Protoveratrines A and B increase sleep apnea index in Sprague-Dawley rats

Trbovic, Sinisa M., Miodrag Radulovacki, and David W. Carley. Protoveratrines A and B increase sleep apneaindex in Sprague-Dawley rats. J. Appl.Physiol. 83(5): 1602-1606, 1997.The action ofprotovertarines A and B, which stimulate carotid sinus baroreceptorsand vagal sensory endings in the heart as well as pulmonary bed, wereassessed on spontaneous and postsigh central sleep apneas in freelymoving Sprague-Dawley rats. During the 6-h recording period, animalswere simultaneously monitored for sleep by using electroencephalogramand electromyogram recordings, for respiration by single-chamberplethysmography, and for blood pressure and heart period by usingradiotelemetry. After administration of 0.2, 0.5, or 1 mg/kg sc ofprotoveratrines, cardiopulmonary changes lasting at least 6 h wereobserved in all three behavioral states [heart period increasedup to 23% in wakefulness, 21% in non-rapid-eye-movement (non-REM)sleep, and 20% in REM sleep; P < 0.005 for each]. At the same time, there was a substantial increase in the number of spontaneous (375% increase;P = 0.04) and postsigh (268%increase, P = 0.0002) apneas. Minuteventilation decreased by up to 24% in wakefulness, 25% in non-REM,and 35% in REM sleep (P < 0.05 foreach). We conclude that pharmacological stimulation of baroreflexespromotes apnea expression in the sleeping rat.

     

Human Ikaros function in activated T cells is regulated by coordinated expression of its largest isoforms

The Ikaros gene is alternately spliced to generate multiple zinc finger proteins involved in gene regulation and chromatin remodeling. Whereas murine studies have provided important information regarding the role of Ikaros in the mouse, little is known of Ikaros function in human. We report functional analyses of the two largest human Ikaros (hIK) isoforms, hIK-VI and hIK-H, in T cells. Abundant expression of hIK-H, the largest described isoform, is restricted to human hematopoietic cells. We find that the DNA binding affinity of hIK-H differs from that of hIK-VI. Co-expression of hIk-H with hIk-VI alters the ability of Ikaros complexes to bind DNA motifs found in pericentromeric heterochromatin (PC-HC). In the nucleus, hIK-VI is localized solely in PC-HC, whereas the hIK-H protein exhibits dual centromeric and non-centromeric localization. Mutational analysis defined the amino acids responsible for the distinct DNA binding ability of hIK-H, as well as the sequence required for the specific subcellular localization of this isoform. In proliferating cells, the binding of hIK-H to the upstream regulatory region of known Ikaros target genes correlates with their positive regulation by Ikaros. Results suggest that expression of hIK-H protein restricts affinity of Ikaros protein complexes toward specific PC-HC repeats. We propose a model, whereby the binding of hIK-H-deficient Ikaros complexes to the regulatory sequence of target genes would recruit these genes to the restrictive pericentromeric compartment, resulting in their repression. The presence of hIK-H in the Ikaros complex would alter its affinity for PC-HC, leading to chromatin remodeling and activation of target genes.     

Food For Thought: Short-Term Fasting Upregulates Glucose Transporters in Neurons and Endothelial Cells,But Not in Astrocytes

Neurochemical Research - Our group previously reported that 6-h fasting increased both insulin II mRNA expression and insulin level in rat hypothalamus. Given that insulin effects on central...     

Involuntary hospitalizations in the psychiatric hospital Jankomir before and following the alterations and amendments made to ZZODS

Schizophrenia and other psychotic disorders as well as the delirium caused by abstinence from alcohol and accute state of drunkenness appear at the very top of the list of factors, which are positively correlated with involuntary hospitalization of patients. This is at the same time a confirmation of the data found in literature considering psychosis an essential factor of involuntary hospitalization; the same referring to the male sex was not, however, confirmed by the results obtained in the first and second research period. Regarding the positive correlation between schizophrenia and other psychotic disturbances, dementia, delirium and other cognitive impairments including the delirium caused by abstinence from alcohol and an accute state of drunkenness on the one side and the high rate of involuntary hospitalization on the other, there is no statistically significant difference between the period preceding and the period following the alterations and amendments to the Law on the protection of patients with mental disorders.     

Diverse substrate recognition mechanisms for rhomboids; thrombomodulin is cleaved by Mammalian rhomboids

The rhomboids are a recently discovered family of intramembrane proteases that are conserved across evolution. Drosophila was the first organism in which they were characterized, where at least Rhomboids 1-3 activate EGF receptor signaling by releasing the active forms of EGF-like growth factors. Subsequent work has begun to shed light on the role of these proteases in bacteria and yeast, but nothing is known about the function of rhomboids in vertebrates beyond evidence that the subclass of mitochondrial rhomboids is conserved. Here, we report that the anticoagulant cell-surface protein thrombomodulin is the first mammalian protein to be a rhomboid substrate in a cell culture assay. The thrombomodulin transmembrane domain (TMD) is cleaved only by vertebrate RHBDL2-like rhomboids. Thrombomodulin TMD cleavage is directed not by sequences within the TMD, as is the case with Spitz but by its cytoplasmic domain, which, at least in some contexts, is necessary and sufficient to determine cleavage by RHBDL2. These data suggest that thrombomodulin could be a physiological substrate for rhomboid. Moreover, the discovery of a second mode of substrate recognition by rhomboids implies mechanistic diversity in this family of intramembrane proteases.     

Conservation of intramembrane proteolytic activity and substrate specificity in prokaryotic and eukaryotic rhomboids

Rhomboid is an intramembrane serine protease responsible for the proteolytic activation of Drosophila epidermal growth factor receptor (EGFR) ligands. Although nothing is known about the function of the approximately 100 currently known rhomboid genes conserved throughout evolution, a recent analysis suggests that a Rhomboid from the pathogenic bacterium Providencia stuartii is involved in the production of a quorum-sensing factor. This suggests that an intercellular signaling mechanism may have been conserved between prokaryotes and metazoans. However, the function of prokaryotic Rhomboids is unknown. We have examined the ability of eight prokaryotic Rhomboids to cleave the three Drosophila EGFR ligands. Despite their striking sequence divergence, Rhomboids from one Gram-positive and four Gram-negative species, including Providencia, specifically cleaved Drosophila substrates, but not similar proteins such as Transforming Growth Factor alpha (TGFalpha) and Delta. Although the sequence similarity between these divergent Rhomboids is very limited, all contain the putative serine catalytic triad residues, and their specific mutation abolished protease activity. Therefore, despite low overall homology, the Rhomboids are a family of ancient, functionally conserved intramembrane serine proteases, some of which also have conserved substrate specificity. Moreover, a function for Rhomboids in activating intercellular signaling appears to have evolved early.     

Variants in Nebulin (NEB) Are Linked to the Development of Familial Primary Angle Closure Glaucoma in Basset Hounds

Several dog breeds are susceptible to developing primary angle closure glaucoma (PACG), which suggests a genetic basis for the disease. We have identified a four-generation Basset Hound pedigree with characteristic autosomal recessive PACG that closely recapitulates PACG in humans. Our aim is to utilize gene mapping and whole exome sequencing approaches to identify PACG-causing sequence variants in the Basset. Extensive clinical phenotyping of all pedigree members was conducted. SNP-chip genotyping was carried out in 9 affected and 15 unaffected pedigree members. Two-point and multipoint linkage analyses of genome-wide SNP data were performed using Superlink-Online SNP-1.1 and a locus was mapped to chromosome 19q with a maximum LOD score of 3.24. The locus contains 12 Ensemble predicted canine genes and is syntenic to a region on chromosome 2 in the human genome. Using exome-sequencing analysis, a possibly damaging, non-synonymous variant in the gene Nebulin (NEB) was found to segregate with PACG which alters a phylogenetically conserved Lysine residue. The association of this variants with PACG was confirmed in a secondary cohort of unrelated Basset Hounds (p = 3.4 × 10^-4, OR = 15.3 for homozygosity). Nebulin, a protein that promotes the contractile function of sarcomeres, was found to be prominently expressed in the ciliary muscles of the anterior segment. Our findings may provide insight into the molecular mechanisms that underlie PACG. The phenotypic similarities of disease presentation in dogs and humans may enable the translation of findings made in this study to patients with PACG.     

The effect of acute heat exposure on rat pituitary corticotroph activation: the role of vasopressin

The increased ambient temperature affects the function of hypothalamic-pituitary-adrenal (HPA) axis. Since the correlation among vasopressin (VP), adrenocorticotropic hormone (ACTH) and corticosterone (CORT) responses to various stressors have been long recognized, the aim of this study was to reveal the aforementioned hormones production and morphology of the pituitary gland after exposure to acute heat. Rats were exposed to high ambient temperature (38 °C) for 20 or 60 minutes. The circulating hormones were determined by an ELISA test or chemiluminescence's method. The results obtained show the elevation in ACTH and CORT secretion depending on the duration of heat exposure. The VP concentration increased only after prolonged exposure to heat (60 min). The pituitary morphology was examined by routine and fluorescent immunohistochemistry as well as electron microscopy. Observed changes in the anterior and posterior pituitary well corresponded to circulating hormones, regarding the volume density of ACTH-immunopositive cells, percentage of ACTH immunopositive area v. total area and number of VP-immunopositive containing varicose fibers per total area. Acute heat exposure also induced changes in shapes of ACTH-immunopositive cells. Cells appeared stellate with numerous slender cytoplasmic processes and degranulated, which is the most obvious after 20 min. In addition, immunopositivity of endothelial and anterior pituitary cells for VP suggests its influence on ACTH secretion.     

Widespread occurrence and genomic context of unusually small polyketide synthase genes in microbial consortia associated with marine sponges

Numerous marine sponges harbor enormous amounts of as-yet-uncultivated bacteria in their tissues. There is increasing evidence that these symbionts play an important role in the synthesis of protective metabolites, many of which are of great pharmacological interest. In this study, genes for the biosynthesis of polyketides, one of the most important classes of bioactive natural products, were systematically investigated in 20 demosponge species from different oceans. Unexpectedly, the sponge metagenomes were dominated by a ubiquitously present, evolutionarily distinct, and highly sponge-specific group of polyketide synthases (PKSs). Open reading frames resembling animal fatty acid genes were found on three corresponding DNA regions isolated from the metagenomes of Theonella swinhoei and Aplysina aerophoba. Their architecture suggests that methyl-branched fatty acids are the metabolic product. According to a phylogenetic analysis of housekeeping genes, at least one of the PKSs belongs to a bacterium of the Deinococcus-Thermus phylum. The results provide new insights into the chemistry of sponge symbionts and allow inference of a detailed phylogeny of the diverse functional PKS types present in sponge metagenomes. Based on these qualitative and quantitative data, we propose a significantly simplified strategy for the targeted isolation of biomedically relevant PKS genes from complex sponge-symbiont associations.     

De novo enzyme design using Rosetta3

The Rosetta de novo enzyme design protocol has been used to design enzyme catalysts for a variety of chemical reactions, and in principle can be applied to any arbitrary chemical reaction of interest. The process has four stages: 1) choice of a catalytic mechanism and corresponding minimal model active site, 2) identification of sites in a set of scaffold proteins where this minimal active site can be realized, 3) optimization of the identities of the surrounding residues for stabilizing interactions with the transition state and primary catalytic residues, and 4) evaluation and ranking the resulting designed sequences. Stages two through four of this process can be carried out with the Rosetta package, while stage one needs to be done externally. Here, we demonstrate how to carry out the Rosetta enzyme design protocol from start to end in detail using for illustration the triosephosphate isomerase reaction.