the master equation formalism is not obtained when concentration fluctuations in chemical reactions are described by the fluctuation-dissipation theory. The known connection between these two theories shows that the discrepancy may be eliminated by taking the properly scaled thermodynamic limit of the master equation prior to achieving steady state. The effect of diffusion on fluctuations is also considered, and exact results are obtained for several reaction mechanisms. At steady states away from equilibrium nonlocal correlations of the Ornstein–Zernike form are found. This agrees with the continuum limit of the master equation formalism used by Gardiner et al.
Photoionization of N2O: Mechanisms of photoionization and ion dissociation
Authors: Berkowitz J. , Eland J. H. D.
Published: February 2003

From high resolution photoionization mass spectrometric data on normal and isotopically substituted N2O it is concluded that the formation of NO+ below the Ã state of N2O+ but above the thermochemical threshold involves the calculated but hitherto unobserved 4A″ state. This state decays to NO+ from low vibrational levels, and mainly to O+ from higher levels. The O+ ion is a more important product of N2O photodissociation than previously thought. The B̃ state of N2O+ is shown to decompose to NO+ and N2+ and the C̃ state to NO+, N2+, N+, and O+. The branching in autoionization from the Ã, B̃, and C̃ Rydberg series to states of different continua has been determined and is found to depend more on the type of series than on the principal quantum number. The shapes of the resonances are strikingly different in the different final channels. The relevance of these findings to ionospheric O++N2 reaction is discussed.
Satellite structure in the x-ray photoelectron spectra of gaseous furan, pyrrole, and thiophen
Authors: Chambers S. A. , Thomas T. D.
Published: February 2003

The intensities and energies of satellite peaks in the x-ray photoelectron spectra of gaseous furan, pyrrole, and thiophen have been measured. Because contributions owing to inelastic scattering of the electrons can be minimized in the gas phase, these investigations give a clearer picture of the satellite structure than do previously reported measurements in the solid phase. CNDO/2 wavefunctions have been used to predict both energies and intensities of the satellite peaks. Although there is qualitative agreement between theory and experiment, quantitative agreement is lacking. The theoretical calculations show that low-energy satellites are those involving transitions from occupied to unoccupied π orbitals. The calculated wavefunctions provide information on charge rearrangement during photoionization. In particular, it is found that not only is there transfer of charge from the heteroatom to the carbons during shake up (expected from properties of the neutral molecule), but also there is charge transfer away from the core hole. Accurate core ionization potentials are reported for these molecules in the gas phase.
Kinetic energy and path curvature in bound state systems
Authors: Switkes Eugene , Russell Eric L. , Skinner James L.
Published: February 2003

The effect of path curvature on kinetic energy in bound state systems has been investigated by solving the Schrödinger equation for a particle confined to an annular region of uniform width. In the one-dimensional limit of a narrow annulus the angular behavior of the particle can be described by linear motion with an effective potential. This potential is related to the radius of curvature R by Veff=−(h/2/2m)(1/4R2). Thus, for a particle confined to a narrow channel high path curvature produces an effective stabilizing influence. An interpretation of this phenomenon is offered and several examples are examined.
Vacuum ultraviolet absorption spectra of binary rare gas mixtures and the properties of heteronuclear rare gas van der Waals molecules
Authors: Freeman Daryl E. , Yoshino K. , Tanaka Y.
Published: February 2003

Vacuum ultraviolet absorption spectra of binary rare gas mixtures Xe/X (X=He, Ne, Ar, or Kr) and Kr/Y (Y=He, Ne, Ar, or Xe) near each of the first two resonance lines of Xe i and Kr i, respectively, have been photographed at high resolution by a 6.65 m spectrograph with a rare gas continuum as background source. The overall formation of spectral features near a resonance line is predominantly towards short wavelengths, and the degree to which discrete bands form near a resonance line generally increases with increasing atomic weight of X or Y. Partial interpretations of the absorption spectra are given in terms of transitions within the heteronuclear rare gas van der Waals molecule XeX or KrY, from the bound ground electronic state (Ω=0) to bound or free excited electronic states (Ω=0,1). Bound excited electronic states are found for KrXe near the first resonance levels of Xe i and Kr i, and for KrXe and ArXe near the second resonance level of Xe i.
Electronic energy levels of benzene below 7 eV
Authors: Doering J. P.
Published: February 2003

The electronic energy levels of benzene below 7 eV have been investigated in an electron impact experiment. Vibrational structure has been resolved for the first three singlet–triplet transitions and the second triplet state has been separated from the overlapping first singlet state. Energies of the vibrational bands of the observed singlet–triplet transitions are (strongest band underlined) S0→T1, 3.67, 3.79, 3.89, 3.98 eV; S0→T2, 4.63, 4.73, 4.85, 4.95 eV; S0→T3, 5.58, 5.69, 5.81 eV. The previous result of Lassettre et al. [E. N. Lassettre, A. Skerbele, M. Dillon, and K. Ross, J. Chem. Phys. 48, 5066 (1968)] which suggested that two electronic states are involved in the 6.2 eV region scattering has been confirmed. These results together with ultraviolet absorption data allow the construction of a rather detailed energy level diagram for benzene up to 7 eV.
Photoionization of dimeric polyatomic molecules: Proton affinities of H2O and HF
Authors: Ng C. Y. , Trevor D. J. , Tiedemann P. W. , Ceyer S. T. , Kronebusch P. L. , Mahan B. H. , Lee Y. T.
Published: February 2003

Photoionization studies of (H2O)2 and (HF)2 producing H3O+ and H2F+ yield 7.18±0.08 eV (165.8±1.8 kcal/mole) and 4.09±0.06 eV (94.3±1.4 kcal/mole) as the proton affinities of H2O and HF, respectively. The measured ionization potential of (H2O)2, 11.21±0.09 eV, along with the known ionization potential of H2O, 12.615±0.001 eV, allow the deduction of a lower bound for the dissociation energy of (H2O)2:1.58±0.13 eV (36±3 kcal/mole). The experiments have demonstrated that photoionization of dimers is one of the most useful general methods for the determination of proton affinities.
The effect of reagent translation on product internal energy distributions: Laser fluorescence study of Al+O2
Authors: Pasternack Louise , Dagdigian Paul J.
Published: February 2003

The effect of incident translational energy on the product internal state distribution has been investigated for the reaction Al+O2→AlO+O. The incident energy is varied through the use of time-of-flight velocity selection of an Al atomic beam, and dye laser fluoresence excitation spectra are reported for AlO produced in a ’’beam-gas’’ arrangement under single-collision conditions. Relative cross sections and effective rotational temperatures are derived for the observed vibrational levels. On the average about 20% of the available energy appears as product vibration and 30% as product rotation. It is found that the AlO internal excitation increases with increasing incident kinetic energy but that ∼80% of the additional kinetic energy is disposed of as product translation and rotation. Comparison of the present results with previous theoretical work suggests that the Al+O2 reaction proceeds on an attractive surface with mixed energy release channeled primarily into product translation and rotation. The observation of a translational energy threshold for the production of AlO in the v=2 level has been employed to derive a lower bound to the AlO dissociation energy D0° (AlO) ?121.0±0.6 kcal/mole. With the help of previous work it is concluded that this lower bound is an equality and a value of 121–122 kcal/mole is recommended for the dissociation energy of aluminum monoxide.
Citron Kinase, a Rho-dependent Kinase, Induces Di-phosphorylation of Regulatory Light Chain of Myosin II
Authors: Yamashiro Shigeko , Totsukawa Go , Yamakita Yoshihiko , Sasaki Yasuharu , Madaule Pascal , Ishizaki Toshimaa , Narumiya Shuh , Matsumura Fumio
Published: February 2003

Citron kinase is a Rho-effector protein kinase that is related to Rho-associated kinases of ROCK/ROK/Rho-kinase family. Both ROCK and citron kinase are suggested to play a role in cytokinesis. However, no substrates are known for citron kinase. We found that citron kinase phosphorylated regulatory light chain (MLC) of myosin II at both Ser-19 and Thr-18 in vitro. Unlike ROCK, however, citron kinase did not phosphorylate the myosin binding subunit of myosin phosphatase, indicating that it does not inhibit myosin phosphatase. We found that the expression of the kinase domain of citron kinase resulted in an increase in MLC di-phosphorylation. Furthermore, the kinase domain was able to increase di-phosphorylation and restore stress fiber assembly even when ROCK was inhibited with a specific inhibitor, Y-27632. The expression of full-length citron kinase also increased di-phosphorylation during cytokinesis. These observations suggest that citron kinase phosphorylates MLC to generate di-phosphorylated MLC in vivo. Although both mono- and di-phosphorylated MLC were found in cleavage furrows, di-phosphorylated MLC showed more constrained localization than did mono-phosphorylated MLC. Because citron kinase is localized in cleavage furrows, citron kinase may be involved in regulating di-phosphorylation of MLC during cytokinesis.
Centrosomes Split in the Presence of Impaired DNA Integrity during Mitosis
Authors: Hut Henderika M.J. , Lemstra Willy , Blaauw Engbert H. , van Cappellen Gert W.A. , Kampinga Harm H. , Sibon Ody C.M.
Published: February 2003

A well-established function of centrosomes is their role in accomplishing a successful mitosis that gives rise to a pair of identical daughter cells. We recently showed that DNA replication defects and DNA damage in Drosophila embryos trigger centrosomal changes, but it remained unclear whether comparable centrosomal responses can be provoked in somatic mammalian cells. To investigate the centrosomal organization in the presence of impaired DNA integrity, live and ultrastructural analysis was performed on γ-tubulin–GFP and EGFP–α-tubulin–expressing Chinese hamster ovary cells. We have shown that during mitosis in the presence of incompletely replicated or damaged DNA, centrosomes split into fractions containing only one centriole. This results in the formation of multipolar spindles with extra centrosome-like structures. Despite the extra centrosomes and the multipolarity of the spindles, cells do exit from mitosis, resulting in severe division errors. Our data provide evidence of a novel mechanism showing how numerous centrosomes and spindle defects can arise and how this can lead to the formation of aneuploid cells.
The Database of Quantitative Cellular Signaling:
management and analysis of chemical kinetic models of signaling
networks
Authors: Sivakumaran Sudhir , Hariharaputran Sridhar , Mishra Jyoti , Bhalla Upinder S.
Published: February 2003

Motivation: Analysis of cellular signaling interactions is expected to pose an enormous informatics challenge, perhaps even larger than analyzing the genome. The complex networks arising from signaling processes are traditionally represented as block diagrams. A key step in the evolution toward a more quantitative understanding of signaling is to explicitly specify the kinetics of all chemical reaction steps in a pathway. Technical advances in proteomics and high-throughput protein interaction assays promise a flood of such quantitative data. While annotations, molecular information and pathway connectivity have been compiled in several databases, and there are several proposals for general cell model description languages, there is currently little experience with databases of chemical kinetics and reaction level models of signaling networks.

Results: The Database of Quantitative Cellular Signaling is a repository of models of signaling pathways. It is intended both to serve the growing field of chemical-reaction level simulation of signaling networks, and to anticipate issues in large-scale data management for signaling chemistry.

Availability: The Database of Quantitative Cellular Signaling is available at http://doqcs.ncbs.res.in. Links to the signaling model simulator, GENESIS/Kinetikit are at http://www.ncbs.res.in/~bhalla/kkit/index.html

Contact: bhalla@ncbs.res.in

* To whom correspondence should be addressed.
Advanced alloy for lead‐free solder balls
Authors: Dittes Marc , Walter Hermann
Published: February 2003

A new alloy for lead‐free solder balls is introduced combining the advantages of a smooth surface after production and a uniform reflectivity after the ball attach process. This property significantly simplifies the inspection process during ball grid array (BGA) processing. By adding small amounts of Indium and Lanthanum to the ternary eutectic SnAgCu‐alloy a pentary, virtually eutectic composition with a melting point of 214°C is realised. It is shown that the processability in terms of reflow behaviour and the shear strength of BGA spheres made of this alloy is comparable to or better than that of ternary SnAgCu‐balls.
Quantitative Role of the Human Papillomavirus Type 16 E5 Gene during the Productive Stage of the Viral Life Cycle
Authors: Genther Sybil M. , Sterling Stephanie , Duensing Stefan , Münger Karl , Sattler Carol , Lambert Paul F.
Published: February 2003

Human papillomaviruses (HPVs) are small circular DNA viruses that cause warts. Infection with high-risk anogenital HPVs, such as HPV type 16 (HPV16), is associated with human cancers, specifically cervical cancer. The life cycle of HPVs is intimately tied to the differentiation status of the host epithelium and has two distinct stages: the nonproductive stage and the productive stage. In the nonproductive stage, which arises in the poorly differentiated basal epithelial compartment of a wart, the virus maintains itself as a low-copy-number nuclear plasmid. In the productive stage, which arises as the host cell undergoes terminal differentiation, viral DNA is amplified; the capsid genes, L1 and L2, are expressed; and progeny virions are produced. This stage of the viral life cycle relies on the ability of the virus to reprogram the differentiated cells to support DNA synthesis. Papillomaviruses encode multiple oncoproteins, E5, E6, and E7. In the present study, we analyze the role of one of these viral oncogenes, E5, in the viral life cycle. To assess the role of E5 in the HPV16 life cycle, we introduced wild-type (WT) or E5 mutant HPV16 genomes into NIKS, a keratinocyte cell line that supports the papillomavirus life cycle. By culturing these cells under conditions that allow them to remain undifferentiated, a state similar to that of basal epithelial cells, we determined that E5 does not play an essential role in the nonproductive stage of the HPV16 life cycle. To determine if E5 plays a role in the productive stage of the viral life cycle, we cultured keratinocyte populations in organotypic raft cultures, which promote the differentiation and stratification of epithelial cells. We found that cells harboring E5 mutant genomes displayed a quantitative reduction in the percentage of suprabasal cells undergoing DNA synthesis, compared to cells containing WT HPV16 DNA. This reduction in DNA synthesis, however, did not prevent amplification of viral DNA in the differentiated cellular compartment. Likewise, late viral gene expression and the perturbation of normal keratinocyte differentiation were retained in cells harboring E5 mutant genomes. These data demonstrate that E5 plays a subtle role during the productive stage of the HPV16 life cycle.
Dimerization and Template Switching in the 5′ Untranslated Region between Various Subtypes of Human Immunodeficiency Virus Type 1
Authors: Andersen Ebbe Sloth , Jeeninga Rienk E. , Damgaard Christian Kroun , Berkhout Ben , Kjems Jørgen
Published: February 2003

The human immunodeficiency virus type 1 (HIV-1) particle contains two identical RNA strands, each corresponding to the entire genome. The 5′ untranslated region (UTR) of each RNA strand contains extensive secondary and tertiary structures that are instrumental in different steps of the viral replication cycle. We have characterized the 5′ UTRs of nine different HIV-1 isolates representing subtypes A through G and, by comparing their homodimerization and heterodimerization potentials, found that complementarity between the palindromic sequences in the dimerization initiation site (DIS) hairpins is necessary and sufficient for in vitro dimerization of two subtype RNAs. The 5′ UTR sequences were used to design donor and acceptor templates for a coupled in vitro dimerization-reverse transcription assay. We showed that template switching during reverse transcription is increased with a matching DIS palindrome and further stimulated proportional to the level of homology between the templates. The presence of the HIV-1 nucleocapsid protein NCp7 increased the template-switching efficiency for matching DIS palindromes twofold, whereas the recombination efficiency was increased sevenfold with a nonmatching palindrome. Since NCp7 did not effect the dimerization of nonmatching palindromes, we concluded that the protein most likely stimulates the strand transfer reaction. An analysis of the distribution of template-switching events revealed that it occurs throughout the 5′ UTR. Together, these results demonstrate that the template switching of HIV-1 reverse transcriptase occurs frequently in vitro and that this process is facilitated mainly by template proximity and the level of homology.
Evaluation of Genetically Engineered Derivatives of a Chinese Strain of Foot-and-Mouth Disease Virus Reveals a Novel Cell-Binding Site Which Functions in Cell Culture and in Animals
Authors: Zhao Qizu , Pacheco Juan M. , Mason Peter W.
Published: February 2003

Adaptation of field isolates of foot-and-mouth disease virus (FMDV) to grow in cells in culture can result in changes in viral properties that include acquisition of the ability to bind to cell surface heparan sulfate (HS). After 13 passages on BHK cells to produce a vaccine, a Cathay topotype isolate of FMDV serotype O from China (O/CHA/90) extended its cell culture host range and bound to heparin-Sepharose, although it did not require cell surface HS as a receptor molecule. To understand these phenomena, we constructed chimeric viruses by using a type A
12
infectious cDNA and the capsid protein-coding regions of O/CHA/90 and its cell culture-adapted derivative (vac-O/CHA/90). Using a set of viruses derived from these chimeras by exchanging portions of the capsid-coding regions, we discovered that a group of amino acid residues that surround the fivefold axis of the icosahedral virion determine host range in cell culture and influence pathogenicity in pigs. These residues included aromatic amino acids at positions 108 and 174 and positively charged residues at positions 83 and 172 in protein 1D. To test if these residues participated in non-integrin-dependent cell binding, the integrin-binding RGD sequence in protein 1D was changed to KGE in two different chimeras. Evaluation of these KGE viruses indicated that growth in cell culture was not dependent on HS. One of these viruses was tested in pigs, where it produced a mild disease and maintained its KGE sequence. These results are discussed in terms of receptor utilization and pathogenesis of this important pathogen.
Structures of Rotavirus Reassortants Demonstrate Correlation of Altered Conformation of the VP4 Spike and Expression of Unexpected VP4-Associated Phenotypes
Authors: Pesavento Joseph B. , Billingsley Angela M. , Roberts Ed J. , Ramig Robert F. , Prasad B. V. Venkataram
Published: February 2003

Numerous prior studies have indicated that viable rotavirus reassortants containing structural proteins of heterologous parental origin may express unexpected phenotypes, such as changes in infectivity and immunogenicity. To provide a structural basis for alterations in phenotypic expression, a three-dimensional structural analysis of these reassortants was conducted. The structures of the reassortants show that while VP4 generally maintains the parental structure when moved to a heterologous protein background, in certain reassortants, there are subtle alterations in the conformation of VP4. The alterations in VP4 conformation correlated with expression of unexpected VP4-associated phenotypes. Interactions between heterologous VP4 and VP7 in reassortants expressing unexpected phenotypes appeared to induce the conformational alterations seen in VP4.
Ebola Virus Transcription Activator VP30 Is a Zinc-Binding Protein
Authors: Modrof Jens , Becker Stephan , Mühlberger Elke
Published: February 2003

Ebola virus VP30 is an essential activator of viral transcription. In viral particles, VP30 is closely associated with the nucleocapsid complex. A conspicuous structural feature of VP30 is an unconventional zinc-binding Cys
3
-His motif comprising amino acids 68 to 95. By using a colorimetric zinc-binding assay we found that the VP30-specific Cys
3
-His motif stoichiometrically binds zinc ions in a one-to-one relationship. Substitution of the conserved cysteines and the histidine within the motif led to a complete loss of the capacity for zinc binding. Functional analyses revealed that none of the tested mutations of the proposed zinc-coordinating residues influenced binding of VP30 to nucleocapsid-like particles but, concerning its role in activating viral transcription, all resulted in a protein that was inactive.
Computer Simulations of Enzyme Catalysis: Methods, Progress, and Insights
Authors: Warshel Arieh
Published: February 2003

▪ Abstract  Understanding the action of enzymes on an atomistic level is one of the important aims of modern biophysics. This review describes the state of the art in addressing this challenge by simulating enzymatic reactions. It considers different modeling methods including the empirical valence bond (EVB) and more standard molecular orbital quantum mechanics/molecular mechanics (QM/MM) methods. The importance of proper configurational averaging of QM/MM energies is emphasized, pointing out that at present such averages are performed most effectively by the EVB method. It is clarified that all properly conducted simulation studies have identified electrostatic preorganization effects as the source of enzyme catalysis. It is argued that the ability to simulate enzymatic reactions also provides the chance to examine the importance of nonelectrostatic contributions and the validity of the corresponding proposals. In fact, simulation studies have indicated that prominent proposals such as desolvation, steric strain, near attack conformation, entropy traps, and coherent dynamics do not account for a major part of the catalytic power of enzymes. Finally, it is pointed out that although some of the issues are likely to remain controversial for some time, computer modeling approaches can provide a powerful tool for understanding enzyme catalysis.
Neuropilin-1 on hematopoietic cells as a source of vascular development
Authors: Yamada Yoshihiro , Oike Yuichi , Ogawa Hisao , Ito Yasuhiro , Fujisawa Hajime , Suda Toshio , Takakura Nobuyuki
Published: February 2003

Neuropilin-1 (NP-1) is a receptor for vascular endothelial growth factor-165 (VEGF165) and acts as a coreceptor that enhances the function of VEGF165 through VEGF receptor-2 (VEGFR-2). Studies using transgenic and knock-out mice of NP-1 indicated that this molecule is important for vascular development as well as neuronal development. We recently reported that clustered soluble NP-1 phosphorylates VEGFR-2 on endothelial cells with a low dose of VEGF165 and rescues the defective vascularity of the NP-1−/− embryo in vitro and in vivo. Here we show that NP-1 is expressed by CD45+ hematopoietic cells in the fetal liver, can bind VEGF165, and phosphorylates VEGFR-2 on endothelial cells. CD45+NP-1+ cells rescued the defective vasculogenesis and angiogenesis in the NP-1−/− P-Sp (para-aortic splanchnopleural mesodermal region) culture, although CD45+NP-1− cells did not. Moreover, CD45+NP-1+ cells together with VEGF165 induced angiogenesis in an in vivo Matrigel assay and cornea neovascularization assay. The extracellular domain of NP-1 consists of “a,” “b,” and “c” domains, and it is known that the “a” and “c” domains are necessary for dimerization of NP-1. We found that both the “a” and “c” domains are essential for such rescue of defective vascularities in the NP-1 mutant. These results suggest that NP-1 enhances vasculogenesis and angiogenesis exogenously and that dimerization of NP-1 is important for enhancing vascular development. In NP-1−/− embryos, vascular sprouting is impaired at the central nervous system (CNS) and pericardium where VEGF is not abundant, indicating that NP-1–expressing cells are required for normal vascular development.
CD100/Plexin-B1 interactions sustain proliferation and survival of normal and leukemic CD5+ B lymphocytes
Authors: Granziero Luisa , Circosta Paola , Scielzo Cristina , Frisaldi Elisa , Stella Stefania , Geuna Massimo , Giordano Silvia , Ghia Paolo , Caligaris-Cappio Federico
Published: February 2003

Growth and survival of chronic B-cell tumors are favored by the malignant cell's capacity to respond to selected microenvironmental stimuli provided by nontumoral bystander cells. To investigate which mechanisms operate in these crosstalks and whether they are malignancy-related or reproduce the mechanisms used by normal B cells we have studied the expression and functional role of semaphorin CD100 (now called Sema4D) in chronic lymphocytic leukemia (CLL) cells and normal CD5+ B cells. We demonstrate here that (1) leukemic and normal CD5+ B lymphocytes uniformly express CD100; (2) the CD100 high-affinity receptor Plexin-B1 is expressed by bone marrow stromal cells, follicular dendritic cells, and activated T lymphocytes, and is thus available to CD100+ lymphocytes in different specific microenvironments; and (3) upon interaction between CD100 and Plexin-B1 both CLL and normal CD5+ B cells increase their proliferative activity and extend their life span. These findings establish that Plexin-B1 is an easily accessible receptor for CD100 within the immune system. The encounter of CD100+ leukemic cells with Plexin-B1 may promote the proliferation and survival of malignant cells. The crosstalk operated by the CD100/Plexin-B1 interaction is not malignancy related but reproduces a mechanism used by normal CD5+ B cells.
NPM-ALK transgenic mice spontaneously develop T-cell lymphomas and plasma cell tumors
Authors: Chiarle Roberto , Gong Jerald Z. , Guasparri Ilaria , Pesci Anna , Cai Jonjing , Liu Jian , Simmons William J. , Dhall Girish , Howes Jennifer , Piva Roberto , Inghirami Giorgio
Published: February 2003

Anaplastic Large CellLymphomas (ALCLs) carry translocations in which the anaplastic lymphoma kinase (ALK) gene is juxtaposed to various genes, the most common of which is the NPM/B23gene. ALK fusion proteins result in the constitutive activation of ALK tyrosine kinase, thereby enhancing proliferation and increasing cell survival. A direct role for NPM-ALK in cellular transformation has been shown in vitro with immortalized cell lines and in vivo using retroviral transfer experiments. Nonetheless, there is no direct evidence of its oncogenic potential in T lymphocytes, which represent the most common target of ALK chimeras. Here, we describe a new mouse model of lymphomagenesis in which human NPM-ALK transcription was targeted to T cells. NPM-ALK transgenic (Tg) mice were born with the expected mendelian distribution, normal lymphoid organs, and a normal number and proportion of helper and suppressor T cells. However, after a short period of latency, all NPM-ALK Tg mice developed malignant lymphoproliferative disorders (mean survival, 18 weeks). NPM-ALK Tg thymic lymphomas displayed a T-cell phenotype characteristic of immature thymocytes and frequently coexpressed surface CD30. A subset of the NPM-ALK Tg mice also developed clonal B-cell plasma cell neoplasms. These tumors arose in peripheral lymphoid organs (plasmacytomas) or within the bone marrow and often led to peripheral neuropathies and limb paralysis. Our NPM-ALK Tg mice are a suitable model to dissect the molecular mechanisms of ALK-mediated transformation and to investigate the efficacy of new therapeutic approaches for the treatment of human ALCL in vivo.