Rerouting of Plant Late Endocytic Trafficking Toward a Pathogen Interface

A number of plant pathogenic and symbiotic microbes produce specialized cellular structures that invade host cells where they remain enveloped by host‐derived membranes. The mechanisms underlying the biogenesis and functions of host–microbe interfaces are poorly understood. Here, we show that plant late endocytic trafficking is diverted toward the extrahaustorial membrane (EHM); a host–pathogen interface that develops in plant cells invaded by Irish potato famine pathogen Phytophthora infestans. A late endosome and tonoplast marker protein Rab7 GTPase RabG3c, but not a tonoplast‐localized sucrose transporter, is recruited to the EHM, suggesting specific rerouting of vacuole‐targeted late endosomes to a host–pathogen interface. We revealed the dynamic nature of this process by showing that, upon activation, a cell surface immune receptor traffics toward the haustorial interface. Our work provides insight into the biogenesis of the EHM and reveals dynamic processes that recruit membrane compartments and immune receptors to this host–pathogen interface.      

Mutation Spectrum of RB1 Gene in Unilateral Retinoblastoma Cases from Tunisia and Correlations with Clinical Features

Retinoblastoma, an embryonic neoplasm of retinal origin, is the most common primary intraocular malignancy in children. Somatic inactivation of both alleles of the RB1 tumor suppressor gene in a retinal progenitor cell through diverse mechanisms including genetic and epigenetic modifications, is the crucial event in initiation of tumorigenesis in most cases of isolated unilateral retinoblastoma. We analyzed DNA from tumor tissue and from peripheral blood to determine the RB1 mutation status and seek correlations with clinical features of 37 unrelated cases of Tunisian origin with sporadic retinoblastoma. All cases were unilateral except one who presented with bilateral disease, in whom no germline coding sequence alteration was identified. A multi-step mutation scanning protocol identified bi-allelic inactivation of RB1 gene in 30 (81%) of the samples tested. A total of 7 novel mutations were identified. There were three tumors without any detectable mutation while a subset contained multiple mutations in RB1 gene. The latter group included tumors collected after treatment with chemotherapy. There were seven individuals with germline mutations and all presented with advanced stage of tumor. There was no difference in age of onset of RB based on the germline mutation status. Thus 20% of the individuals with sporadic unilateral RB in this series carried germline mutations and indicate the importance of genetic testing all children with sporadic retinoblastoma. These findings help to characterize the spectrum of mutations present in the Tunisian population and can improve genetic diagnosis of retinoblastoma.     

Dietary Supplement Enriched in Antioxidants and Omega-3 Protects from Progressive Light-Induced Retinal Degeneration

In the present study, we have evaluated one of the dietary supplements enriched with antioxidants and fish oil used in clinical care for patient with age-related macular degeneration. Rats were orally fed by a gastric canula daily with 0.2 ml of water or dietary supplement until they were sacrificed. After one week of treatment, animals were either sacrificed for lipid analysis in plasma and retina, or used for evaluation of rod-response recovery by electroretinography (ERG) followed by their sacrifice to measure rhodopsin content, or used for progressive light-induced retinal degeneration (PLIRD). For PLIRD, animals were transferred to bright cyclic light for one week. Retinal damage was quantified by ERG, histology and detection of apoptotic nuclei. Animals kept in dim-cyclic-light were processed in parallel. PLIRD induced a thinning of the outer nuclear layer and a reduction of the b-wave amplitude of the ERG in the water group. Retinal structure and function were preserved in supplemented animals. Supplement induced a significant increase in omega-3 fatty acids in plasma by 168% for eicosapentaenoic acid (EPA), 142% for docosapentaenoic acid (DPA) and 19% for docosahexaenoic acid (DHA) and a decrease in the omega-6 fatty acids, DPA by 28%. In the retina, supplement induced significant reduction of linolenic acid by 67% and an increase in EPA and DPA by 80% and 72%, respectively, associated with significant decrease in omega-6 DPA by 42%. Supplement did not affect rhodopsin content or rod-response recovery. The present data indicate that supplement rapidly modified the fatty acid content and induced an accumulation of EPA in the retina without affecting rhodopsin content or recovery. In addition, it protected the retina from oxidative stress induced by light. Therefore, this supplement might be beneficial to slow down progression of certain retinal degeneration.     

<Emphasis Type="Italic">Saccharothrix ghardaiensis</Emphasis> sp. nov., an actinobacterium isolated from Saharan soil

The taxonomic position of a new Saccharothrix strain, designated MB46^T, isolated from a Saharan soil sample collected in Mzab region (Ghardaïa province, South Algeria) was established following a polyphasic approach. The novel microorganism has morphological and chemical characteristics typical of the members of the genus Saccharothrix and formed a phyletic line at the periphery of the Saccharothrix espanaensis subcluster in the 16S rRNA gene dendrograms. Results of the 16S rRNA gene sequence comparisons revealed that strain MB46^T shares high degrees of similarity with S. espanaensis DSM 44229^T (99.2%), Saccharothrix variisporea DSM 43911^T (98.7%) and Saccharothrix texasensis NRRL B-16134^T (98.6%). However, the new strain exhibited only 12.5–17.5% DNA relatedness to the neighbouring Saccharothrix spp. On the basis of phenotypic characteristics, 16S rRNA gene sequence comparisons and DNA-DNA hybridizations, strain MB46^T is concluded to represent a novel species of the genus Saccharothrix, for which the name Saccharothrix ghardaiensis sp. nov. (type strain MB46^T = DSM 46886^T = CECT 9046^T) is proposed.     

The chloroplast protein RPH1 plays a role in the immune response of Arabidopsis to Phytophthora brassicae

Plant immune responses to pathogens are often associated with enhanced production of reactive oxygen species (ROS), known as the oxidative burst, and with rapid hypersensitive host cell death (the hypersensitive response, HR) at sites of attempted infection. It is generally accepted that the oxidative burst acts as a promotive signal for HR, and that HR is highly correlated with efficient disease resistance. We have identified the Arabidopsis mutant rph1 ( resistance to Phytophthora 1 ), which is susceptible to the oomycete pathogen Phytophthora brassicae despite rapid induction of HR. The susceptibility of rph1 was specific for P. brassicae and coincided with a reduced oxidative burst, a runaway cell-death response, and failure to properly activate the expression of defence-related genes. From these results, we conclude that, in the immune response to P. brassicae , (i) HR is not sufficient to stop the pathogen, (ii) HR initiation can occur in the absence of a major oxidative burst, (iii) the oxidative burst plays a role in limiting the spread of cell death, and (iv) RPH1 is a positive regulator of the P. brassicae -induced oxidative burst and enhanced expression of defence-related genes. Surprisingly, RPH1 encodes an evolutionary highly conserved chloroplast protein, indicating a function of this organelle in activation of a subset of immune reactions in response to P. brassicae . The disease resistance-related role of RPH1 was not limited to the Arabidopsis model system. Silencing of the potato homolog StRPH1 in a resistant potato cultivar caused susceptibility to the late blight pathogen Phytophthora infestans .     

Molecular optimization of rabies virus glycoprotein expression in Pichia pastoris

In this work, different approaches were investigated to enhance the expression rabies virus glycoprotein (RABV‐G) in the yeast Pichia pastoris; this membrane protein is responsible for the synthesis of rabies neutralizing antibodies. First, the impact of synonymous codon usage bias was examined and an optimized RABV‐G gene was synthesized. Nevertheless, data showed that the secretion of the optimized RABV‐G gene was not tremendously increased as compared with the non‐optimized one. In addition, similar levels of RABV‐G were obtained when α‐factor mating factor from Saccharomyces cerevisiae or the acid phosphatase PHO1 was used as a secretion signal. Therefore, sequence optimization and secretion signal were not the major bottlenecks for high‐level expression of RABV‐G in P. pastoris. Unfolded protein response (UPR) was induced in clones containing high copy number of RABV‐G expression cassette indicating that folding was the limiting step for RABV‐G secretion. To circumvent this limitation, co‐overexpression of five factors involved in oxidative protein folding was investigated. Among these factors only PDI1, ERO1 and GPX1 proved their benefit to enhance the expression. The highest expression level of RABV‐G reached 1230 ng ml^?1. Competitive neutralizing assay confirmed that the recombinant protein was produced in the correct conformational form in this host.     

Target cell APOBEC3C can induce limited G-to-A mutation in HIV-1

The evolutionary success of primate lentiviruses reflects their high capacity to mutate and adapt to new host species, immune responses within individual hosts, and, in recent years, antiviral drugs. APOBEC3G (A3G) and APOBEC3F (A3F) are host cell DNA-editing enzymes that induce extensive HIV-1 mutation that severely attenuates viral replication. The HIV-1 virion infectivity factor (Vif), expressed in vivo, counteracts the antiviral activity of A3G and A3F by inducing their degradation. Other APOBECs may contribute more to viral diversity by inducing less extensive mutations allowing viral replication to persist. Here we show that in APOBEC3C (A3C)-expressing cells infected with the patient-derived HIV-1 molecular clones 210WW, 210WM, 210MW, and 210MM, and the lab-adapted molecular clone LAI, viral G-to-A mutations were detected in the presence of Vif expression. Mutations occurred primarily in the GA context and were relatively infrequent, thereby allowing for spreading infection. The mutations were absent in cells lacking A3C but were induced after transient expression of A3C in the infected target cell. Inhibiting endogenous A3C by RNA interference in Magi cells prevented the viral mutations. Thus, A3C is necessary and sufficient for G-to-A mutations in some HIV-1 strains. A3C-induced mutations occur at levels that allow replication to persist and may therefore contribute to viral diversity. Developing drugs that inhibit A3C may be a novel strategy for delaying viral escape from immune or antiretroviral inhibition.     

First characterization of congenital myasthenic syndrome type 5 in North Africa

Molecular Biology Reports - Congenital myasthenic syndromes (CMS) are associated with defects in the structure and the function of neuromuscular junctions. These rare disorders can result from...     

Diversity of Degradation Pathways of Some Aromatic Compounds by Phenotype and Genotype Testing in Acinetobacter Strains

Phenotype and genotype testing were combined in this study to identify the diversity of five Acinetobacter strains to degrade some low-molecular-weight aromatic compounds. Three groups were identified: Group one, A. calcoaceticus PHEA-2 and A. baumanii UMI-95, which had the ability to degrade catechol and protocatchuate, and could catabolize vanillic acid and phenol; Group two, Acinetobacter sp. MM1, which had the ability to degrade protocatchuate, but not catechol, just could catabolize vanillic acid; and Group three, A. lwoffi UMI-8, A. junii 85, which degraded neither catechol nor protocatchuate, not could catabolize vanillic acid or phenol. This revised version was published online in July 2006 with corrections to the Cover Date.