261 research outputs found
Bi-allelic variants in RNF170 are associated with hereditary spastic paraplegia.
Alterations of Ca2+ homeostasis have been implicated in a wide range of neurodegenerative diseases. Ca2+ efflux from the endoplasmic reticulum into the cytoplasm is controlled by binding of inositol 1,4,5-trisphosphate to its receptor. Activated inositol 1,4,5-trisphosphate receptors are then rapidly degraded by the endoplasmic reticulum-associated degradation pathway. Mutations in genes encoding the neuronal isoform of the inositol 1,4,5-trisphosphate receptor (ITPR1) and genes involved in inositol 1,4,5-trisphosphate receptor degradation (ERLIN1, ERLIN2) are known to cause hereditary spastic paraplegia (HSP) and cerebellar ataxia. We provide evidence that mutations in the ubiquitin E3 ligase gene RNF170, which targets inositol 1,4,5-trisphosphate receptors for degradation, are the likely cause of autosomal recessive HSP in four unrelated families and functionally evaluate the consequences of mutations in patient fibroblasts, mutant SH-SY5Y cells and by gene knockdown in zebrafish. Our findings highlight inositol 1,4,5-trisphosphate signaling as a candidate key pathway for hereditary spastic paraplegias and cerebellar ataxias and thus prioritize this pathway for therapeutic interventions
Enhanced T cell engraftment after retroviral delivery of an antiviral gene in HIV-infected individuals
Implantation Serine Proteinase 1 Exhibits Mixed Substrate Specificity that Silences Signaling via Proteinase-Activated Receptors
Implantation S1 family serine proteinases (ISPs) are tryptases involved in embryo hatching and uterine implantation in the mouse. The two different ISP proteins (ISP1 and ISP2) have been detected in both pre- and post-implantation embryo tissue. To date, native ISP obtained from uterus and blastocyst tissues has been isolated only as an active hetero-dimer that exhibits trypsin-like substrate specificity. We hypothesised that in isolation, ISP1 might have a unique substrate specificity that could relate to its role when expressed alone in individual tissues. Thus, we isolated recombinant ISP1 expressed in Pichia pastoris and evaluated its substrate specificity. Using several chromogenic substrates and serine proteinase inhibitors, we demonstrate that ISP1 exhibits trypsin-like substrate specificity, having a preference for lysine over arginine at the P1 position. Phage display peptide mimetics revealed an expanded but mixed substrate specificity of ISP1, including chymotryptic and elastase activity. Based upon targets observed using phage display, we hypothesised that ISP1 might signal to cells by cleaving and activating proteinase-activated receptors (PARs) and therefore assessed PARs 1, 2 and 4 as potential ISP1 targets. We observed that ISP1 silenced enzyme-triggered PAR signaling by receptor-disarming. This PAR-disarming action of ISP1 may be important for embryo development and implantation
Effects of passive smoking on heart rate variability, heart rate and blood pressure: an observational study
Increased immune response elicited by DNA vaccination with a synthetic gp120 sequence with optimized codon usage
DNA vaccination elicits humoral and cellular immune responses and has been shown to confer protection against several viral, bacterial, and parasitic pathogens. Here we report that optimized codon usage of an injected DNA sequence considerably increases both humoral and cellular immune responses. We recently generated a synthetic human immunodeficiency virus type 1 gp120 sequence in which most wild-type codons were replaced with codons from highly expressed human genes (syngp120). In vitro expression of syngp120 is considerably increased in comparison to that of the respective wild-type sequence. In BALB/c mice, DNA immunization with syngp120 resulted in significantly increased antibody titers and cytotoxic T-lymphocyte reactivity, suggesting a direct correlation between expression levels and the immune response. Moreover, syngp120 is characterized by rev-independent expression and a low risk of recombination with viral sequences. Thus, synthetic genes with optimized codon usage represent a novel strategy to increase the efficacy and safety of DNA vaccination
Diagnostische Bedeutung der Proteinbindung von Plasmacortisol, bestimmt durch Dextrangelfiltration
1. Mittels Dextrangelfiltration wurde nach Inkubation von markiertem Cortisol und Plasma der proteingebundene und der sog. freie Anteil (%) des endogenen Plasmacortisols ermittelt und bei gleichzeitiger fluorimetrischer Bestimmung der 11-OHCS auch die Menge proteingebundenen, bzw. sog. freien Cortisols (µg-%) berechnet.
2. Die diagnostische Brauchbarkeit der Methode wurde bei Patienten mit Nebennierenrindeninsuffizienz, mit Hypophysentumoren, nach Hypophysektomie, mit Cushing-Syndrom mit der fluorimetrischen Bestimmung der 11-OHCS verglichen. Die einfache Bestimmung der Cortisolbindung war bei hypophysektomierten Patienten der Bestimmung der 11-OHCS überlegen und entsprach der aufwendigeren ACTH-Belastung.
3. Falsch hohe fluorimetrische 11-OHCS-Spiegel im Plasma unter Spirolacton- oder Oestrogenbehandlung und in der Gravidität lassen sich durch Bestimmung der Cortisolbindung klären. Bei Schilddrüsenüberfunktion war das sog. freie Cortisol im Plasma relativ und absolut vermehrt, bei Schilddrüsenunterfunktion fand sich eine Zunahme des plasmaproteingebundenen Cortisols.1. Following incubation of labeled cortisol and plasma the percentages of protein bound and socalled free endogenous cortisol were determined by means of dextran gel filtration.
2. The diagnostic value of this method was compared with fluorimetric determinations of 11-OHCS for patients with adrenal insufficiency, Cushing-Syndrome, pituitary tumors and after hypophysectomy. In hypophysectomized patients the simple determination of protein bound cortisol was found to correlate well with diagnostic ACTH-infusion tests and to be more sensitive than fluorimetric determinations of 11-OHCS in 9 a.m. plasma.
3. Falsely elevated fluorimetric values of plasma 11-OHCS in patients treated with spirolactone or estrogens, resp. during pregnancy may be recognized through determination of cortisol binding. — In thyrotoxicosis socalled free cortisol was elevated, both relatively and absolutely; in hypothyroidism an increase of protein bound cortisol was found
Genetic load and transgenic mitigating genes in transgenic Brassica rapa (field mustard) × Brassica napus (oilseed rape) hybrid populations
<p>Abstract</p> <p>Background</p> <p>One theoretical explanation for the relatively poor performance of <it>Brassica rapa </it>(weed) × <it>Brassica napus </it>(crop) transgenic hybrids suggests that hybridization imparts a negative genetic load. Consequently, in hybrids genetic load could overshadow any benefits of fitness enhancing transgenes and become the limiting factor in transgenic hybrid persistence. Two types of genetic load were analyzed in this study: random/linkage-derived genetic load, and directly incorporated genetic load using a transgenic mitigation (TM) strategy. In order to measure the effects of random genetic load, hybrid productivity (seed yield and biomass) was correlated with crop- and weed-specific AFLP genomic markers. This portion of the study was designed to answer whether or not weed × transgenic crop hybrids possessing more crop genes were less competitive than hybrids containing fewer crop genes. The effects of directly incorporated genetic load (TM) were analyzed through transgene persistence data. TM strategies are proposed to decrease transgene persistence if gene flow and subsequent transgene introgression to a wild host were to occur.</p> <p>Results</p> <p>In the absence of interspecific competition, transgenic weed × crop hybrids benefited from having more crop-specific alleles. There was a positive correlation between performance and number of <it>B. napus </it>crop-specific AFLP markers [seed yield vs. marker number (r = 0.54, P = 0.0003) and vegetative dry biomass vs. marker number (r = 0.44, P = 0.005)]. However under interspecific competition with wheat or more weed-like conditions (i.e. representing a situation where hybrid plants emerge as volunteer weeds in subsequent cropping systems), there was a positive correlation between the number of <it>B. rapa </it>weed-specific AFLP markers and seed yield (r = 0.70, P = 0.0001), although no such correlation was detected for vegetative biomass. When genetic load was directly incorporated into the hybrid genome, by inserting a fitness-mitigating dwarfing gene that that is beneficial for crops but deleterious for weeds (a transgene mitigation measure), there was a dramatic decrease in the number of transgenic hybrid progeny persisting in the population.</p> <p>Conclusion</p> <p>The effects of genetic load of crop and in some situations, weed alleles might be beneficial under certain environmental conditions. However, when genetic load was directly incorporated into transgenic events, e.g., using a TM construct, the number of transgenic hybrids and persistence in weedy genomic backgrounds was significantly decreased.</p
Formation of Trans-Activation Competent HIV-1 Rev:RRE Complexes Requires the Recruitment of Multiple Protein Activation Domains
The HIV-1 Rev trans-activator is a nucleocytoplasmic shuttle protein that is essential for virus replication. Rev directly binds to unspliced and incompletely spliced viral RNA via the cis-acting Rev Response Element (RRE) sequence. Subsequently, Rev oligomerizes cooperatively and interacts with the cellular nuclear export receptor CRM1. In addition to mediating nuclear RNA export, Rev also affects the stability, translation and packaging of Rev-bound viral transcripts. Although it is established that Rev function requires the multimeric assembly of Rev molecules on the RRE, relatively little is known about how many Rev monomers are sufficient to form a trans-activation competent Rev:RRE complex, or which specific activity of Rev is affected by its oligomerization. We here analyzed by functional studies how homooligomer formation of Rev affects the trans-activation capacity of this essential HIV-1 regulatory protein. In a gain-of-function approach, we fused various heterologous dimerization domains to an otherwise oligomerization-defective Rev mutant and were able to demonstrate that oligomerization of Rev is not required per se for the nuclear export of this viral trans-activator. In contrast, however, the formation of Rev oligomers on the RRE is a precondition to trans-activation by directly affecting the nuclear export of Rev-regulated mRNA. Moreover, experimental evidence is provided showing that at least two protein activation domains are required for the formation of trans-activation competent Rev:RRE complexes. The presented data further refine the model of Rev trans-activation by directly demonstrating that Rev oligomerization on the RRE, thereby recruiting at least two protein activation domains, is required for nuclear export of unspliced and incompletely spliced viral RNA
Nucleolus: the fascinating nuclear body
Nucleoli are the prominent contrasted structures of the cell nucleus. In the nucleolus, ribosomal RNAs are synthesized, processed and assembled with ribosomal proteins. RNA polymerase I synthesizes the ribosomal RNAs and this activity is cell cycle regulated. The nucleolus reveals the functional organization of the nucleus in which the compartmentation of the different steps of ribosome biogenesis is observed whereas the nucleolar machineries are in permanent exchange with the nucleoplasm and other nuclear bodies. After mitosis, nucleolar assembly is a time and space regulated process controlled by the cell cycle. In addition, by generating a large volume in the nucleus with apparently no RNA polymerase II activity, the nucleolus creates a domain of retention/sequestration of molecules normally active outside the nucleolus. Viruses interact with the nucleolus and recruit nucleolar proteins to facilitate virus replication. The nucleolus is also a sensor of stress due to the redistribution of the ribosomal proteins in the nucleoplasm by nucleolus disruption. The nucleolus plays several crucial functions in the nucleus: in addition to its function as ribosome factory of the cells it is a multifunctional nuclear domain, and nucleolar activity is linked with several pathologies. Perspectives on the evolution of this research area are proposed
- …