HIV-1 Vpr Triggers Mitochondrial Destruction by Impairing Mfn2-Mediated ER-Mitochondria Interaction

Human immunodeficiency virus 1 (HIV-1) viral protein R (Vpr) has been shown to induce host cell death by increasing the permeability of mitochondrial outer membrane (MOM). The mechanism underlying the damage to the mitochondria by Vpr, however, is not clearly illustrated. In this study, Vpr that is introduced, via transient transfection or lentivirus infection, into the human embryonic kidney cell line HEK293, human CD4+ T lymphoblast cell line SupT1, or human primary CD4+ T cells serves as the model system to study the molecular mechanism of Vpr-mediated HIV-1 pathogenesis. The results show that Vpr injures MOM and causes a loss in membrane potential (MMP) by posttranscriptionally reducing the expression of mitofusin 2 (Mfn2) via VprBP-DDB1-CUL4A ubiquitin ligase complex, gradually weakening MOM, and increasing mitochondrial deformation. Vpr also markedly decreases cytoplasmic levels of dynamin-related protein 1 (DRP1) and increases bulging in mitochondria-associated membranes (MAM), the specific regions of endoplasmic reticulum (ER) which form physical contacts with the mitochondria. Overexpression of Mfn2 and DRP1 significantly decreased the loss of MMP and apoptotic cell death caused by Vpr. Furthermore, by employing time-lapse confocal fluorescence microscopy, we identify the transport of Vpr protein from the ER, via MAM to the mitochondria. Taken together, our results suggest that Vpr-mediated cellular damage may occur on an alternative protein transport pathway from the ER, via MAM to the mitochondria, which are modulated by Mfn2 and DRP1

Transposon based tagging: IRAP, REMAP, and iPBS

Retrotransposons are a major component of virtually all eukaryotic genomes, which makes them useful as molecular markers. Various molecular marker systems have been developed that exploit the ubiquitous nature of these genetic elements and their property of stable integration into dispersed chromosomal loci that are polymorphic within species. To detect polymorphisms for retrotransposon insertions, marker systems generally rely on PCR amplification between the retrotransposon termini and some component of flanking genomic DNA. The main methods of IRAP, REMAP, RBIP, and SSAP all detect the polymorphic sites at which the retrotransposon DNA is integrated into the genome. Marker systems exploiting these methods can be easily developed and are inexpensively deployed in the absence of extensive genome sequence data. Here, we describe protocols for the IRAP, REMAP and iPBS techniques, including methods for PCR amplification with a single primer or with two primers, agarose gel electrophoresis of the product using optimal electrophoresis buffers, we also describe iPBS techniques for the rapid isolation of retrotransposon termini and full-length elements.Peer reviewe

Cardiovascular and lifestyle risk factors in lumbar radicular pain or clinically defined sciatica: a systematic review

Lumbar radicular pain is a fairly common health problem, yet its risk factors are far from clear. There are no published systematic reviews on associations between cardiovascular or lifestyle risk factors and lumbar radicular pain or sciatica. The aim of this systematic literature review was to assess associations between these risk factors and lumbar radicular pain or sciatica. We conducted a systematic search of the Medline database for all original articles on lumbar radicular pain or sciatica published until August 2006. Twenty-two papers from 19 studies were included in the review. Overweight or obesity was associated with sciatica in most of the case-control and cohort studies. Some studies showed an increased risk of lumbar radicular pain in smokers with a long smoking history or in those with high levels of physical activity. A few case-control studies showed an association between serum C-reactive protein and sciatica. No consistent associations were found for serum lipids levels or high blood pressure. In summary, the associations of overweight, long smoking history, high physical activity and a high serum C-reactive protein level with lumbar radicular pain or sciatica were substantiated by the present review. However, more prospective studies are needed in order to further clarify these associations and the mechanisms of action