Probing electron-phonon excitations in molecular junctions by quantum interference.

Electron-phonon coupling is a fundamental inelastic interaction in condensed matter and in molecules. Here we probe phonon excitations using quantum interference in electron transport occurring in short chains of anthraquinone based molecular junctions. By studying the dependence of molecular junction's conductance as a function of bias voltage and temperature, we show that inelastic scattering of electrons by phonons can be detected as features in conductance resulting from quenching of quantum interference. Our results are in agreement with density functional theory calculations and are well described by a generic two-site model in the framework of non-equilibrium Green's functions formalism. The importance of the observed inelastic contribution to the current opens up new ways for exploring coherent electron transport through molecular devices.

[Cytogenetic maps of lampbrush chromosomes of newts of the genus Pleurodeles: an algorithm of lampbrush chromosome identification in Pleurodeles waltl by immunocytochemical marker-loop staining with polyclonal anti-Ro52 antisera]. / Tsitogeneticheskie karty khromosom-lampovykh shchetok tritonov roda Pleurodeles: algoritm identifikatsii lampovykh shchetok Pleurodeles waltl s ispol'zovaniem immunokhimicheskogo okrashivaniia markernykh petel' poliklonal'noi syvorotki protiv proteina Ro52.

Our work was aimed at developing a simple and effective method of identification of most or all chromosomes of Pleurodeles newts. To this end, we used DAPI staining of the chromomeres of newt lampbrush chromosomes and immunochemical reactions between the ribonucleoprotein (RNP) marker loops and polyclonal antibodies against human zinc-finger protein Ro52 (52-kDa Ro/SS-A). A method has been developed to obtain newt lampbrush chromosome preparations. Cytological maps of P. waltl chromosomes (Spanish population/subspecies) showing distributions of chromomeres and marker loops along the chromosome length were constructed.

Oligomer-oligomer versus oligomer-monomer C(2)-C(2)' coupling reactions in polypyrrole growth.

The C(2)-C(2)' coupling reactions of oligopyrrole radical-cations of increasing length generated by electrochemical oxidation have been modeled by transition state calculations. The modeling approach takes into account solvent effects and (i) shows that the coupling distance in the transition state decreases with oligomer length, (ii) demonstrates that dimerization rates in the gas phase decrease with oligomer length but increase in water, (iii) suggests that in a less solvating medium the dimerization rates could be equivalent, (iv) indicates that in all solvents quaterpyrrole and sexipyrrole formation is faster through a coupling reaction between oligomer and monomer radical-cations than two oligomer radical-cations, and (v) suggests that for the formation of a long oligopyrrole from oligopyrrole-pyrrole reactions the mechanism might involve the coupling of the oligopyrrole dication with a non-oxidized pyrrole unit instead of the coupling of two radical-cations or that of the oligopyrrole dication with a pyrrole radical-cation.

Conducting-polymer electrochemical switching as an easy means for control of the molecular properties of grafted transition metal complexes.

Copper(II) 3',4'-bis(N,N'-oxamato)terthiophene has been synthesized and electropolymerized. The copper(II)-complex centers are not affected by the polymerization process, which involves coupling between Calpha carbon atoms of the terthiophene units and leads to a new conjugated polymer consisting of polythiophene chains bearing bis(oxamato)-Cu(II) groups regioregularly grafted onto the polymer backbone. The polymer is stable with respect to polythiophene electroactivity, and no demetallation or modification of the Cu oxidation state occurs over a large potential range. In this material, the two moieties exhibit direct electronic interaction, which makes it possible to use the conductive polymer backbone as a molecular wire or a nanocontact capable of inputting to the bis(oxamato)-Cu(II) groups through the polythiophene-switching reaction. FTIR, XPS, and XAS spectroscopies have been used to study the effect of the state of the conducting polymer upon the properties of the copper(II) center (electron density, ligand field strength, size of cavity, force constants of some bonds). These properties can be controlled to some extent by the potential applied to this device. From the point of view of the copper(II) center, this effect is similar to the grafting of substituents with various electronic properties.

240Pu/239Pu isotopic ratios and 239 + 240Pu total measurements in surface and deep waters around Mururoa and Fangataufa atolls compared with Rangiroa atoll (French Polynesia).

The average values of 240Pu/239Pu mass isotopic ratios of plutonium deposited in Mururoa and Fangataufa atoll sediments by French atmospheric nuclear tests range from 3.5 to 5%. In order to assess the near field and far field influence of those deposits in the open ocean, two water profiles were measured for 239 + 240Pu and 240Pu/239Pu using, for the first time, an Inductively Coupled Plasma Mass Spectrometer which was developed to achieve femtogram detection limits. One site was located at the limit of the French territorial waters, which is 22 km distant from Mururoa. The second site was located close to Rangiroa atoll, at a distance of approximately 1200-km from French nuclear test sites. The sample volumes were approximately 500 litres and plutonium was purified prior to mass spectrometry and alpha spectrometry measurements. In Rangiroa, the 239 + 240Pu profile is comparable with those already determined in world open oceans but the maximum detected activity, 9 mBq/m3 at 500-600 m is a lot lower than those measured in the northern hemisphere. 240Pu/239Pu ratios were measured between 500 and 1000 m and were not statistically different from the typical 0.18 +/- 0.01 ratio which characterises the global fallout. Consequently, any influence of plutonium from the tests in Mururoa and Fangataufa is not apparent at Rangiroa. The vertical distribution of 239 + 240Pu near Mururoa shows similar changes with depth but with a slight increase in concentration. 240Pu/239Pu mass ratios vary with depth, from 7 to 10% in the upper 500 m and in the deep waters (below 1000 m) to 15-16% between 600 and 1000 m. A contribution from plutonium deposited in the sediments at Mururoa and Fangataufa is observed at the limit of territorial waters, especially in surface and deep waters.

XL43 and XL75: two novel RING finger-containing genes expressed during oogenesis and embryogenesis in Xenopus laevis.

This article reports on the isolation of two new Xenopus genes that encode two putative zinc finger proteins. The XL43 and XL75 proteins belong to the RBCC family, and also contains the rfp-like domain. XL43 and XL75 RNAs are found in the ovary, and myc-tagged proteins are detected in mRNA injected oocytes. Whole-mount in situ hybridization of embryos revealed that these two clones are expressed exclusively in neurectodermic and mesodermic tissues. The data suggest that XL43 and XL75 genes could play an important role in the frog's early development, perhaps as a transcription factor. This RBCC family, a subclass of the RING finger family, comprises proteins with known cellular transformation properties. All members possess, besides one RING finger motif, one or two B-boxes, each having a pair of zinc fingers, and a coiled coil domain (Borden, K.L. B. et al., 1996. Proc. Natl. Acad. Sci. USA 93, 1601-1606). Among this group, some members possess, besides the RING-B box-coiled coil (RBCC) motifs, a long C-terminal domain referred to as the rfp-like domain. Although its effective role has not been elucidated yet, this last domain could play an important role by binding a ligand (Bellini, M. et al., 1993. EMBO J. 12, 107-114).

B24 protein stored in lampbrush spheres is involved in early cleavage in urodele amphibians.

A cDNA encoding a protein (B24) belonging to the Mcm/P1 family was isolated from the newt Triturus carnifex. In eukaryotes, the members of the Mcm/P1 family are essential factors in the DNA replication process. B24 protein (TcMcm3) is present in salamandrid ovarian oocytes and early embryos; its role was tested by injecting specific anti-B24 monoclonal antibodies into the cytoplasm of one blastomere of two-cell stage embryos. The injected blastomere encountered cleavage arrest either soon after the injection or following one or two divisions; later, it degenerated. Instead, the uninjected blastomere went on developing and organizing a hemi-embryo, which does not grow beyond the tailbud stage. These results are consistent with the hypothesis that the B24 protein is involved in DNA replication at cleavage. The B24 protein studied here appears to play a specific role in early development; other variants of the Mcm3 group seem to be employed by different adult tissues.

The I protein of the heterogeneous nuclear ribonucleoprotein complex is a novel dog nuclear autoantigen.

In eukaryotic cells, heterogeneous nuclear RNA is associated with a set of abundant nuclear proteins to form complex ribonucleoprotein structures (hnRNP). Autoantibodies to hnRNP G protein have been previously reported in German shepherd dogs with lupus-like syndrome. In the present study, we describe the characterization of a novel antigen recognized by a serum from a schnauzer dog with a non-erosive polyarthritis. The autoantibodies give, by indirect immunofluorescence, a nuclear pattern with staining close to one of the nucleoli. Immunoblotting and immunoprecipitation data reveal that the autoantigens are in fact two closely related basic proteins (average pI 8.7) with apparent molecular weights of 56 kDa (p56) and 59 kDa (p59). The results of immunoprecipitation with anti-hnRNP antibodies and DNA affinity column chromatography strongly suggest that these autoantigens correspond to hnRNP I proteins. This point was confirmed by cloning and sequencing a cDNA clone encoding the complete sequence of the antigens. In addition, we found that anti-hnRNP I antibodies preferentially stain certain loops of the Pleurodeles waltl lampbruch chromosomes. These data, added to previous ones on anti-p43/hnRNP G protein in German shepherd dogs with lupus-like syndrome, confirm the interest of this category of antibodies to hnRNP proteins in autoimmune disorders.

A zinc-binding domain is required for targeting the maternal nuclear protein PwA33 to lampbrush chromosome loops.

In oocytes of the newt Pleurodeles waltl, the maternal nuclear protein PwA33 occurs on the lampbrush chromosomes and in some nucleoplasmic particles of the germinal vesicle. PwA33 is a modular protein and we used site-directed mutagenesis to alter the sequences encoding two metal-binding regions, the C3HC4 (or RING finger) and B-box motifs. Several mutant clones were generated and their synthetic transcripts were injected into Pleurodeles oocytes for in vivo analysis. In the oocyte, all translation products localized in the germinal vesicle. Proteins encoded by RING finger mutant clones were distributed in a pattern identical to that of the wild type protein, but when His266 of the B-box was mutated, PwA33 failed to localize in the lampbrush chromosomes and the nucleoplasmic particles. Using an in vitro colorimetric assay, we demonstrated that PwA33 is a zinc-binding protein and that mutations in the RING finger and B-Box altered its metal-binding properties. The RING finger motif bound two Zn2+ ions and the binding ratios of several mutants were consistent with the tertiary structure recently proposed for this motif. The B-box coordinated one Zn2+ and this binding was inhibited by the His266 mutation. The failure of the His266 mutation to bind zinc and to localize properly within the germinal vesicle suggests that an intact B-box is required for normal functioning of the PwA33 protein in the oocyte.

Localization of antigens PwA33 and La on lampbrush chromosomes and on nucleoplasmic structures in the oocyte of the urodele Pleurodeles waltl: light and electron microscopic immunocytochemical studies.

Monoclonal antibodies A33/22 and La11G7 have been used to study the distribution of the corresponding antigens, PwA33 and La, on the lampbrush chromosome loops and nucleoplasmic structures of P. waltl oocytes, using immunofluorescence, confocal laser scanning microscopy and immunogold labeling. The results obtained with these antibodies have been compared with those obtained with the Sm-antigen-specific monoclonal antibody Y12. All these monoclonal antibodies (mAbs) labeled the matrices of the majority of normal loops along their whole length. Nucleoplasmic RNP granules showed a strong staining with the mAbs La11G7 and Y12 throughout their mass, but with the mAb A33/22, they showed only a weak peripheral labeling in the form of patches on their surface. This patchy labeling was confirmed by confocal laser scanning microscopy. Electron microscopy revealed that this patchy labeling might be due to a hitherto undescribed type of submicroscopic granular structure, around 100 nm in either dimension, formed by 10-nm particles. Such granules were observed either attached to the RNP granules or free in the nucleoplasm, but rarely in relation with the normal loop matrices. These 100-nm granules may have a role in the movement of proteins and snRNPs inside the oocyte nuclei for storage, recycling, and/or degradation. Our results also suggest that all the microscopically visible free RNP granules of the nucleoplasm of P. waltl oocytes correspond to B snurposomes. The granules forming the B (globular) loops showed a labeling pattern similar to that of B snurposomes; their possible relationship is discussed.

Asymmetrical DNA and AT/GC base content of differential sector of Pleurodeles waltl sexual bivalent: a quantitative fluorescence imaging analysis in lampbrush chromosomes.

The mitotic Z and W sex chromosomes in Pleurodeles seem to be identical. Earlier morphological and molecular analyses of lampbrush paired chromosomes in the female meiosis showed clearly that 20% of the chromosomal length located in the middle part of the sex bivalent (bivalent IV) is heteromorphic. We investigated here the base content and composition of the DNA axes in the heteromorphic region by quantitative fluorescence imaging using various base-specific (DAPI, Hoechst 33342 and chromo-mycin A3) or base-nonspecific (ethidium bromide) fluorescent DNA probes. Our results show a significantly higher percentage of AT bases in Z than in W differential sectors. In addition the entire base content of Z appears slightly higher than that of W.

Identification of an amphibian oocyte nuclear protein as a candidate for a role in embryonic DNA replication.

Monoclonal antibody B24/3 recognizes a nuclear protein of 104 kD in germinal vesicles of newt oocytes. Immunohistostaining of oocytes at different stages of growth shows an accumulation of B24 protein throughout oogenesis. During development B24 protein is located inside embryonic cell nuclei from the onset of cleavage onwards. It gradually decreases from gastrulation and disappears at the tailbud stage. The NvB24 17.1 clone was isolated from an ovary expression library of the newt Notophthalmus viridescens and then sequenced: the open reading frame is capable of encoding a polypeptide of 744 amino acids. Northern blot experiments have shown that the 17.1 clone recognizes a single transcript of about 3 Kb in the ovary. In situ hybridization experiments showed that B24 mRNA transcription starts from previtellogenic oocytes, and is followed by the appearance and gradual accumulation of B24 protein in germinal vesicles of medium and large size oocytes. Keeping in mind the sequence similarity shown by the B24 protein to the mouse P1 protein as well as to the budding yeast Mcm3 and fission yeast cdc21 proteins, B24 protein can be speculated to play a role in the events of DNA replication during early amphibian embryogenesis. As B24 antigen is located in the sphere organelles both inserted on the lampbrush chromosomes and free in the oocyte nucleoplasm, an additional possible role of B24 protein could be related to assembling and/or storing snRNPs during oogenesis.

hnRNP G: sequence and characterization of a glycosylated RNA-binding protein.

The autoantigen p43 is a nuclear protein initially identified with autoantibodies from dogs with a lupus-like syndrome. Here we show that p43 is an RNA-binding protein, and identify it as hnRNP G, a previously described component of heterogeneous nuclear ribonucleoprotein complexes. We demonstrate that p43/hnRNP G is glycosylated, and identify the modification as O-linked N-acetylglucosamine. A full-length cDNA clone for hnRNP G has been isolated and sequenced, and the predicted amino acid sequence for hnRNP G shows that it contains one RNP-consensus RNA binding domain (RBD) at the amino terminus and a carboxyl domain rich in serines, arginines and glycines. The RBD of human hnRNP G shows striking similarities with the RBDs of several plant RNA-binding proteins.

The dichotomy between germ line and somatic line, and the origin of cell mortality.

The germ cells of extant animals are potentially immortal, whereas somatic cells are mortal, that is, they are able to carry out only a finite number of divisions. In this article we propose an evolutionary interpretation of these differences. We assume that germ cells of the earliest metazoans inherited immortality from their unicellular ancestor, while somatic cells acquired mortality by gaining new functions. It follows that cell mortality was under genetic control from the beginning of metazoan life.

A putative zinc-binding protein on lampbrush chromosome loops.

We used mAb A33/22, which recognizes a nuclear protein on the loops of amphibian lampbrush chromosomes, to select cDNA clone PwA33 from an expression library of the newt Pleurodeles waltl. A myc-tagged transcript of clone PwA33 was injected into Pleurodeles oocytes. The translation product localized in the germinal vesicle (GV) and was distributed on the lampbrush loops in a pattern identical to that of the endogenous protein. PwA33 encodes a 71 kDa protein with three distinct domains: a region rich in Cys/His residues that may form zinc fingers, a coiled-coil domain with potential for dimerization and a third 'rfp-like' domain that is shared by several other nuclear proteins. The putative zinc fingers and the coiled-coil domain resemble features in known nucleic acid-binding regulatory proteins. These structures, coupled with a distinctive pattern of expression in embryonic tissues, suggest that A33 may function as a regulatory protein during early development. It is unlikely that the large store of A33 in the GV is bound to DNA. Instead, its association with the nascent transcripts on the lampbrush chromosome loops suggests a role in pre-mRNA synthesis or processing.

Lampbrush W and Z heterochromosome characterization with a monoclonal antibody and heat-induced chromosomal markers in the newt Pleurodeles waltl: W chromosome plays a role in female sex determination.

Two subsets of lateral loops scattered on lampbrush chromosomes of the newt Pleurodeles waltl were characterized. One group was identified by labelling with a monoclonal antibody (A1). The second group was identified by the ability of the loops to be induced by heat treatment. Three loops of each subset were mapped on a short region of the two homologues of lampbrush bivalent IV. These regions appear to be heteromorphic because the six loops are always heterozygous. Five loops are found on one homologue and the sixth on the partner. The distribution of these markers in phenotypic females corresponding to the three sexual genotypes ZW, WW and ZZ shows an absolute correlation of the five loop group with the W chromosome and of the other loop with the Z chromosome. Therefore the heteromorphic regions of the homologues correspond to the differential segments of the heterochromosomes. The identification of a trisomic ZZW female suggests that the W chromosome bears female sex determinants. Furthermore the results show that heat induces loop development and that under normal conditions giant loop development is influenced by the sexual genotype.

Light and electron microscopic immunolocalization of two nuclear antigens in the liver of Pleurodeles waltl using monoclonal antibodies.

The distribution of 2 nuclear antigens in the interphase nuclei of liver of Pleurodeles waltl was determined with the help of monoclonal antibodies, using immunofluorescence for light microscopy and indirect immunoperoxidase and immunogold labeling procedures for electron microscopic localization. The antibodies C36/1 and A33/22 label antigens with relative molecular masses of 270 kDa and 80 kDa, and isoelectric points of 7.0 and 6.4, respectively. The liver of urodels is characterized by the presence of a peripheral layer of hematopoietic cells around the parenchymatous tissue formed by typical hepatocytes. The antibody C36/1 labels the nuclei of both types of cells, whereas the antibody A33/22 labels the nuclei of hepatocytes but not those of the peripheral hematopoietic cells. With both these antibodies, labeling, whenever observed, is restricted to fibrillar structures in the interchromatin space, i.e., to peri- and inter-chromatin fibrils; condensed chromatin, nucleoli, and nuclear envelope are not labeled.

Monoclonal antibodies to lampbrush chromosome antigens of Pleurodeles waltlii.

Germinal vesicles of oocytes from Pleurodeles waltlii were used for immunization of BALB/c mice to obtain hybridomas secreting monoclonal antibodies. The hybridomas were screened for reactivity of their antibodies against lampbrush chromosomes of oocytes, as revealed by indirect immunostaining. Antibodies labelling the lampbrush chromosomes were also tested on histological sections of oocytes, embryos, and larvae of Pleurodeles. Characterization of the antigens was accomplished through immunoblotting of two-dimensional electrophoretic gels of germinal vesicle proteins. The ten monoclonal antibodies giving a positive reaction were classed into five groups. Group 1, exemplified by antibody A33, recognizes all the lampbrush chromosome transcribing sites (loops). Moreover, it differentially labels the cell nuclei during embryonic and larval development. Group 2, antibody B71, also stains all the loops of the lampbrush chromosomes, but does not react with cell nuclei of embryos and larvae. Group 3, antibody A1, labels specific loops, some of which are heterozygous in the strain of P. waltlii used. These heterozygosities have allowed us to localize and to characterize a chromosomal segment on bivalent IV which is heteromorphic in the two partners of the bivalent. We suggest that this heteromorphism represents a morphological distinction between Z and W heterochromosomes. Moreover, this antibody reacts with only one transcription unit along a loop that contains several units. Group 4, antibody B24, stains the only two structures in the lampbrush chromosomes of P. waltlii that do not have a loop organization, the mass "M" and the spheres. Group 5, antibody A35, reacts with the chromomeres. The antigens corresponding to antibodies A33 and B24 have been identified as proteins, which have apparent molecular weights of 80 and 104 kilodaltons, respectively. They correspond to proteins abundant in the germinal vesicles. All the antibodies described here cross-react with the lampbrush chromosomes of five other species of Urodeles.

Fibronectin synthesis during oogenesis and early development of the amphibian Pleurodeles waltlii.

Fibronectin (FN) synthesis during oogenesis and early embryogenesis of the amphibian Pleurodeles waltlii was investigated. The isotopically labelled amino acids [3H]leucine and [35S]methionine were incubated with oocytes or microinjected into embryos. Newly synthesized FN was analysed by polyacrylamide gel electrophoresis, using the high resolution two-dimensional gel system described by O'Farrell. With this method and fluorography we demonstrate that FN synthesis begins during oogenesis. De novo synthesized FN appears during cleavage and gastrulation. Using actinomycin D we show the presence of maternal messenger RNA coding for FN. It is translated during the cleavage and gastrulation stages.