Mouse glandular kallikrein genes. Structure and partial sequence analysis of the kallikrein gene locus

Mouse glandular kallikreins are encoded by a family of closely linked genes which are located on chromosome 7 at a site corresponding to the genetically defined Tam-1, Prt-4, and Prt-5 loci. We have characterized 24 kallikrein genes by genomic cloning and restriction mapping of 310 kilobase pairs of BALB/c mouse DNA. Most of these genes are highly homologous, have the same exon/intron organization, and are linked in clusters of up to 11 genes. Partial sequence analysis of the kallikrein genes has facilitated identification of those members of the family for which protein sequence data exist and assignment of those which are pseudogenes or encode proteins of unknown function. We find that a maximum of 14 mouse kallikrein genes have the potential to encode functional proteins.     

Interaction among hepatocyte-stimulating factors, interleukin 1, and glucocorticoids for regulation of acute phase plasma proteins in human hepatoma (HepG2) cells

Human hepatoma (HepG2) cells respond to unfractionated conditioned media of human squamous carcinoma (COLO-16) cells and lipopolysaccharide-stimulated human peripheral blood monocytes by increasing the synthesis of alpha 1-acid glycoprotein, haptoglobin, complement C3, alpha 1-antichymotrypsin, alpha 1-antitrypsin, and fibrinogen, while decreasing the synthesis of albumin. The regulation of the acute phase proteins is mediated by hepatocyte-stimulating factors (HSF) and interleukin 1 (IL-1) present in the conditioned medium. Purified HSF-I from COLO-16 cells stimulates preferentially alpha 1-acid glycoprotein synthesis, whereas COLO-HSF-II stimulates preferentially the synthesis of haptoglobin, fibrinogen, and alpha 1-antitrypsin. HSF from monocytes, which has been identified as interferon-beta 2 (B cell stimulating factor-2), displayed the same activity as COLO-HSF-II. Dexamethasone alone had no effect on acute phase plasma protein synthesis but enhanced the response to various HSF severalfold. IL-1 had a relatively low stimulatory activity on the synthesis of alpha 1-acid glycoprotein, haptoglobin, and alpha 1-antichymotrypsin but strongly reduced the basal expression of fibrinogen. The only synergistic action between IL-1 and HSF (or interferon-beta 2) was noted for the synthesis of alpha 1-acid glycoprotein. Tumor necrosis factor active on other hepatic cells failed to modulate significantly the expression of any plasma proteins in HepG2 cells. These studies showed that for an optimal HepG2-cell response a combination of HSF (or interferon-beta 2), IL-1, and dexamethasone is needed. This finding might indicate the identity of some of those hormones involved in regulation of the hepatic acute phase response in vivo.     

Estimating the number of genetic elements that defer senescence inDrosophila

Summary Although many different physiological and biochemical changes characterize the process of senescence, little is understood of the genetic elements that determine its age of onset. We provide here the first estimates of the number of genetic factors that extend longevity inDrosophila melanogaster. Life span was measured in F₁, F₂ and backcrosses of true-breeding long and short-lived stocks ofD. melanogaster, established by selection. Estimates of the number of effective factors delaying senescence range from about 0.3 to 1.5, indicating control by a single factor. The distribution of longevity shows this to arise as selection acts on the short-lived parental stock. Life span is extended at the cost of early fecundity.     

Lipids in the laminated layer of liver, lung and daughter hydatid cysts of equine Echinococcus granulosus (Cestoda)

Lipids extracted from the laminated layers of horse liver and lung hydatids, including a daughter liver cyst, were analysed using TLC. No differences in lipid composition was detected in 11 liver cysts, whether from the same or different livers, and di- and triacylglycerols, cholesterol, wax and steryl esters, oleic acid, sphingomyelin, phosphatidyl choline, phosphatidyl inositol and ceramide hexosides were detected. The daughter cyst differed from its "parent" cyst in lacking diacylglycerols and wax and steryl esters. The lung cyst differed from the liver cysts in that cholesterol, wax and steryl esters and diacylglycerols were not detected.     

A proton nuclear magnetic resonance assignment and secondary structure determination of recombinant human thioredoxin

Two-dimensional 1H NMR spectroscopy has been applied to a structural analysis of the reduced form of a recombinant human thioredoxin, a ubiquitous dithiol oxidoreductase recently isolated from an immunocompetent lymphoblastoid cell line. The sequential assignment of the spectrum, including all proline residues, has been accomplished by using experiments to demonstrate through-bond and through-space connectivities. The secondary structure has been determined by a qualitative interpretation of nuclear Overhauser effects, NH exchange data, and 3JHN alpha coupling constants. The secondary structure was found to be similar to that of the X-ray structure of Escherichia coli thioredoxin, consisting of a mixed five-stranded beta-sheet surrounded by four alpha-helices. The assignment and structural characterization of human thioredoxin was facilitated by the increased resolution and sensitivity afforded by a magnetic field strength of 600 MHz and required the use of two temperatures and two pH conditions to resolve ambiguities caused by a duplication of resonances. This duplication, extending from Phe-41 to Val-59, and including Lys-3-Ile-5, Val-24, Val-25, Asn-39, and Ile-101-Glu-103, appears to be due to heterogeneity arising from the presence or absence of the N-terminal methionine.     

Theoretical calculation of tautomer equilibria in solution: 4-(5-)methylimidazole

A combination of quantum mechanical calculations and molecular dynamics simulations has been used to calculate the tautomer ratio of 4-(5-)methyl imidazole in solution, and the results are in good agreement with experiment.     

GABA A -receptors: Structural requirements and sites of gene expression in mammalian brain

GABA_A-receptors, the major synaptic targets for the neutotransmitter GABA, are gated chloride channels. By their allosteric drug-induced modulation they serve as molecular control elements through which the levels of anxiety, vigilance, muscle tension and epileptiform activity can be regulated. Despite their functional prominence, the structural requirements of fully functional GABA_A-receptors are still elusive. Expression of cDNAs coding for the ₁- and ₁-subunits of rat brain yielded GABA-gated chloride channels which were modulated by barbiturates but displayed only agonistic responses to ligands of the benzodiazepine receptor. GABA_A-receptors with fully functional benzodiazepine receptor sites were formed when the ₁- and ₁-subunits were coexpressed with the ₂-subunit of rat brain. These receptors, however, failed to show cooperativity of GABA in gating the channel. In order to determine the subunit repertoire available for receptor assembly in different neuronal populations in vivo, the sites of subunit gene expression were (₁, ₂, ₃, ₅, ₆, ₁, ₂, ₃, ₂) mapped by in situ hybridization histochemistry in brain sections. The mRNAs of the ₁-, ₁- and ₂-subunits were co-localized e.g. in mitral cells of olfactory bulb, pyramidal cells of hippocampus as well as granule cells of dentate gyrus and cerebellum. The lack of colocalization in various other brain areas points to an extensive receptor heterogeneity. The presence of multiple GABA_A-receptors in brain may contribute to synaptic plasticity, differential responsiveness of neurons to GABA and to variations in drug profiles.Special issue dedicated to Dr. Erminio Costa