Stabilizing ER Ca2+ Channel Function as an Early Preventative Strategy for Alzheimer’s Disease

Alzheimer’s disease (AD) is a devastating neurodegenerative condition with no known cure. While current therapies target late-stage amyloid formation and cholinergic tone, to date, these strategies have proven ineffective at preventing disease progression. The reasons for this may be varied, and could reflect late intervention, or, that earlier pathogenic mechanisms have been overlooked and permitted to accelerate the disease process. One such example would include synaptic pathology, the disease component strongly associated with cognitive impairment. Dysregulated Ca²⁺ homeostasis may be one of the critical factors driving synaptic dysfunction. One of the earliest pathophysiological indicators in mutant presenilin (PS) AD mice is increased intracellular Ca²⁺ signaling, predominantly through the ER-localized inositol triphosphate (IP₃) and ryanodine receptors (RyR). In particular, the RyR-mediated Ca²⁺ upregulation within synaptic compartments is associated with altered synaptic homeostasis and network depression at early (presymptomatic) AD stages. Here, we offer an alternative approach to AD therapeutics by stabilizing early pathogenic mechanisms associated with synaptic abnormalities. We targeted the RyR as a means to prevent disease progression, and sub-chronically treated AD mouse models (4-weeks) with a novel formulation of the RyR inhibitor, dantrolene. Using 2-photon Ca²⁺ imaging and patch clamp recordings, we demonstrate that dantrolene treatment fully normalizes ER Ca²⁺ signaling within somatic and dendritic compartments in early and later-stage AD mice in hippocampal slices. Additionally, the elevated RyR2 levels in AD mice are restored to control levels with dantrolene treatment, as are synaptic transmission and synaptic plasticity. Aβ deposition within the cortex and hippocampus is also reduced in dantrolene-treated AD mice. In this study, we highlight the pivotal role of Ca²⁺ aberrations in AD, and propose a novel strategy to preserve synaptic function, and thereby cognitive function, in early AD patients.     

Towards interbreed IBD fine mapping of the mh locus: Double-muscling in the Asturiana de los Valles breed involves the same locus as in the Belgian Blue cattle breed

The Spanish ``Asturiana' cattle breed is characterized by the segregation of a genetically determined muscular hypertrophy referred to as double-muscling or ``culones'. We demonstrate by linkage analysis that this muscular hypertrophy involves the mh locus previously shown to cause double-muscling in the Belgian Blue cattle breed, pointing towards locus homogeneity of this trait across both breeds. Moreover, using a twopoint and multipoint maximum likelihood approach, we show that flanking microsatellite markers are in linkage disequilibrium with the mh locus in both breeds albeit with different alleles. Finally, we discuss how allelic homogeneity across breeds might be exploited to achieve efficient genetic fine-mapping of the mh locus. Received: 13 September 1996 / Accepted: 20 January 1997     

The crystal structure of the beta-lactamase of Streptomyces albus G at 0.3 nm resolution.

The crystal structure of the beta-lactamase of Streptomyces albus G has been solved at 0.3 nm resolution by X-ray-diffraction methods. The enzyme is a typical two-domain protein. One domain consists of five alpha-helices, and the other is five-stranded beta-sheet with alpha-helices on both sides of the sheet. The active-site serine residue (Ser-48) is within a cleft located between the two domains.     

Lysyl-tRNA synthetase from Escherichia coli K12. Chromatographic heterogeneity and the lysU-gene product.

In contrast with most aminoacyl-tRNA synthetases, the lysyl-tRNA synthetase of Escherichia coli is coded for by two genes, the normal lysS gene and the inducible lysU gene. During its purification from E. coli K12, lysyl-tRNA synthetase was monitored by its aminoacylation and adenosine(5')tetraphospho(5')adenosine (Ap4A) synthesis activities. Ap4A synthesis was measured by a new assay using DEAE-cellulose filters. The heterogeneity of lysyl-tRNA synthetase (LysRS) was revealed on hydroxyapatite; we focused on the first peak, LysRS1, because of its higher Ap4A/lysyl-tRNA activity ratio at that stage. Additional differences between LysRS1 and LysRS2 (major peak on hydroxyapatite) were collected. LysRS1 was eluted from phosphocellulose in the presence of the substrates, whereas LysRS2 was not. Phosphocellulose chromatography was used to show the increase of LysRS1 in cells submitted to heat shock. Also, the Mg2+ optimum in the Ap4A-synthesis reaction is much higher for LysRS1. LysRS1 showed a higher thermostability, which was specifically enhanced by Zn2+. These results in vivo and in vitro strongly suggest that LysRS1 is the heat-inducible lysU-gene product.     

A locus involved in kanamycin, chloramphenicol and L-serine resistance is located in the bglY-galU region of the Escherichia coli K12 chromosome

Summary Spontaneous mutants of Escherichia coli K12 displaying an increased level of the kanamycin resistance conferred by plasmid pGR71 were selected. Several mutants obtained in this way apparently carry large chromosomal deletions extending into galU and/or bglY (27 min). This positive selection of deletions allowed detection of a new locus located between galU and bglY. Deletions of this locus are responsible for increased resistance to kanamycin (Irk), decreased resistance to l-serine in minimal medium (Drs) and decreased resistance to chloramphenicol (Drc) when a cat gene is present in the bacteria.     

Limitation of DNA-4',6-diamidine-2-phenylindole assay in the presence of an excess of tRNA

4',6-Diamidino-2-phenylindole, DAPI, forms fluorescent complexes with DNA. This property has been used to quantify DNA on the basis of fluorometric test. However, the fluorescence quantum yield of DAPI increases also with tRNA. DNA estimation needs particular care in the presence of tRNA. For DNA containing 50% adenine-thymine (AT), DAPI can be used if DNA represents at least 3.4% of the total nucleotide material. This percentage varies with the AT/guanosine-cytosine content. When the fraction of DNA decreases further, the DAPI assay can no longer be used.     

Intrinsic potential for high fetal hemoglobin production in a Druze family with β-thalassemia is due to an unlinked genetic determinant

Summary The mechanism for elevated production of fetal hemoglobin (Hb F) in a Druze patient with °-thalassemia intermedia was investigated. Heterozygous family members exhibited normal Hb F levels, suggesting that the increase in -gene expression in the propositus may be partly due to anemic stress. Erythroid progenitors of these family members cultured in vitro [burst forming units (erythroid); (BFUe)] showed elevated synthesis of Hb F, indicating the existence of a genetically determined intrinsic capacity for high Hb F production in this family. The propositus was found to be homozygous for a IVS2-position 1 mutation, on the background of Mediterranean haplotype I, which is not known to be linked to high Hb F production. Moreover, extensive molecular studies of the -globin gene cluster, including sequence analysis of the promoter regions of the -globin genes, did not reveal any cisacting mechanism that could account for the high Hb F production in the propositus. A young niece of the propositus with °-thalassemia major was recently discovered, who was homozygous for the same -globin allele and haplotype as the propositus. However, unlike her uncle, she does not have a high Hb F level and presents with a severe clinical course. Her inability to produce high Hb F suggests that the genetic determinant for increased -gene expression in the propositus is unlinked to the -globin gene cluster.     

N2-Succinylornithine in ornithine catabolism of Pseudomonas aeruginosa

Most Pseudomonas aeruginosa PAO mutants which were unable to utilize l-arginine as the sole carbon and nitrogen source (aru mutants) under aerobic conditions were also affected in l-ornithine utilization. These aru mutants were impaired in one or several enzymes involved in the conversion of N²-succinylornithine to glutamate and succinate, indicating that the latter steps of the arginine succinyltransferase pathway can be used for ornithine catabolism. Addition of aminooxyacetate, an inhibitor of the N²-succinylornithine 5-aminotransferase, to resting cells of P. aeruginosa in ornithine medium led to the accumulation of N²-succinylornithine. In crude extracts of P. aeruginosa an ornithine succinyltransferase (l-ornithine:succinyl-CoA N²-succinyltransferase) activity could be detected. An aru mutant having reduced arginine succinyltransferase activity also had correspondingly low levels of ornithine succinyltransferase. Thus, in P. aeruginosa, these two activities might be due to the same enzyme, which initiates aerobic arginine and ornithine catabolism.Abbreviations OAT ornithine 5-aminotransferase - SOAT N²-succinylornithine 5-aminotransferase - Oru ornithine utilization - Aru arginine utilization     

Co-expression of the proto-oncogene fos (c-fos) and an embryonic interferon (ovine trophoblastin) by sheep conceptuses during implantation.

Expression of the c-fos proto-oncogene by ovine conceptuses was analyzed by Northern and slot blots and indirect immunohistofluorescence in relation to the expression of the embryonic interferon-alpha (oTP) during implantation. c-fos was expressed initially in the trophoblast, and then in the allantois, when this tissue began to develop (day 17). In the embryonic tissues, the c-fos proto-oncogene was weakly expressed up to day 22 and increased thereafter. In the trophoblast, the expression of c-fos proto-oncogene was transient, occurring when the oTP gene was transcribed at a maximal level at the beginning of implantation (days 14-15), and decreased thereafter, following the pattern of oTP gene expression. This decline is due essentially to the arrest of c-fos and oTP gene expression by the trophoblastic cells which established cellular contacts with the uterine epithelium during the implantation process.     

Inhibition of chlorophyll synthesis inHordeum vulgare by 3-amino 2,3-dihydrobenzoic acid (gabaculin)

Gabaculin (3-amino 2,3-dihydrobenzoic acid) is shown to be a very potent inhibitor of chlorophyll formation inHordeum vulgate. Exposure of leaf segments to 30/M gabaculin results in an 80% inhibition of chlorophyll synthesis, and this is paralleled by a decrease in carotenoid. Dual-inhibitor studies with dioxoheptanoic acid, which is an inhibitor of inolaevulinic acid dehydratase, show that gabaculin inhibits an earlier step than dioxoheptanoic acid and affects -aminolaevulinic acid synthesis rather than its subsequent metabolism.