The Chop Gene Contains an Element for the Positive Regulation of the Mitochondrial Unfolded Protein Response

We have previously reported on the discovery of a mitochondrial specific unfolded protein response (mtUPR) in mammalian cells, in which the accumulation of unfolded protein within the mitochondrial matrix results in the transcriptional activation of nuclear genes encoding mitochondrial stress proteins such as chaperonin 60, chaperonin 10, mtDnaJ, and ClpP, but not those encoding stress proteins of the endoplasmic reticulum (ER) or the cytosol. Analysis of the chaperonin 60/10 bidirectional promoter showed that the CHOP element was required for the mtUPR and that the transcription of the chop gene is activated by mtUPR. In order to investigate the role of CHOP in the mtUPR, we carried out a deletion analysis of the chop promoter. This revealed that the transcriptional activation of the chop gene by mtUPR is through an AP-1 (activator protein-1) element. This site lies alongside an ERSE element through which chop transcription is activated in response to the ER stress response (erUPR). Thus CHOP can be induced separately in response to 2 different stress response pathways. We also discuss the potential signal pathway between mitochondria and the nucleus for the mtUPR

RNA and DNA Bacteriophages as Molecular Diagnosis Controls in Clinical Virology: A Comprehensive Study of More than 45,000 Routine PCR Tests

Real-time PCR techniques are now commonly used for the detection of viral genomes in various human specimens and require for validation both external and internal controls (ECs and ICs). In particular, ICs added to clinical samples are necessary to monitor the extraction, reverse transcription, and amplification steps in order to detect false-negative results resulting from PCR-inhibition or errors in the technical procedure. Here, we performed a large scale evaluation of the use of bacteriophages as ICs in routine molecular diagnosis. This allowed to propose simple standardized procedures (i) to design specific ECs for both DNA and RNA viruses and (ii) to use T4 (DNA) or MS2 (RNA) phages as ICs in routine diagnosis. Various technical formats for using phages as ICs were optimised and validated. Subsequently, T4 and MS2 ICs were evaluated in routine real-time PCR or RT-PCR virological diagnostic tests, using a series of 8,950 clinical samples (representing 36 distinct specimen types) sent to our laboratory for the detection of a variety of DNA and RNA viruses. The frequency of inefficient detection of ICs was analyzed according to the nature of the sample. Inhibitors of enzymatic reactions were detected at high frequency in specific sample types such as heparinized blood and bone marrow (>70%), broncho-alveolar liquid (41%) and stools (36%). The use of T4 and MS2 phages as ICs proved to be cost-effective, flexible and adaptable to various technical procedures of real-time PCR detection in virology. It represents a valuable strategy for enhancing the quality of routine molecular diagnosis in laboratories that use in-house designed diagnostic systems, which can conveniently be associated to the use of specific synthetic ECs. The high rate of inhibitors observed in a variety of specimen types should stimulate the elaboration of improved technical protocols for the extraction and amplification of nucleic acids

Clinical Forms of Chikungunya in Gabon, 2010

Chikungunya fever (CHIK) is a disease caused by a virus transmitted to humans by infected mosquitos. The virus is responsible for multiple outbreaks in tropical and temperate areas worldwide, and is now a global concern. Clinical and biological features of the disease are poorly described, especially in Africa, where the disease is neglected because it is considered benign. During a recent CHIK outbreak that occurred in southeast Gabon, we prospectively studied clinical and biological features of 270 virologically confirmed cases. Fever and arthralgias were the predominant symptoms. Furthermore, variable and distinct clinical pictures including pure febrile, pure arthralgic and unusual forms (neither fever nor arthralgias) were detected. No severe forms or deaths were reported. These findings suggest that, during CHIK epidemics, some patients may not have classical symptoms (fever and arthralgias). Local surveillance is needed to detect any changes in the pathogenicity of this virus

Sp1 Expression Is Disrupted in Schizophrenia; A Possible Mechanism for the Abnormal Expression of Mitochondrial Complex I Genes, NDUFV1 and NDUFV2

The prevailing hypothesis regards schizophrenia as a polygenic disease, in which multiple genes combine with each other and with environmental stimuli to produce the variance of its clinical symptoms. We investigated whether the ubiquitous transcription factor Sp1 is abnormally expressed in schizophrenia, and consequently can affect the expression of genes implicated in this disorder. promoter by binding to its three GC-boxes. Both activation and binding were inhibited by mithramycin.These findings suggest that abnormality in Sp1, which can be the main activator/repressor or act in combination with additional transcription factors and is subjected to environmental stimuli, can contribute to the polygenic and clinically heterogeneous nature of schizophrenia

2nd International External Quality Control Assessment for the Molecular Diagnosis of Dengue Infections

Dengue viruses (DENV) are the most widespread arthropod-borne viruses which have shown an unexpected geographic expansion, as well as an increase in the number and severity of outbreaks in the last decades. In this context, the accurate diagnosis and reliable surveillance of dengue infections are essential. The laboratory diagnosis of dengue relies on the use of several methods detecting markers of DENV infection present in patient serum. Molecular diagnosis methods are usually rapid, sensitive, and simple when correctly standardized. Moreover, PCR-based diagnosis techniques are able to readily detect DENV during the acute phase of the disease and may assume an important role in dengue diagnosis and surveillance. Different reverse transcriptase PCR (RT-PCR) methods have been developed and are currently available and should be standardized in each laboratory to maintain high quality performance. In this work an External quality assessment (EQA) activity has been carried out to evaluate the accuracy and quality of laboratory data for the molecular diagnosis and surveillance of dengue, which involved worldwide dengue reference laboratories. In conclusion, RT-PCR techniques for dengue diagnosis applied by the participating laboratories demonstrated the need of further improvement in most laboratories

Chikungunya Virus Infection

Chikungunya virus (CHIKV) is an alphavirus transmitted by mosquitoes, mostly Aedes aegypti and Aedes albopictus. After half a century of focal outbreaks of acute febrile polyarthralgia in Africa and Asia, the disease unexpectedly spread in the past decade with large outbreaks in Africa and around the Indian Ocean and rare autochthonous transmission in temperate areas. This emergence brought new insights on its pathogenesis, notably the role of the A226V mutation that improved CHIKV fitness in Ae. albopictus and the possible CHIKV persistence in deep tissue sanctuaries for months after infection. Massive outbreaks also revealed new aspects of the acute stage: the high number of symptomatic cases, unexpected complications, mother-to-child transmission, and low lethality in debilitated patients. The follow-up of patients in epidemic areas has identified frequent, long-lasting, rheumatic disorders, including rare inflammatory joint destruction, and common chronic mood changes associated with quality-of-life impairment. Thus, the globalization of CHIKV exposes countries with Aedes mosquitoes both to brutal outbreaks of acute incapacitating episodes and endemic long-lasting disorders

Monitoring Winter and Summer Abundance of Cetaceans in the Pelagos Sanctuary (Northwestern Mediterranean Sea) Through Aerial Surveys

Systematic long-term monitoring of abundance is essential to inform conservation measures and evaluate their effectiveness. To instigate such work in the Pelagos Sanctuary in the Mediterranean, two aerial surveys were conducted in winter and summer 2009. A total of 467 (131 in winter, 336 in summer) sightings of 7 species was made. Sample sizes were sufficient to estimate abundance of fin whales in summer (148; 95% CI = 87–254) and striped dolphins in winter (19,462; 95% CI = 12 939–29 273) and in summer (38 488; 95% CI = 27 447–53 968). Numbers of animals within the Sanctuary are significantly higher in summer, when human activities and thus potential population level impacts are highest. Comparisons with data from past shipboard surveys suggest an appreciable decrease in fin whales within the Sanctuary area and an appreciable increase in striped dolphins. Aerial surveys proved to be more efficient than ship surveys, allowing more robust estimates, with smaller CIs and CVs. These results provide essential baseline data for this marine protected area and continued regular surveys will allow the effectiveness of the MPA in terms of cetacean conservation to be evaluated and inform future management measures. The collected data may also be crucial in assessing whether ship strikes, one of the main causes of death for fin whales in the Mediterranean, are affecting the Mediterranean population

Clinical and Virological Factors Influencing the Performance of a NS1 Antigen-Capture Assay and Potential Use as a Marker of Dengue Disease Severity

Dengue is the most prevalent arthropod-borne disease in tropical regions. The clinical manifestation may vary from asymptomatic to potentially fatal dengue shock syndrome. Early laboratory confirmation of dengue diagnosis is essential since many symptoms are not specific. Dengue non-structural protein 1 (NS1) may be used in simple antigen-capture ELISA for early detection of dengue virus infection. Our result demonstrated that the Platelia NS1 antigen detection kit had a quite low overall sensitivity. However, sensitivity rises significantly when used in combination with MAC-ELISA. When taking into account the various forms of dengue infection, the NS1 antigen detection was found relatively high in patients sampled during the first 3 days of fever onset, in patients with primary infection, DENV-1 infection, with high level of viremia and in mild form of dengue fever. In asymptomatically infected individuals, RT-PCR assay has proved to be more sensitive than NS1 antigen detection. Moreover, the NS1 antigen level correlated significantly with high viremia and low level of NS1 antigen was associated with more severe disease

UDP-N-Acetylglucosamine 2-Epimerase/N-Acetylmannosamine Kinase (GNE) Binds to Alpha-Actinin 1: Novel Pathways in Skeletal Muscle?

Hereditary inclusion body myopathy (HIBM) is a rare neuromuscular disorder caused by mutations in GNE, the key enzyme in the biosynthetic pathway of sialic acid. While the mechanism leading from GNE mutations to the HIBM phenotype is not yet understood, we searched for proteins potentially interacting with GNE, which could give some insights about novel putative biological functions of GNE in muscle. We used a Surface Plasmon Resonance (SPR)-Biosensor based assay to search for potential GNE interactors in anion exchanged fractions of human skeletal muscle primary culture cell lysate. Analysis of the positive fractions by in vitro binding assay revealed alpha-actinin 1 as a potential interactor of GNE. The direct interaction of the two proteins was assessed in vitro by SPR-Biosensor based kinetics analysis and in a cellular environment by a co-immunoprecipitation assay in GNE overexpressing 293T cells. Furthermore, immunohistochemistry on stretched mouse muscle suggest that both GNE and alpha-actinin 1 localize to an overlapping but not identical region of the myofibrillar apparatus centered on the Z line. The interaction of GNE with alpha-actinin 1 might point to its involvement in alpha-actinin mediated processes. In addition these studies illustrate for the first time the expression of the non-muscle form of alpha-actinin, alpha-actinin 1, in mature skeletal muscle tissue, opening novel avenues for its specific function in the sarcomere. Although no significant difference could be detected in the binding kinetics of alpha-actinin 1 with either wild type or mutant GNE in our SPR biosensor based analysis, further investigation is needed to determine whether and how the interaction of GNE with alpha-actinin 1 in skeletal muscle is relevant to the putative muscle-specific function of alpha-actinin 1, and to the muscle-restricted pathology of HIBM

PGC-1alpha Down-Regulation Affects the Antioxidant Response in Friedreich's Ataxia

BACKGROUND: Cells from individuals with Friedreich's ataxia (FRDA) show reduced activities of antioxidant enzymes and cannot up-regulate their expression when exposed to oxidative stress. This blunted antioxidant response may play a central role in the pathogenesis. We previously reported that Peroxisome Proliferator Activated Receptor Gamma (PPARgamma) Coactivator 1-alpha (PGC-1alpha), a transcriptional master regulator of mitochondrial biogenesis and antioxidant responses, is down-regulated in most cell types from FRDA patients and animal models. METHODOLOGY/PRINCIPAL FINDINGS: We used primary fibroblasts from FRDA patients and the knock in-knock out animal model for the disease (KIKO mouse) to determine basal superoxide dismutase 2 (SOD2) levels and the response to oxidative stress induced by the addition of hydrogen peroxide. We measured the same parameters after pharmacological stimulation of PGC-1alpha. Compared to control cells, PGC-1alpha and SOD2 levels were decreased in FRDA cells and did not change after addition of hydrogen peroxide. PGC-1alpha direct silencing with siRNA in control fibroblasts led to a similar loss of SOD2 response to oxidative stress as observed in FRDA fibroblasts. PGC-1alpha activation with the PPARgamma agonist (Pioglitazone) or with a cAMP-dependent protein kinase (AMPK) agonist (AICAR) restored normal SOD2 induction. Treatment of the KIKO mice with Pioglitazone significantly up-regulates SOD2 in cerebellum and spinal cord. CONCLUSIONS/SIGNIFICANCE: PGC-1alpha down-regulation is likely to contribute to the blunted antioxidant response observed in cells from FRDA patients. This response can be restored by AMPK and PPARgamma agonists, suggesting a potential therapeutic approach for FRDA.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe