bluuebunny/crossref_metadata_2025 huggingface dataset:
Expression and growth inhibitory effect of decapentaplegic Vg-related protein 6: evidence for a regulatory role in keratinocyte differentiation.
Authors: Drozdoff V , Wall N A , Pledger W J
Published: May 2006

Decapentaplegic Vg-related protein 6 (DVR-6 or bone morphogenetic protein BMP-6) is a member of the DVR subgroup of the transforming growth factor beta superfamily, a large group of multifunctional signaling polypeptides that are expressed as secreted disulfide-bonded dimers proteolytically cleaved from larger precursors. The predominant expression of DVR-6 in the differentiating postmitotic layers of stratified squamous epithelia strongly suggests a role for DVR-6 in regulation of epithelial differentiation. In primary mouse keratinocytes induced to differentiate by suspension culture in methylcellulose, new expression of DVR-6 mRNA and protein was detected within 8 h among a majority of the suspended cells, which preceded the induction of expression of the suprabasal keratins K1 and K10. To test the hypothesis that DVR-6 is a keratinocyte growth regulatory factor, a retroviral expression vector expressing human DVR-6 was used to infect attached cultures of undifferentiated basal cells. Expression of DVR-6 in primary mouse keratinocytes before differentiation resulted in the secretion of prepro and processed (pro region) forms in the conditioned medium and a dramatic inhibition of cell growth. These findings suggest that inhibition of cell growth by DVR-6 may be a primary step in keratinocyte differentiation.
Identification of plant genetic loci involved in a posttranscriptional mechanism for meiotically reversible transgene silencing.
Authors: Dehio C , Schell J
Published: May 2006

Numerous reports describe phenomena of transgene silencing in plants, yet the underlying genetic and molecular mechanisms are poorly understood. We observed that regeneration of Arabidopsis thaliana plants transgenic for the rolB gene of Agrobacterium rhizogenes results in a selection for transgene silencing. Transgene silencing could be monitored in this system by reversion of the visible RolB phenotype. We report a phenotypic, molecular, and genetic characterization of a meiotically reversible transgene silencing phenomenon observed in a rolB transgenic line. In this line, the rolB gene is expressed strongly and uniformly in seedlings, but in the course of further development, the rolB gene is silenced erratically at a frequency that depends on the dosage of rolB. The silenced state is mitotically stable, while complete resetting of rolB gene expression occurs in seedlings of the following generation. The silencing of rolB correlates with a dramatic reduction of steady-state rolB transcripts, while rolB nuclear run-off transcripts are only moderately reduced. Therefore, rolB gene silencing seems to act predominantly at the posttranscriptional level. The process of rolB gene silencing was found to be affected by two extragenic modifier loci that influence both the frequency and the timing of rolB gene silencing during plant development. These genetic data demonstrate a direct involvement of defined plant genes in this form of gene silencing.
Selection of multiple human immunodeficiency virus type 1 variants that encode viral proteases with decreased sensitivity to an inhibitor of the viral protease.
Authors: Kaplan A H , Michael S F , Wehbie R S , Knigge M F , Paul D A , Everitt L , Kempf D J , Norbeck D W , Erickson J W , Swanstrom R
Published: May 2006

Inhibitors of the human immunodeficiency virus type 1 (HIV-1) protease represent a promising addition to the available agents used to inhibit virus replication in a therapeutic setting. HIV-1 is capable of generating phenotypic variants in the face of a variety of selective pressures. The potential to generate variants with reduced sensitivity to a protease inhibitor was examined by selecting for virus growth in cell culture in the presence of the protease inhibitor A-77003. Virus variants grew out in the presence of the inhibitor, and these variants encoded proteases with reduced sensitivity to the inhibitor. Variants were identified that encoded changes in each of the three subsites of the protease that interact with the inhibitor. HIV-1 displays significant potential for altering its interaction with this protease inhibitor, suggesting the need for multiple protease inhibitors with varying specificities.
Targeted mutation of the CREB gene: compensation within the CREB/ATF family of transcription factors.
Authors: Hummler E , Cole T J , Blendy J A , Ganss R , Aguzzi A , Schmid W , Beermann F , Schütz G
Published: May 2006

The cAMP response element binding protein (CREB) has been implicated as a key regulator in the transcriptional control of many genes. To assess the functional importance of CREB in vivo and its role in development, we used gene targeting to generate mice with a disruption of the CREB gene. Homozygous mutant mice appeared healthy and exhibited no impairment of growth or development. In this report we demonstrate that CREB and two other members of the CREB/ATF family, cAMP response element modulation protein (CREM) and activating transcription factor 1 (ATF1), appear to form a unique subgroup within this extensive class of transcription factors. Examination of CREM mRNA and protein levels in CREB mutant mice demonstrated overexpression of CREM in all tissues examined, but no change in ATF1 levels. These data demonstrate that CREB is not the sole mediator of cAMP-dependent transcriptional regulation and probably acts in concert with a specific subset of cAMP response element-binding proteins to transduce the cAMP signal and, in its absence, these same proteins can compensate for CREB function in vivo.
Arabidopsis floral homeotic gene BELL (BEL1) controls ovule development through negative regulation of AGAMOUS gene (AG).
Authors: Ray A , Robinson-Beers K , Ray S , Baker S C , Lang J D , Preuss D , Milligan S B , Gasser C S
Published: May 2006

Ovules are the developmental precursors of seeds. In angiosperms the ovules are enclosed within the central floral organs, the carpels. We have identified a homeotic mutation in Arabidopsis, "bell" (bel1), which causes transformation of ovule integuments into carpels. In situ hybridization analysis shows that this mutation leads to increased expression of the carpel-determining homeotic gene AGAMOUS (AG) in the mutant ovules. Introduction of a constitutively expressed AG transgene into wild-type plants causes the ovules to resemble those of bel1 mutants. We propose that the BEL1 gene product directs normal integument development, in part by suppressing AG expression in this structure. Our results allow expansion of the current model of floral organ identity to include regulation of ovule integument identity.
Likelihood ratios for DNA identification.
Authors: Collins A , Morton N E
Published: May 2006

Likelihood ratio (LR) tests are provided for the three alternatives to DNA identity: exclusion, coincidence, and kinship. The coincidence test uses the radius of coalescence to conserve the observed frequency of single band phenotypes. Genotype probabilities under kinship are derived for mating groups, specified relatives, and structured populations; and unbiased estimates of the genetic parameters are provided. The LR is made robust to gene frequency errors by specifying the mean matching probability, and the tolerable loss of information this entails is determined by LR theory. This straightforward application of the seminal work of Jerzy Neyman and Sewall Wright strongly supports the use of LRs and kinship for presentation of DNA evidence by expert witnesses and committees.
Ablation of E2A in recombinant adenoviruses improves transgene persistence and decreases inflammatory response in mouse liver.
Authors: Engelhardt J F , Ye X , Doranz B , Wilson J M
Published: May 2006

First-generation recombinant adenoviruses that lack E1 sequences have shown tremendous promise in animal and human models of gene therapy. Important limitations of these vectors are that recombinant gene expression is transient and inflammation occurs at the site of gene transfer. Our hypothesis for generating vectors with increased persistence is that present recombinant adenoviruses express viral proteins that stimulate cellular immune responses leading to destruction of the infected cells and repopulation of the organ with non-transgene-containing cells. This model predicts that further crippling of the virus will improve persistence and diminish pathology. We describe in this report second-generation recombinant adenoviruses harboring a beta-galactosidase-expressing transgene in which a temperature-sensitive mutation has been introduced into the E2A gene of an E1-deleted recombinant. At nonpermissive temperature, this virus fails to express late gene products, even when E1 is expressed in trans. The biology of this recombinant was studied in vivo in the context of mouse liver, a setting that is permissive for adenovirus type 5 replication. Animals that received the second-generation virus expressed the transgene for at least 70 days, whereas expression of the first-generation virus was no longer than 14 days. In addition, the inflammatory response, as measured by infiltration of CD8+ T cells, was blunted and delayed in livers infected with second-generation virus. These studies illustrate that modifications that disrupt structural protein expression in recombinant adenoviruses may be useful in enhancing their utility for gene therapy.
Double-stranded RNA-dependent protein kinase activates transcription factor NF-kappa B by phosphorylating I kappa B.
Authors: Kumar A , Haque J , Lacoste J , Hiscott J , Williams B R
Published: May 2006

The induction of interferon (IFN) genes by viruses or double-stranded RNA (dsRNA) requires the assembly of a complex set of transcription factors on responsive DNA elements of IFN gene promoters. One of the factors necessary for regulating IFN-beta gene transcription is nuclear factor NF-kappa B, the activation of which is triggered by dsRNA. It has previously been suggested that the dsRNA-activated p68 protein kinase (PKR) may act as an inducer-receptor, transducing the signal from dsRNA to NF-kappa B through phosphorylation of the inhibitor I kappa B. We present direct evidence that PKR can phosphorylate I kappa B-alpha (MAD-3) and activate NF-kappa B DNA binding activity in vitro. We further show that dsRNA induces an unusual phosphorylated form of I kappa B-alpha. The expression of a transdominant mutant PKR is able to perturb the dsRNA-mediated signaling pathway in vivo, suggesting a role for this kinase in IFN-beta gene induction.
DNA-binding and trans-activation properties of Drosophila E2F and DP proteins.
Authors: Dynlacht B D , Brook A , Dembski M , Yenush L , Dyson N
Published: May 2006

The temporal activation of E2F transcriptional activity appears to be an important component of the mechanisms that prepare mammalian cells for DNA replication. Regulation of E2F activity appears to be a highly complex process, and the dissection of the E2F pathway will be greatly facilitated by the ability to use genetic approaches. We report the isolation of two Drosophila genes that can stimulate E2F-dependent transcription in Drosophila cells. One of these genes, dE2F, contains three domains that are highly conserved in the human homologs E2F-1, E2F-2, and E2F-3. Interestingly, one of these domains is highly homologous to the retinoblastoma protein (RB)-binding sequences of human E2F genes. The other gene, dDP, is closely related to the human DP-1 and DP-2 genes. We demonstrate that dDP and dE2F interact and cooperate to give sequence-specific DNA binding and optimal trans-activation. These features suggest that endogenous Drosophila E2F, like human E2F, may be composed of heterodimers and may be regulated by RB-like proteins. The isolation of these genes will provide important reagents for the genetic analysis of the E2F pathway.
Phosphorylation and inactivation of protein phosphatase 1 by cyclin-dependent kinases.
Authors: Dohadwala M , da Cruz e Silva E F , Hall F L , Williams R T , Carbonaro-Hall D A , Nairn A C , Greengard P , Berndt N
Published: May 2006

Protein phosphatase 1 and protein phosphatase 2A contain potential phosphorylation sites for cyclin-dependent kinases. In the present study we found that rabbit skeletal muscle protein phosphatase 1, as well as recombinant protein phosphatase 1 alpha and protein phosphatase 1 gamma 1, but not protein phosphatase 2A, was phosphorylated and inhibited by cdc2/cyclin A and cdc2/cyclin B. Phosphopeptide mapping and phospho amino acid analysis suggested that the phosphorylation site was located at a C-terminal threonine. Neither cdc2/cyclin A nor cdc2/cyclin B phosphorylated an active form of protein phosphatase 1 alpha in which Thr-320 had been mutated to alanine, indicating that the phosphorylation occurred at this threonine residue. Furthermore, protein phosphatase 1, but not protein phosphatase 2A, activity was found to change during the cell cycle of human MG-63 osteosarcoma cells. The observed oscillations in protein phosphatase 1 activity during the cell cycle may be due, at least in part, to phosphorylation of protein phosphatase 1 by cyclin-dependent kinases. Together, the results suggest a mechanism for direct regulation of protein phosphatase 1 activity.
Active site of the mRNA-capping enzyme guanylyltransferase from Saccharomyces cerevisiae: similarity to the nucleotidyl attachment motif of DNA and RNA ligases.
Authors: Fresco L D , Buratowski S
Published: May 2006

Nascent mRNA chains are capped at the 5' end by the addition of a guanylyl residue to form a G(5')ppp(5')N ... structure. During the capping reaction, the guanylyltransferase (GTP:mRNA guanylyltransferase, EC 2.7.7.50) is reversibly and covalently guanylylated. In this enzyme-GMP (E-GMP) intermediate, GMP is linked to the epsilon-amino group of a lysine residue via a phosphoamide bond. Lys-70 was identified as the GMP attachment site of the Saccharomyces cerevisiae guanylyltransferase (encoded by the CEG1 gene) by guanylylpeptide sequencing. CEG1 genes with substitutions at Lys-70 were unable to support viability in yeast and produced proteins that were not guanylylated in vitro. The CEG1 active site exhibits sequence similarity to the active sites of viral guanylyltransferases and polynucleotide ligases, suggesting similarity in the mechanisms of nucleotidyl transfer catalyzed by these enzymes.
Alternative splicing generates secretory isoforms of human CD1.
Authors: Woolfson A , Milstein C
Published: May 2006

Human CD1 genes are a family of five non-polymorphic genes that, although homologous to both class I and II major histocompatibility complex genes, map to chromosome 1. Only three of the antigens, CD1a, -b, and -c, have been clustered with monoclonal antibodies. They are noncovalently associated with beta 2-microglobulin and may function as nonclassical antigen-presenting molecules. Here we analyze their expression in mouse myeloma transfectants and human thymocytes and find mRNA splicing complexity. This manifests itself as incomplete splicing, alternative splicing, utilization of cryptic splice sites, and the generation of alternative reading frames. In the case of CD1A transfectants, we demonstrate that the major protein product is secreted and show by amino acid sequence analysis that this is derived from an unspliced transcript. A second major CD1a component appears to be retained intracellularly. The production of alternatively spliced transcripts in the thymus is not a feature of all CD1 genes. Although in the case of CD1A only the transcript encoding the cell surface CD1a isoform is found, CD1C and -E produce complex intrathymic splicing patterns. The CD1C transcripts predict the expression of a secreted CD1c isoform in the human thymus, which we detect in CD1C transfectant culture supernatants. CD1 gene expression is thus characterized by considerable splicing complexity, and the difference between the splicing patterns found in different environments suggests that this is tissue specific.
Phylogeny from function: evidence from the molecular fossil record that tRNA originated in replication, not translation.
Authors: Maizels N , Weiner A M
Published: May 2006

We propose a phylogeny for the evolution of tRNA that is based on the ubiquity and conservation of tRNA-like structures in the replication of contemporary genomes. This phylogeny is unique in suggesting that the function of tRNA in replication dates back to the very beginnings of life on earth, before the advent of templated protein synthesis. The origin we propose for tRNA has distinct implications for the order in which other components of the modern translational apparatus evolved. We further suggest that the "top half" of modern tRNA-a coaxial stack of the acceptor stem on the T psi C arm--is the ancient structural and functional domain and that the "bottom half" of tRNA--a coaxial stack of the dihydrouracil arm on the anticodon arm--arose later to provide additional specificity.
Proterozoic and early Cambrian protists: evidence for accelerating evolutionary tempo.
Authors: Knoll A H
Published: May 2006

In rocks of late Paleoproterozoic and Mesoproterozoic age (ca. 1700-1000 million years ago), probable eukaryotic microfossils are widespread and well preserved, but assemblage and global diversities are low and turnover is slow. Near the Mesoproterozoic-Neoproterozoic boundary (1000 million years ago), red, green, and chromophytic algae diversified; molecular phylogenies suggest that this was part of a broader radiation of "higher" eukaryotic phyla. Observed diversity levels for protistan microfossils increased significantly at this time, as did turnover rates. Coincident with the Cambrian radiation of marine invertebrates, protistan microfossils again doubled in diversity and rates of turnover increased by an order of magnitude. Evidently, the Cambrian diversification of animals strongly influenced evolutionary rates, within clades already present in marine communities, implying an important role for ecology in fueling a Cambrian explosion that extends across kingdoms.
Dynamics of adaptation and diversification: a 10,000-generation experiment with bacterial populations.
Authors: Lenski R E , Travisano M
Published: May 2006

We followed evolutionary change in 12 populations of Escherichia coli propagated for 10,000 generations in identical environments. Both morphology (cell size) and fitness (measured in competition with the ancestor) evolved rapidly for the first 2000 generations or so after the populations were introduced into the experimental environment, but both were nearly static for the last 5000 generations. Although evolving in identical environments, the replicate populations diverged significantly from one another in both morphology and mean fitness. The divergence in mean fitness was sustained and implies that the populations have approached different fitness peaks of unequal height in the adaptive landscape. Although the experimental time scale and environment were microevolutionary in scope, our experiments were designed to address questions concerning the origin as well as the fate of genetic and phenotypic novelties, the repeatability of adaptation, the diversification of lineages, and thus the causes and consequences of the uniqueness of evolutionary history. In fact, we observed several hallmarks of macroevolutionary dynamics, including periods of rapid evolution and stasis, altered functional relationships between traits, and concordance of anagenetic and cladogenetic trends. Our results support a Wrightian interpretation, in which chance events (mutation and drift) play an important role in adaptive evolution, as do the complex genetic interactions that underlie the structure of organisms.
Selection of DNA clones with enhancer sequences.
Authors: Asoh S , Lee-Kwon W , Mouradian M M , Nirenberg M
Published: May 2006

A method is described for selection of DNA clones that contain enhancer sequences that activate gene expression. An Escherichia coli-rodent cell shuttle vector, pPyE0, was used that contains polyoma viral DNA without the polyoma enhancer region. Replication of pPyE0 DNA in mouse cells is markedly reduced due to deletion of the polyoma enhancer region. Insertion of mouse genomic DNA fragments that contain putative enhancer sequences into pPyE0 adjacent to the polyoma origin of replication restored, to varying extents, the ability of the recombinant plasmid DNA to replicate in mouse cells. Recombinant plasmids that replicate well in mouse cells, therefore, are amplified selectively. Transfection of mouse neuroblastoma or fibroblast cells that constitutively synthesize polyoma large tumor antigen with a library of mouse genomic DNA fragments inserted in pPyE0 yielded many recombinant plasmids. DNA inserts from each of the 16 clones that were examined stimulated the expression of an enhancerless chloramphenicol acetyltransferase reporter gene. The DNA inserts from 4 clones that were studied resulted in 4- to 13-fold increases in chloramphenicol acetyltransferase mRNA in transfected mouse cells. Nucleotide sequence analysis led to the identification of 5 genomic DNA clones that were obtained by selection. All of the homologies found were to regions of DNA that are thought to be involved in the regulation of gene expression.
Sertoli cell differentiation on basement membrane is mediated by the c-fos protooncogene.
Authors: Papadopoulos V , Dym M
Published: May 2006

When Sertoli cells of the seminiferous epithelium in the testis were cultured on Matrigel (a reconstituted basement membrane), laminin, or one of the biologically active laminin-derived peptides (YIGSR, SIKVAV, or RGD), they exhibited dramatic changes in morphology accompanied by changes in protein secretion and gene expression, including a rapidly induced stimulation of c-fos mRNA. To examine the role of c-fos in Sertoli cell attachment, spreading, and differentiation on extracellular matrix, we constructed sense and antisense c-fos phosphorothioate oligodeoxynucleotides (ODNs). Sertoli cells, in small clumps of 10-20 cells, cultured on Matrigel or laminin in the presence of ODNs antisense to c-fos (30 micrograms/ml) did not spread for the first 10-20 hr. After that time, normal spreading occurred, probably as a result of ODN degradation. Cells cultured in medium supplemented with ODNs sense to c-fos (30 micrograms/ml), or without ODNs, spread within 30 min to 1 hr, and after 12 hr a monolayer was established. Furthermore, incubation of Sertoli cells with ODNs antisense to c-fos resulted in a significant reduction of c-fos protein levels, whereas treatment with ODNs sense to c-fos barely affected c-fos protein expression. These data indicate that c-fos may mediate the events involved in Sertoli cell attachment and spreading upon contact of the cells with extracellular matrix.
Glycopeptide enkephalin analogues produce analgesia in mice: evidence for penetration of the blood-brain barrier.
Authors: Polt R , Porreca F , Szabò L Z , Bilsky E J , Davis P , Abbruscato T J , Davis T P , Harvath R , Yamamura H I , Hruby V J
Published: May 2006

Most peptides have not proved useful as neuroactive drugs because they are blocked by the blood-brain barrier and do not reach their receptors within the brain. Intraperitoneally administered L-serinyl beta-D-glucoside analogues of [Met5]enkephalin (glycopeptides) have been shown to be transported across the blood-brain barrier to bind with targeted mu- and delta-opioid receptors in the mouse brain. The opioid nature of the binding has been demonstrated with intracerebroventricularly administered naloxone. Paradoxically, glucosylation decreases the lipophilicity of the peptides while promoting transport across the lipophilic endothelial layer. It is suggested that glucose transporter GLUT-1 is responsible for the transport of the peptide message. Profound and long-lasting analgesia has been observed in mice (tail-flick and hot-plate assays) with two of the glycopeptide analogues when administered intraperitoneally.
Transmembrane topology of two kainate receptor subunits revealed by N-glycosylation.
Authors: Wo Z G , Oswald R E
Published: May 2006

Glutamate receptors are the primary excitatory neurotransmitter receptors in vertebrate brain and are of critical importance to a wide variety of neurological processes. Recent reports suggest that ionotropic glutamate receptors may have a unique transmembrane topology not shared by other ligand-gated ion channels. We report here the cloning of cDNAs from goldfish brain encoding two homologous kainate receptors with protein molecular masses of 41 kDa. Using a cell-free translation/translocation system, we show that (i) a portion of these receptors previously thought to be a large intracellular loop is actually located extracellularly and (ii) the putative second transmembrane region of the receptor thought to line the ion channel may not be a true membrane-spanning domain. An alternative model for the transmembrane topology of kainate receptors is proposed that could potentially serve as a framework for future detailed study of the structure of this important class of neurotransmitter receptors.
Prolactin activates the interferon-regulated p91 transcription factor and the Jak2 kinase by tyrosine phosphorylation.
Authors: David M , Petricoin E F , Igarashi K , Feldman G M , Finbloom D S , Larner A C
Published: May 2006

The prolactin (PRL) receptor is a member of the family of cytokine receptors that lack intrinsic tyrosine kinase activity but contain two conserved cysteines in their N-terminal regions and a WSXWS motif adjacent to their transmembrane domains. In a manner similar to the interferons (IFNs), exposure of cells to PRL results in tyrosine phosphorylation of several cellular proteins and the rapid transcriptional induction of the IFN regulatory factor 1 gene. In this communication, we demonstrate that treatment of rat Nb2 lymphoma cells with PRL activates a latent protein factor so that it binds to an enhancer in the IFN regulatory factor 1 gene. This enhancer has been shown to be required for IFN-gamma-activated expression of this gene. PRL-induced assembly of the DNA binding complex, PRL-stimulated factor, required tyrosine phosphorylation. PRL-stimulated factor contained at least one protein that was antigenically similar to the p91 transcription factor, a component of several transcription complexes required for cytokine-activated gene expression. PRL not only induced the tyrosine phosphorylation of p91 but also induced tyrosine phosphorylation of Jak2, a tyrosine kinase required for IFN-gamma-activated gene expression. These results provide evidence for a signaling mechanism, some of whose components are shared by both PRL and IFN-gamma receptors, that results in the expression of early response genes.
Isolation and characterization of the gene encoding 2,3-oxidosqualene-lanosterol cyclase from Saccharomyces cerevisiae.
Authors: Shi Z , Buntel C J , Griffin J H
Published: May 2006

The ERG7 gene encoding oxidosqualene-lanosterol cyclase [(S)-2,3-epoxysqualene mutase (cyclizing, lanosterol forming), EC 5.4.99.7] from Saccharomyces cerevisiae has been cloned by genetic complementation of a cyclase-deficient erg7 strain. The DNA sequence of this gene has been determined and found to contain an open reading frame of 2196 nt (including stop codon) that encodes a predicted protein of 731 amino acids. The predicted molecular mass of the S. cerevisiae cyclase, 83.4 kDa, is similar to the predicted molecular masses of the oxidosqualene-lanosterol cyclase from Candida albicans and the oxidosqualene-cycloartenol cyclase from Arabidopsis thaliana, as well as to the molecular masses assigned to vertebrate oxidosqualene-lanosterol cyclases; however, it is substantially larger than the molecular mass assigned to purified S. cerevisiae cyclase. At the level of DNA and predicted amino acid sequences, the S. cerevisiae and C. albicans cyclases share 56% and 63% identity, respectively. Tryptophan and tyrosine residues are unusually abundant in the predicted amino acid sequences of (oxido)-squalene cyclases, leading to a hypothesis that electron-rich aromatic side chains from these residues are essential features of cyclase active sites.
Cloning of four cyclins from maize indicates that higher plants have three structurally distinct groups of mitotic cyclins.
Authors: Renaudin J P , Colasanti J , Rime H , Yuan Z , Sundaresan V
Published: May 2006

While a large number of cyclins have been described in animals and yeasts, very limited information is available regarding cyclins in plants. We describe here the isolation of cDNA clones encoding four putative mitotic cyclins from maize. All four cyclins were able to induce maturation of Xenopus oocytes, demonstrating that they can act as mitotic cyclins in this system. Northern analysis showed that all four cyclins were expressed only in actively dividing tissues and organs, with a stronger correlation between expression and mitotic activity than is observed with cdc2. The deduced protein sequences suggest that the four maize cyclins belong to the cyclin A and B families identified from animal and yeast studies but that they cannot be described easily as either A-type or B-type cyclins. However, comparison with previously cloned plant cyclins shows that cyclins in higher plants form three distinct structural groups that have been conserved in both monocotyledonous and dicotyledonous species and that cyclins from all three groups are present within a single plant species.
Increase of synaptic density and memory retention by a peptide representing the trophic domain of the amyloid beta/A4 protein precursor.
Authors: Roch J M , Masliah E , Roch-Levecq A C , Sundsmo M P , Otero D A , Veinbergs I , Saitoh T
Published: May 2006

The secreted form (sAPP) of the Alzheimer amyloid beta/A4 protein precursor (APP) has been shown to be involved in the in vitro regulation of fibroblast growth and neurite extension from neuronal cells. The active site of sAPP responsible for these functions is within a small domain just C-terminal to the Kunitz-type protease inhibitor (KPI) insertion site. We report here that a 17-mer peptide, containing this active domain of sAPP, can induce cellular and behavioral changes when infused into rat brains. After 2 weeks of APP 17-mer peptide infusion, the animals were tested for reversal learning and memory retention and were sacrificed for morphological examination of brains. We found that administration of the APP 17-mer peptide resulted in an 18% increase in the number of presynaptic terminals in the frontoparietal cortex. At the behavioral level, 17-mer-infused animals with nonimpaired learning capability showed an increased memory retention that seemed to interfere with reversal learning performance. This APP 17-mer effect on memory retention was not observed in animals with impaired initial learning capacity. These results suggest that APP is involved in memory retention through its effect on synaptic structure.
Birth of normal young after electrofusion of mouse oocytes with round spermatids.
Authors: Ogura A , Matsuda J , Yanagimachi R
Published: May 2006

Normally, round spermatids, the youngest male germ cells with a set of haploid chromosomes, cannot fertilize mature oocytes. However, when mouse spermatids were fused with oocytes, some of the resulting zygotes developed into normal fertile mice of either sex. This demonstrates that the nuclei of spermatids can provide the paternally imprinted chromosomes needed for full embryonic development and also that the complex postmeiotic modifications involved in sperm formation in the testis and their maturation in the epididymis merely serve to facilitate natural delivery of the paternal genome. This finding may find an application in the treatment of male infertility due to defective spermiogenesis/sperm maturation.
Continuous network of endoplasmic reticulum in cerebellar Purkinje neurons.
Authors: Terasaki M , Slater N T , Fein A , Schmidek A , Reese T S
Published: May 2006

Purkinje neurons in rat cerebellar slices injected with an oil drop saturated with 1,1'-dihexadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate [DiIC16(3) or DiI] to label the endoplasmic reticulum were observed by confocal microscopy. DiI spread throughout the cell body and dendrites and into the axon. DiI spreading is due to diffusion in a continuous bilayer and is not due to membrane trafficking because it also spreads in fixed neurons. DiI stained such features of the endoplasmic reticulum as densities at branch points, reticular networks in the cell body and dendrites, nuclear envelope, spines, and aggregates formed during anoxia nuclear envelope, spines, and aggregates formed during anoxia in low extracellular Ca2+. In cultured rat hippocampal neurons, where optical conditions provide more detail, DiI labeled a clearly delineated network of endoplasmic reticulum in the cell body. We conclude that there is a continuous compartment of endoplasmic reticulum extending from the cell body throughout the dendrites. This compartment may coordinate and integrate neuronal functions.
Characterization of the promoter region of the gene for the rat neutral and basic amino acid transporter and chromosomal localization of the human gene.
Authors: Yan N , Mosckovitz R , Gerber L D , Mathew S , Murty V V , Tate S S , Udenfriend S
Published: May 2006

The promoter region of the rat kidney neutral and basic amino acid transporter (NBAT) gene has been isolated and sequenced. The major transcription initiation site was mapped by primer extension. The entire promoter region and a set of 5' deletions within it were expressed at a high level in LLC-PK1 cells using the luciferase indicator gene. Positive and negative regulatory elements in the promoter region were observed. A human genomic clone of the transporter was also obtained and was used to localize the NBAT gene at the p21 region of chromosome 2.
Nitric oxide and cGMP cause vasorelaxation by activation of a charybdotoxin-sensitive K channel by cGMP-dependent protein kinase.
Authors: Archer S L , Huang J M , Hampl V , Nelson D P , Shultz P J , Weir E K
Published: May 2006

Nitric oxide (NO)-induced relaxation is associated with increased levels of cGMP in vascular smooth muscle cells. However, the mechanism by which cGMP causes relaxation is unknown. This study tested the hypothesis that activation of Ca-sensitive K (KCa) channels, mediated by a cGMP-dependent protein kinase, is responsible for the relaxation occurring in response to cGMP. In rat pulmonary artery rings, cGMP-dependent, but not cGMP-independent, relaxation was inhibited by tetraethylammonium, a classical K-channel blocker, and charybdotoxin, an inhibitor of KCa channels. Increasing extracellular K concentration also inhibited cGMP-dependent relaxation, without reducing vascular smooth muscle cGMP levels. In whole-cell patch-clamp experiments, NO and cGMP increased whole-cell K current by activating KCa channels. This effect was mimicked by intracellular administration of (Sp)-guanosine cyclic 3',5'-phosphorothioate, a preferential cGMP-dependent protein kinase activator. Okadaic acid, a phosphatase inhibitor, enhanced whole-cell K current, consistent with an important role for channel phosphorylation in the activation of NO-responsive KCa channels. Thus NO and cGMP relax vascular smooth muscle by a cGMP-dependent protein kinase-dependent activation of K channels. This suggests that the final common pathway shared by NO and the nitrovasodilators is cGMP-dependent K-channel activation.
Enterokinase, the initiator of intestinal digestion, is a mosaic protease composed of a distinctive assortment of domains.
Authors: Kitamoto Y , Yuan X , Wu Q , McCourt D W , Sadler J E
Published: May 2006

Enterokinase is a protease of the intestinal brush border that specifically cleaves the acidic propeptide from trypsinogen to yield active trypsin. This cleavage initiates a cascade of proteolytic reactions leading to the activation of many pancreatic zymogens. The full-length cDNA sequence for bovine enterokinase and partial cDNA sequence for human enterokinase were determined. The deduced amino acid sequences indicate that active two-chain enterokinase is derived from a single-chain precursor. Membrane association may be mediated by a potential signal-anchor sequence near the amino terminus. The amino terminus of bovine enterokinase also meets the known sequence requirements for protein N-myristoylation. The amino-terminal heavy chain contains domains that are homologous to segments of the low density lipoprotein receptor, complement components C1r and C1s, the macrophage scavenger receptor, and a recently described motif shared by the metalloprotease meprin and the Xenopus A5 neuronal recognition protein. The carboxyl-terminal light chain is homologous to the trypsin-like serine proteases. Thus, enterokinase is a mosaic protein with a complex evolutionary history. The amino acid sequence surrounding the amino terminus of the enterokinase light chain is ITPK-IVGG (human) or VSPK-IVGG (bovine), suggesting that single-chain enterokinase is activated by an unidentified trypsin-like protease that cleaves the indicated Lys-Ile bond. Therefore, enterokinase may not be the "first" enzyme of the intestinal digestive hydrolase cascade. The specificity of enterokinase for the DDDDK-I sequence of trypsinogen may be explained by complementary basic-amino acid residues clustered in potential S2-S5 subsites.
Common major histocompatibility complex class II markers in clinical variants of cicatricial pemphigoid.
Authors: Yunis J J , Mobini N , Yunis E J , Alper C A , Deulofeut R , Rodriguez A , Foster C S , Marcus-Bagley D , Good R A , Ahmed A R
Published: May 2006

Cicatricial pemphigoid (CP) is a chronic autoimmune blistering disease affecting multiple mucous membranes derived from stratified squamous epithelium and occasionally the skin. CP has a wide spectrum of disease manifestations. Patients with oral pemphigoid (OP) have a benign self-limited disease in which pathological changes are restricted to the oral mucosa. On the other hand, patients with ocular cicatricial pemphigoid (OCP), a chronic condition marked with relapses and remissions, have ocular involvement and also perhaps involvement of other mucous membranes. All clinical subsets are characterized by the presence of a similar anti-basement zone autoantibody. The factors that determine the development of one form of CP or the other are not known. In a previous study, we described the association between OCP and the DQB1*0301 allele (P = 0.006). In this study, we have analyzed 22 Caucasian patients with OP and their family members for major histocompatibility complex DRB generic, DQA1, and DQB1 allele associations by PCR-sequence-specific oligonucleotide probe hybridization. The results were compared to those obtained from 17 Caucasian patients with OCP and to control Caucasian alleles and haplotypes. The DQB1*0301 allele frequency was 38.6% in OP, 52.9% in OCP, and 17.8% in controls. Statistically significant associations were detected between the DQB1*0301 allele and both OP (P = 0.0047) and OCP (P < 0.0001). In addition, DRB1*04 showed a statistically significant association (P = 0.005) with OCP when compared to controls. Analysis of major histocompatibility complex class II haplotypes showed significant statistical associations between both OCP and OP and the HLA-DRB1*04, DRB4*0101, DQA1*03, DQB1*0301 haplotype (P < 0.0001 and P = 0.0012, respectively). Our results indicate that DQB1*0301 is a marker of both oral and ocular forms of CP. The analysis of the amino acid sequence of the DQB1 alleles present in both OP and OCP suggested that amino acid residues at position 57 and positions 71-77 may also be markers of CP.
High levels of a retinoic acid-generating dehydrogenase in the meso-telencephalic dopamine system.
Authors: McCaffery P , Dräger U C
Published: May 2006

Retinoic acid is synthesized from retinaldehyde by several different dehydrogenases, which are arranged in conserved spatial and developmentally regulated patterns. Here we show for the mouse that a class-1 aldehyde dehydrogenase, characterized by oxidation and disulfiram sensitivity, is found in the brain at high levels only in the basal forebrain. It is present in axons and terminals of a subpopulation of dopaminergic neurons of the mesostriatal and mesolimbic system, forming a retinoic acid-generating projection from the ventral tegmentum to the corpus striatum and the shell of the nucleus accumbens. In the striatum the projection is heaviest to dorsal and rostral regions, declining gradually toward ventral. The enzyme is expressed early in development, shortly after appearance of tyrosine hydroxylase. Other dopaminergic neurons in the brain, as well as the chromaffin cells of the adrenal medulla, do not contain this dehydrogenase. The presence of this enzyme may be a factor in the long-term success of transplants of dopaminergic cells to the corpus striatum in Parkinson disease, and it may play a role in parkinsonism and catatonia due to disulfiram (Antabuse) neurotoxicity.
Increased disease susceptibility of transgenic tobacco plants with suppressed levels of preformed phenylpropanoid products.
Authors: Maher E A , Bate N J , Ni W , Elkind Y , Dixon R A , Lamb C J
Published: May 2006

It has been proposed that natural products synthesized by plants contribute to their resistance to pests and pathogens. We show here that transgenic tobacco plants with suppressed levels of the phenylpropanoid biosynthetic enzyme phenylalanine ammonia-lyase (L-phenylalanine ammonia-lyase, EC 4.3.1.5) and correspondingly low levels of chlorogenic acid, the major soluble leaf phenylpropanoid product, exhibit more rapid and extensive lesion development than wild-type plants after infection by the virulent fungal pathogen Cercospora nicotianae. These observations provide direct evidence that phenylpropanoid products contribute to disease limitation. No induction of transcripts encoding phenylalanine ammonia-lyase or the lignin branch pathway enzyme caffeic acid O-methyltransferase was observed during the infection and there was no perturbation in the pattern of soluble phenylpropanoids. Hence, increased disease susceptibility does not involve inhibition of a pathogen-induced response but likely reflects inhibition of the developmental accumulation of chlorogenic acid. Demonstration of the contribution of such preformed protectants to plant health identifies attractive targets for manipulation by breeding or gene transfer to reduce the quantitative impact of disease.
Cyclin D1 overexpression vs. retinoblastoma inactivation: implications for growth control evasion in non-small cell and small cell lung cancer.
Authors: Schauer I E , Siriwardana S , Langan T A , Sclafani R A
Published: May 2006

The cyclin-dependent kinases and their associated regulatory cyclins control cell cycle progression and cell growth. Antibodies against these proteins were used to determine their levels in several lung tumor-derived cell lines and a "normal" immortalized bronchoepithelial cell line in order to investigate their potential roles in the etiology of lung cancer. All the cell lines expressed roughly equal levels of cdk-1; cdk-2; PSTAIRE-sequence containing kinases; proliferating cell nuclear antigen; and cyclins A, B1, and E. Cyclin D1, however, was present at 4- to 100-fold higher levels in 11 of 12 non-small cell lung cancer cell lines than in the bronchoepithelial line and all but one of the small cell lung cancer lines. Furthermore, immunoblots of the retinoblastoma gene product, pRB, revealed a perfect correlation between pRB levels and tumor type with normal levels of phosphorylation-competent pRB in all of the non-small cell lung cancer lines and undetectable levels of pRB in all of the small cell lung cancer lines. These data suggest the possibility that small cell and non-small cell lung cancer may evade normal growth controls by different mechanisms: loss of the proliferation inhibitor pRB in small cell lung cancer and overexpression of the growth promoting cyclin D1 in non-small cell lung cancer.
Transcriptional activation by TAL1 and FUS-CHOP proteins expressed in acute malignancies as a result of chromosomal abnormalities.
Authors: Sánchez-García I , Rabbitts T H
Published: May 2006

Proteins that appear to participate in transcriptional control of gene expression are increasingly implicated in leukemias and malignant solid tumors. We report here that the N-terminal domains of the proteins TAL1 (ectopically activated in T-cell acute leukemias after chromosomal abnormalities caused by V-D-J recombinase error) (V, variable; D, diversity; J, joining) and FUS-CHOP (a liposarcoma tumor-specific fusion protein that is produced as a result of a chromosomal translocation) can function as transcription activators of specific responsive reporter genes. The result with TAL1 provides evidence that transcriptional activation can be mediated by a gene activated by translocation in T-cell acute leukemias. In the case of the liposarcoma, transactivation by the FUS-CHOP protein occurs because the FUS transcriptional activation domain is added to the DNA-binding CHOP protein normally lacking such activity. Therefore, the association of transcriptional activation and DNA-binding elements is a common consequence in proteins activated or newly created as fusion proteins after chromosomal translocations in acute leukemias and in malignant solid tumors.
Host fidelity is an effective premating barrier between sympatric races of the apple maggot fly.
Authors: Feder J L , Opp S B , Wlazlo B , Reynolds K , Go W , Spisak S
Published: May 2006

Models of sympatric speciation for phytophagous insects posit a central role for host plant-associated mating as a premating isolating mechanism in lieu of geographic barriers to gene flow. Here, by means of three mark-and-recapture studies, we confirm that host fidelity (i.e., the tendency of an insect to reproduce on the same host species that it used in earlier life-history stages) restricts gene flow between sympatric apple- and hawthorn-infesting races of Rhagoletis pomonella (Diptera: Tephritidae) to approximately 6% per generation. Genetically based differences in host preference, adult eclosion under the "correct" host species, and allochronic isolation contribute to host fidelity in various degrees in the races. The results verify that host-associated adaptation can produce reproductive isolation as a correlated character (a key premise of sympatric speciation). The study also represents one of the few or perhaps only example in animals where the intra-specific isolating effects of specific phenotypes have been quantified in nature.
Agrobacterium tumefaciens transfers single-stranded transferred DNA (T-DNA) into the plant cell nucleus.
Authors: Tinland B , Hohn B , Puchta H
Published: May 2006

Transferred DNA (T-DNA) is transferred as a single-stranded derivative from Agrobacterium to the plant cell nucleus. This conclusion is drawn from experiments exploiting the different properties of single- and double-stranded DNA to perform extrachromosomal homologous recombination in plant cells. After transfer from Agrobacterium to plant cells, T-DNA molecules recombined much more efficiently if the homologous sequences were of opposite polarity than if they were of the same polarity. This observation reflects the properties of single-stranded DNA; single-stranded DNA molecules of opposite polarity can anneal directly, whereas single-stranded DNA molecules of the same polarity first have to become double stranded to anneal. Judging from the relative amounts of single- to double-stranded T-DNA derivatives undergoing recombination, we infer that the T-DNA derivatives enter the plant nucleus in their single-stranded form.