The heat shock cognate 80 gene of tomato is flanked by matrix attachment regions

Matrix attachment regions (MARs) are thought to participate in the organization and segregation of independent chromosomal loop domains. Although there are several reports on the action of MARs in the context of heterologous genes, information is more limited on the role of MARs associated with plant genes. Transgenic studies suggest that the upstream, intron and downstream regions of the developmentally regulated heat shock cognate 80 gene (HSC80) of tomato participate in chromatin organization. In this study, we tested the in vitro affinity of the HSC80 gene to chromosomal scaffolds prepared from shoot apices of tomato. We found that a 1.5 kb upstream region and a 1.4 kb downstream region, but not the intron region, are MARs. These MARs interact with tomato and pea scaffolds and bind regardless of the expression status of HSC80 in the tissue from which the nuclei were isolated. Comparison to two known yeast MARs ARS1 and CENIII, showed that the HSC80 5MAR binds more avidly to tomato scaffolds than ARS1, while no binding of CENIII was observed. Competition binding between the two HSC80 MARs indicated that the 5 MAR can outcompete the 3 MAR and not vice versa. Last, we observed that the interaction of the 3 MAR with the scaffold could result in an electrophoretic mobility shift resistant to SDS, protease, and phenol treatment. In conclusion, MARs whose binding properties can be clearly differentiated are closely flanking the HSC80 gene. The discovery of MARs in regions which have a distinct function in HSC80 transgenes but not in transient expression assays, is consistent with a chromosomal scaffold role in HSC80 gene regulation.     

Stratigraphy,facies and synsedimentary tectonics in the Jurassic Rosso Ammonitico Veronese (Altopiano di Asiago,NE Italy)

Summary The red, pelagic limestones of the Rosso Ammonitico Veronese (Upper Bajocian-Tithonian) of the Altopiano di Asiago area can be subdivided into eight facies. They differ from each other in structure (bedding style, presence and type of nodularity) and texture (nature of components, grain-vs mud-support, compactional features). Several discontinuities could be recognized, based on sedimentological or biostratigraphic evidence. In the context of a drowned platform, where sediments essentially consist of skeletal remains of both planktonic and benthic organism, the different facies are interpreted as reflecting a varying influence of currents on the winnowing of micrite and on triggering early cementation. Early cementation in turn, controlled the patterns of bioturbation and the response of sediments to later compaction and pressure-dissolution. At times, microbial mats, of unidentified nature, were important in trapping and binding sediment, giving rise to early lithified nodules and layers of stromatolitic aspect. The Rosso Ammonitico Veronese can be subdivided into three units: lower Rosso Ammonitico (RAI: Upper Bajocian-Lower Callovian), middle Rosso Ammonitico (RAM: Upper Callovian-Middle Oxfordian), and upper Rosso Ammonitico (RAS: Lower Kimmeridgian-Tithonian). Frequent ammonite moulds allow the precise dating of the base and top of each unit, and the documentation of facies heteropies and hiatusses in the more fossiliferous RAS. The lower unit (RAM) is massive and essentially nodular; the middle unit (RAM) is well bedded, non-nodular, and cherty; the upper unit (RAS) is richer in clay and typically nodular. The RAI and the RAS are present everywhere, though significant facies and thickness changes affect particularly the RAS; the RAM is much more variable, ranging from 0 to 10 metres. These variations, that may be gradual or abrupt, are inter-preted as the result of Middle-Late Callovian block-faulting which generated an irregular sea floor topography where the swells were more exposed to currents that continuously removed sediments, inducing long-lasting periods of nondeposition. Sediments preferentially accumulated in the adjacent lows. A confirmation of this hypothesis is provided by evidence of synsedimentary tectonics, described for the first time in the Rosso Ammonitico Veronese. Neptunian dykes, both vertical and horizontal, are developed at the top of the RAI and are filled with laminated sediments or collapse breccias. Glides of metre-size blocks and slumps are present at the top of the RAI and at the base of the RAM, respectively. Cm-thick layers of mud supported breccias are intercalated in the upper part of the RAI and within the RAM: they are interpreted as seismites. All these features document a tensional regime that generated fractures in more or less lithified sediments and failure along steep fault scarps or gently dipping slopes of tilted fault blocks. Recognition of this Callovian-Oxfordian tectonic activity shows that, after the Bajocian drowning, the Trento Plateau did not simply experience a uniform and general foundering: a small-scale block-faulting was still active and affected the pattern of facies distribution.     

Conservation of the alternative oxidase and enhancement of cyanide-resistant respiration in suspension-cultured pear fruit cells by inhibitors of macromolecular synthesis

The contribution of the alternative pathway to the respiration of suspension-cultured pear ( Pyrus communis cv. Passa Crasanne) cells was enhanced, often severalfold, within 2 to 4 days following the addition of cycloheximide, actinomycin D, or 2-(4-methyl-2,6-dinitroanalino)- N -methyl propionamide (D-MDMP). Concomitant inhibition of cellular protein synthesis by cycloheximide and actinomycin D was transient and incomplete. However, inhibition by D-MDMP was virtually complete (>97%) and persisted over several days. [³⁵S]-labelling and polyacrylamide gel separation indicated that cycloheximide precluded the appearance of discernable new proteins in mitochondria. Probes with monoclonal antibodies revealed a conservation of alternative oxidase protein levels in the mitochondria of inhibitor-treated cells. The data, appraised within the complexities of cell-culture dynamics, lead to the conclusion that the observed increases in capacity for cyanide-resistant respiration are the consequence, likely indirect, of inhibited protein synthesis with resultant retention and activation of constitutive alternative oxidase.     

Intra- and interobserver concordance in scoring Harris lines: A test on bone sections and radiographs

Little attention has been devoted to assessing the reproducibility of (paleo)pathological observations. Harris lines (HL) are among the markers most used to determine chronology of stresses suffered during growth. Nevertheless, their scoring entails remarkable methodological difficulty. Bone sections (S) and radiographs (R) of 29 adult tibiae of archeological provenance (medieval) were scored for HL by five observers. At regular intervals of time, each observer gave two independent counts on both series. Results show a) a substantial interobserver disagreement of HL estimates for both sectional and radiographic records, and b) a high level of intraobserver error. © 1994 Wiley-Liss, Inc.     

Fine mapping of the Autosomal Dominant Split Hand/Split Foot Locus on Chromosome 7, Band q21.3-q22.1

Split hand/split foot (SHFD) is a human developmental defect characterized by missing digits, fusion of remaining digits, and a deep median cleft in the hands and feet. Cytogenetic studies of deletions and translocations associated with this disorder have indicated that an autosomal dominant split hand/split foot locus (gene SHFD1) maps to 7q21-q22. To characterize the SHFD1 locus, somatic cell hybrid lines were constructed from cytogenetically abnormal individuals with SHFD. Molecular analysis resulted in the localization of 93 DNA markers to one of 10 intervals surrounding the SHFD1 locus. The translocation breakpoints in four SHFD patients were encompassed by the smallest region of overlap among the SHFD-associated deletions. The order of DNA markers in the SHFD1 critical region has been defined as PON–D7S812–SHFD1–D7S811–ASNS. One DNA marker, D7S811, detected altered restriction enzyme fragments in three patients with translocations when examined by pulsed-field gel electro-phoresis (PFGE). These data map SHFD1, a gene that is crucial for human limb differentiation, to a small interval in the q21.3-q22.1 region of human chromosome 7.     

CHROMOSOMAL VERSUS MITOCHONDRIAL DNA EVOLUTION: TRACKING THE EVOLUTIONARY HISTORY OF THE SOUTHWESTERN EUROPEAN POPULATIONS OF THE SOREX ARANEUS GROUP (MAMMALIA,INSECTIVORA)

The shrews of the Sorex araneus group have undergone a spectacular chromosome evolution. The karyotype of Sorex granarius is generally considered ancestral to those of Sorex coronatus and S. araneus. However, a sequence of 777 base pairs of the cytochrome b gene of the mitochondrial DNA (mtDNA) produces a quite different picture: S. granarius is closely related to the populations of S. araneus from the Pyrenees and from the northwestern Alps, whereas S. coronatus and S. araneus from Italy and the southern Alps represent two well-separated lineages. It is suggested that mtDNA and chromosomal evolution are in this case largely independant processes. Whereas mtDNA haplotypes are closely linked to the geographical history of the populations, chromosomal mutations were probably transmitted from one population to another. Available data suggest that the impressive chromosome polymorphism of this group is quite a recent phenomenon.     

Vertebrate homeobox genes

In the former part of the review the principal available data aboutHox genes, their molecular organisation and their expression in vertebrate embryos, with particular emphasis for mammals, are briefly summarized.In the latter part we analysed the expression of four mouse homeobox genes related to twoDrosophila genes expressed in the developing head of the fly: Emx1 and Emx2, related toems, and Otx1 and Otx2, related tootd.     

Oxidation and aromatic ring cleavage of 4-methoxy and 3,4-dimethoxycennamic acid by Lentinus edodes

Several products of metabolism and aromatic ring cleavage of 3-methoxy and 3,4-dimethoxycinnamic acid from ligninolytic cultures of Lentinus edodes were isolated and identified.     

A novel 2-oxoglutarate-dependent dioxygenase involved in vindoline biosynthesis: characterization,purification and kinetic properties

The enzyme, desacetoxyvindoline 4-hydroxylase, was purified to apparent homogeneity from Catharanthus roseus by ammonium sulfate precipitation and successive chromatography on Sephadex G-100, green 19-agarose, hydroxylapatite, -kg sepharose and Mono Q. The 4-hydroxylase was characterized by its strict specificity for position 4 of desacetoxyvindoline suggesting it to catalyze the second to last step in vindoline biosynthesis. The molecular mass of the native and denatured 4-hydroxylase was 45 kDa and 44.7 kDa, respectively, suggesting that the native enzyme is a monomer. Two-dimensional isoelectric focusing under denaturing conditions resolved the purified 4-hydroxylase into three charge isoforms of pIs 4.6, 4.7 and 4.8. The purified 4-hydroxylase exhibited no requirement for divalent cations, but inactive enzyme was reactivated in a time-dependent manner by incubation with ferrous ions. The enzyme was not inhibited by EDTA or SH-group reagents at concentrations up to 10 mM. The mechanism of action of desacetoxyvindoline 4-hydroxylase was investigated. The results of substrate interaction kinetics and product inhibition studies suggest an Ordered Ter Ter mechanism where -kg is the first substrate to bind followed by the binding of O₂ and desacetoxyvindoline. Their K _m values for -kg, O₂ and desacetoxyvindoline are 45 M, 45 M and 0.03 M, respectively. The first product to be released was deacetylvindoline followed by CO₂ and succinate, respectively.Abbreviations -kg -ketoglutarate or 2-oxoglutarate - NMT N-methyltransferase - SAM S-adenosyl-l-methionine - TLC thin layer chromatography - VBL vinblastine - VCR vincristine     

Polygalacturonase-inhibiting protein accumulates in Phaseolus vulgaris L. in response to wounding, elicitors and fungal infection

Polygalacturonase-inhibiting protein (PGIP) is a cell wall-associated protein that specifically binds to and inhibits the activity of fungal endopolygalacturonases. The Phaseolus vulgaris gene encoding PGIP has been cloned and characterized. Using a fragment of the cloned pgip gene as a probe in Northern blot experiments, it is demonstrated that the pgip mRNA accumulates in suspension-cultured bean cells following addition of elicitor-active oligogalacturonides or fungal glucan to the medium. Rabbit polyclonal antibodies specific for PGIP were generated against a synthetic peptide designed from the N-terminal region of PGIP; the antigenicity of the peptide was enhanced by coupling to KLH. Using the antibodies and the cloned pgip gene fragment as probes in Western and Northern blot experiments, respectively, it is shown that the levels of PGIP and its mRNA are increased in P. vulgaris hypocotyls in response to wounding or treatment with salicylic acid. Using gold-labeled goat-anti-rabbit secondary antibodies in EM studies, it has also been demonstrated that, in bean hypocotyls infected with Colletotrichum lindemuthianum, the level of PGIP preferentially increases in those cells immediately surrounding the infection site. The data support the hypothesis that synthesis of PGIP constitutes an active defense mechanism of plants that is elicited by signal molecules known to induce plant defense genes.