The C-Terminal Domain of the mGluR1 Metabotropic Glutamate Receptor Affects Sensitivity to Agonists

Abstract: The metabotropic glutamate receptor (mGluR) subtype 1 exists as at least three variants (−1a, −1b, and −1c) generated by alternative splicing at the C-terminal domain. Fluorometric Ca²⁺ measurements were used to compare the concentration dependency of agonist-induced rises in intracellular free Ca²⁺ concentration ([Ca²⁺]_i) in human embryonic HEK 293 cells transiently expressing rat mGluR1a, mGluR1b, or mGluR1c. The rank order of agonist potencies was quisqualate ≫ (2 S, 1' S, 2' S )-2-(carboxycyclopropyl)glycine (L-CCG-I) > (1 S, 3 R )-1-aminocyclopentane-1,3-dicarboxylic acid [(1 S, 3 R )-ACPD] and did not differ among the splice variants. However, agonists were consistently more potent at mGluR1a than at mGluR1c and mGluR1b. In the same system, we characterized the agonist pharmacology of two chimeric rat mGluR3/1 receptors where the first and/or the second intracellular loop(s) and the C-terminal domain were exchanged with the corresponding mGluR1a or mGluR1c sequences and that were previously shown to mediate elevations in [Ca²⁺]_i in response to agonists. The potency of agonists was higher at the chimera having the C-terminus of mGluR1a as compared with those having the mGluR1c C-terminus. Both chimeric mGluR3/1 receptors had the same rank order of agonist potencies: L-CCG-I ≫ (1 S, 3 R )-ACPD ∼ quisqualate. These data support the hypothesis that the C-terminal domain of mGluRs plays a role in determining the potency of agonists for inducing mGluR-mediated functional responses.     

A heuristic algorithm for the loading problem in flexible manufacturing systems

The paper considers the loading problem in flexible manufacturing systems (FMSs). This problem involves the assignment to the machine tools of all operations and associated cutting tools required for part types that have been selected to be produced simultaneously. The loading problem is first formulated as a linear mixed 0–1 program with the objective to minimize the greatest workload assigned to each machine. A heuristic procedure is presented in which an assignment of operations to machine tools is obtained by solving a parameterized generalized assignment problem with an objective function that approximates the use of tool slots required by the operations assigned to the machines. The algorithm is coded in FORTRAN and tested on an IBM-compatible personal computer. Computational results are presented for different test problems to demonstrate the efficiency and effectiveness of the suggested procedure.     

Interaction of poly(ADP-ribose)polymerase with DNA polymerase α

Homogeneously purified poly(ADP-ribose) polymerase (PARP) specifically stimulated the activity of immunoaffinity-purified calf or human DNA polymerase by about 6 to 60-fold. Apparently, poly(ADP-ribosyl)ation of DNA polymerase was not necessary for the stimulation. The effects of PARP on DNA polymerase were biphasic: at very low concentrations of DNA, it rather inhibited its activity, whereas, at higher DNA concentrations, PARP greatly stimulated it. The autopoly(ADP-ribosyl)ation of PARP suppressed both its stimulatory and inhibitory effects. By immunoprecipitation with an anti-DNA polymerase antibody, it was clearly shown that PARP may be physically associated with DNA polymerase . Stimulation of DNA polymerase may be attributed to the physical association between the two, rather than to the DNA-binding capacity of PARP, since the PARP fragment containing only the DNA binding domain showed little stimulatory activity. The existence of PARP-DNA polymerase complexes were also detected in crude extracts of calf thymus.     

DNA binding factor GT-2 from Arabidopsis

Complementary DNA clones encoding a DNA-binding factor have been obtained from Arabidopsis by DNA hybridization with a GT-2 factor cDNA clone from rice. The GT-2 gene appears to be present as a single copy in the Arabidopsis genome and is transcribed as a 2.1 kb mRNA which is not light-regulated. The longest open reading frame in the sequenced clones predicts a protein of 65 kDa, beginning with the first in-frame methionine. The protein contains basic, acidic, and proline/glutamine-rich motifs and has significant amino acid sequence homology to the rice GT-2 factor, including three regions of 50–75 amino acids each of greater than 60% identity. Two of these regions are predicted to form similar trihelix structures postulated to be involved in selective binding to specific variations of a GT-box motif DNA sequence found in the promoter regions of several plant genes. Except for weak similarity to a tobacco GT-box binding factor, GT-1a/B2F, Arabidopsis GT-2 has no similarity to other sequences in the databases. DNA-binding studies show that Arabidopsis GT-2 has binding characteristics similar to those of the rice GT-2 factor, but dissimilar to those of the tobacco GT-1a/B2F factor. The data indicate that a DNA-binding factor containing domains of similar structure and target-sequence specificity has been conserved between monocots and dicots.     

Cloning and characterization of the gene encoding an esterase from Spirulina platensis

The gene encoding a 23 kDA serine esterase from the cyanobacterium Spirulina platensis has been identified, cloned, characterized and expressed in Escherichia coli. The primary structure of the esterase deduced from the DNA sequence displayed 32% sequence identity with the carboxylesterase (esterase II) encoded by estB of Pseudomonas fluorescens; the highest degree of homology is found in a stretch of 11 identical or highly conserved amino acid residues corresponding to the GXSXG consensus motif found in the catalytic site of many serine proteases, lipases and esterases.     

Widespread formation of intracellular calcium carbonates by the bloom-forming cyanobacterium Microcystis

The formation of intracellular amorphous calcium carbonates (iACC) has been recently observed in a few cultured strains of Microcystis, a potentially toxic bloom-forming cyanobacterium found worldwide in freshwater ecosystems. If iACC-forming Microcystis are abundant within blooms, they may represent a significant amount of particulate Ca. Here, we investigate the significance of iACC biomineralization by Microcystis. First, the presence of iACC-forming Microcystis cells has been detected in several eutrophic lakes, indicating that this phenomenon occurs under environmental conditions. Second, some genotypic (presence/absence of ccyA, a marker gene of iACC biomineralization) and phenotypic (presence/absence of iACC) diversity have been detected within a collection of strains isolated from one single lake. This illustrates that this trait is frequent but also variable within Microcystis even at a single locality. Finally, one-third of publicly available genomes of Microcystis were shown to contain the ccyA gene, revealing a wide geographic and phylogenetic distribution within the genus. Overall, the present work shows that the formation of iACC by Microcystis is common under environmental conditions. While its biological function remains undetermined, this process should be further considered regarding the biology of Microcystis and implications on the Ca geochemical cycle in freshwater environments.     

The Glucose-Related Decrease in Polar Auxin Transport During Ripening and its Possible Role in Grapevine Berry Coloring

Journal of Plant Growth Regulation - Auxin is a hormone that delays ripening in part by reducing anthocyanin content and impairing color development. Auxin content declines during the ripening...     

COMPARATIVE ONTOGENY OF A WILD CUCURBIT AND ITS DERIVED CULTIVAR

Most previous studies of evolutionary modification of form in plants have focused primarily on individual organs or flowers. Few have investigated the role of evolutionary changes in timing or position at the level of whole plant ontogeny. This study compares ontogenies of the primary shoots of two subspecies of Cucurbita argyrosperma, one a cultivar and the other its wild progenitor. Differences in flowering times between these subspecies suggested that the cultivar may have evolved from the wild subspecies via heterochronic processes leading to paedomorphosis. Analyses showed that both subspecies are similar in vegetative architecture and rates of leaf production. Earlier flowering in the cultivar, both in terms of position and absolute time, appears to have arisen through progenesis. Initial observations of leaf blade morphology led to the hypothesis that paedomorphosis and gigantism also may have been involved in the evolution of leaf blade shape in the cultivar: all leaves of the cultivar are larger and visually similar in shape to early leaves of the wild subspecies. However, quantitative analysis revealed that leaves of the cultivar are neither geometrically, nor solely allometrically larger versions of early leaves of the progenitor. Leaf shape in the cultivar exhibits novel features as well as effects of allometry shared with the progenitor, hence a simple hypothesis of paedomorphic evolution of leaf shape is not supported.     

Solid-phase synthesis of triple-helical collagen-model peptides

Summary The triple-helical conformation of collagen has been proposed to be important for mediation of cellular activities, such as adhesion and activation, extracellular matrix assembly, and enzyme function. We have developed synthetic protocols that allow for the study of biological activities of specific collagen sequences in triple-helical conformation. These methods primarily involve solid-phase assembly and covalent linkage of three peptide chains. The resultant triple-helical peptides have sufficient thermal stabilities to permit structural and biological characterization under physiological conditions. The present article critically reviews the various approaches for constructing synthetic triple-helices.This paper is based on a presentation given at the Symposium on Peptide Structure and Design as part of the 31st Annual ACS Western Regional Meeting held in San Diego, CA, USA, October 18–21, 1995.