Modulation of an early step in the secretory machinery in hippocampal nerve terminals

In hippocampal neurons, neurotransmitter release can be regulated by protein kinase A (PKA) through a direct action on the secretory machinery. To identify the site of PKA modulation, we have taken advantage of the ability of the neurotoxin Botulinum A to cleave the synaptic protein SNAP-25. Cleavage of this protein decreases the Ca2+ responsiveness of the secretory machinery by partially uncoupling Ca2+-sensing from fusion per se. This is expressed as a shift toward higher Ca2+ levels of the Ca2+ to neurotransmitter release relationship and as a perturbation of synaptic delay under conditions where secretion induced by the Ca2+-independent secretagogue ruthenium red is unimpaired. We find that SNAP-25 cleavage also perturbs PKA-dependent modulation of secretion; facilitation of ruthenium red-evoked neurotransmitter release by the adenylyl cyclase activator forskolin is blocked completely after Botulinum toxin A action. Together with our observation that forskolin modifies the Ca2+ to neurotransmitter release relationship, our results suggest that SNAP-25 acts as a functional linker between Ca2+ detection and fusion and that PKA modulates an early step in the secretory machinery related to calcium sensing to facilitate synaptic transmission.

Cyclin A activates the DNA polymerase δ-dependent elongation machinery in vitro: A parvovirus DNA replication model

Replication of the single-stranded linear DNA genome of parvovirus minute virus of mice (MVM) starts with complementary strand synthesis from the 3′-terminal snap-back telomere, which serves as a primer for the formation of double-stranded replicative form (RF) DNA. This DNA elongation reaction, designated conversion, is exclusively dependent on cellular factors. In cell extracts, we found that complementary strand synthesis was inhibited by the cyclin-dependent kinase inhibitor p21WAF1/CIP1 and rescued by the addition of proliferating cell nuclear antigen, arguing for the involvement of DNA polymerase (Pol) δ in the conversion reaction. In vivo time course analyses using synchronized MVM-infected A9 cells allowed initial detection of MVM RF DNA at the G1/S phase transition, coinciding with the onset of cyclin A expression and cyclin A-associated kinase activity. Under in vitro conditions, formation of RF DNA was efficiently supported by A9 S cell extracts, but only marginally by G1 cell extracts. Addition of recombinant cyclin A stimulated DNA conversion in G1 cell extracts, and correlated with a concomitant increase in cyclin A-associated kinase activity. Conversely, a specific antibody neutralizing cyclin A-dependent kinase activity, abolished the capacity of S cell extracts for DNA conversion. We found no evidence for the involvement of cyclin E in the regulation of the conversion reaction. We conclude that cyclin A is necessary for activation of complementary strand synthesis, which we propose as a model reaction to study the cell cycle regulation of the Pol δ-dependent elongation machinery.

Competition between a sterol biosynthetic enzyme and tRNA modification in addition to changes in the protein synthesis machinery causes altered nonsense suppression

The Saccharomyces cerevisiae Mod5 protein catalyzes isopentenylation of A to i6A on tRNAs in the nucleus, cytosol, and mitochondria. The substrate for Mod5p, dimethylallyl pyrophosphate, is also a substrate for Erg20p that catalyzes an essential step in sterol biosynthesis. Changing the distribution of Mod5p so that less Mod5p is present in the cytosol decreases i6A on cytosolic tRNAs and alters tRNA-mediated nonsense suppression. We devised a colony color/growth assay to assess tRNA-mediated nonsense suppression and used it to search for genes, which, when overexpressed, affect nonsense suppression. We identified SAL6, TEF4, and YDL219w, all of which likely affect nonsense suppression via alteration of the protein synthesis machinery. We also identified ARC1, whose product interacts with aminoacyl synthetases. Interestingly, we identified ERG20. Midwestern analysis showed that yeast cells overproducing Erg20p have reduced levels of i6A on tRNAs. Thus, Erg20p appears to affect nonsense suppression by competing with Mod5p for substrate. Identification of ERG20 reveals that yeast have a limited pool of dimethylallyl pyrophosphate. It also demonstrates that disrupting the balance between enzymes that use dimethylallyl pyrophosphate as substrate affects translation.

Delays in setting workplace standards for cancer-causing and other dangerous substances, Department of Labor, Department of Health, Education, and Welfare : report to the Congress /

Publication date stamped on cover.

The American cotton spinner and managers' and carders' guide : a practical treatise on cotton spinning : giving the dimensions and speed of machinery, draught and twist calculations, etc., with notices of recent improvements : together with rules and examples for making changes in the size and numbers of roving and yarn /

Cover title: American cotton-spinner.

Sulfur and its importance in food production, conclusions and recommendations of the Special Subcommittee on Fertilizer and Farm Machinery of the Committee on Agriculture, House of Representatives, based on hearings before the subcommittee February 20-23, 1951.

Mode of access: Internet.

Directory of manufacturers of and dealers in engineers and contractors machinery and supplies; a classified list of machinery and supplies manufactured by advertisers in the sixth annual contractors̓ number of the Engineering Record.

2d ed.

Radford's cyclopedia of cement construction; a general reference work on up-to-date practice in the manufacture and testing of cements; the selection of concreting materials, tools, and machinery; the proportioning, mixing, and depositing of concrete, and its application to all types and reinforced; together with analysis of the principles of constructive design, cost estimating, and the allied branches of stone and brick masonry and steel construction; based on the practical experience of a large staff of experts in actual construction work ...

"Prepared under the supervision of William A. Radford, editor-in-chief ... Alfred Sidney Johnson ... editor in charge."

Cotton ginning machinery & equipment in the United States, 1940.

Mode of access: Internet.

American politics. Political parties and party problems in the United States; a sketch of American party history and of the development and operations of party machinery, together with a consideration of certain party problems in their relations to political morality,

Contains short bibliographies.

Power for operating machinery in printing houses, a treatise on the methods of applying power to printing presses and allied machinery, with particular reference to electric drive./

Mode of access: Internet.

Workplace violence prevention strategies and research needs : report from the conference Partnering in Workplace Violence Prevention: Translating Research to Practice, November 15-17, 2004, Baltimore, Maryland.

"September 2006."

Sugar machinery; a descriptive treatise devoted to the machinery and processes used in the manufacture of cane and beet sugars,

Mode of access: Internet.