Browsing by Author "Ramdohr, Pablo"

Now showing 1 - 4 of 4
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    Activity of the human immunodeficiency virus type 1 cell cycle-dependent internal ribosomal entry site is modulated by IRES trans-acting factors
    (OXFORD UNIV PRESS, 2011) Vallejos, Maricarmen; Deforges, Jules; Plank, Terra Dawn M.; Letelier, Alejandro; Ramdohr, Pablo; Abraham, Christopher G.; Valiente Echeverria, Fernando; Kieft, Jeffrey S.; Sargueil, Bruno; Lopez Lastra, Marcelo
    The 5' leader of the human immunodeficiency virus type 1 (HIV-1) genomic RNA harbors an internal ribosome entry site (IRES) that is functional during the G2/M phase of the cell cycle. Here we show that translation initiation mediated by the HIV-1 IRES requires the participation of trans-acting cellular factors other than the canonical translational machinery. We used 'standard' chemical and enzymatic probes and an 'RNA SHAPE' analysis to model the structure of the HIV-1 5' leader and we show, by means of a footprinting assay, that G2/M extracts provide protections to regions previously identified as crucial for HIV-1 IRES activity. We also assessed the impact of mutations on IRES function. Strikingly, mutations did not significantly affect IRES activity suggesting that the requirement for pre-formed stable secondary or tertiary structure within the HIV-1 IRES may not be as strict as has been described for other viral IRESes. Finally, we used a proteomic approach to identify cellular proteins within the G2/M extracts that interact with the HIV-1 5' leader. Together, data show that HIV-1 IRES-mediated translation initiation is modulated by cellular proteins.
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    Andes Virus Antigens Are Shed in Urine of Patients with Acute Hantavirus Cardiopulmonary Syndrome
    (AMER SOC MICROBIOLOGY, 2009) Godoy, Paula; Marsac, Delphine; Stefas, Elias; Ferrer, Pablo; Tischler, Nicole D.; Pino, Karla; Ramdohr, Pablo; Vial, Pablo; Valenzuela, Pablo D. T.; Ferres, Marcela; Veas, Francisco; Lopez Lastra, Marcelo
    Hantavirus cardiopulmonary syndrome (HCPS) is a highly pathogenic emerging disease (40% case fatality rate) caused by New World hantaviruses. Hantavirus infections are transmitted to humans mainly by inhalation of virus-contaminated aerosol particles of rodent excreta and secretions. At present, there are no antiviral drugs or immunotherapeutic agents available for the treatment of hantaviral infection, and the survival rates for infected patients hinge largely on early virus recognition and hospital admission and aggressive pulmonary and hemodynamic support. In this study, we show that Andes virus (ANDV) interacts with human apolipoprotein H (ApoH) and that ApoH-coated magnetic beads or ApoH-coated enzyme-linked immunosorbent assay plates can be used to capture and concentrate the virus from complex biological mixtures, such as serum and urine, allowing it to be detected by both immunological and molecular approaches. In addition, we report that ANDV-antigens and infectious virus are shed in urine of HCPS patients.
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    The 5'-untranslated region of the mouse mammary tumor virus mRNA exhibits cap-independent translation initiation
    (OXFORD UNIV PRESS, 2010) Vallejos, Maricarmen; Ramdohr, Pablo; Valiente Echeverria, Fernando; Tapia, Karla; Rodriguez, Felipe E.; Lowy, Fernando; Pablo Huidobro Toro, J.; Dangerfield, John A.; Lopez Lastra, Marcelo
    In this study, we demonstrate the identification of an internal ribosome entry site (IRES) within the 5'-untranslated region (5'-UTR) of the mouse mammary tumor virus (MMTV). The 5'-UTR of the full-length mRNA derived from the infectious, complete MMTV genome was cloned into a dual luciferase reporter construct containing an upstream Renilla luciferase gene (RLuc) and a downstream firefly luciferase gene (FLuc). In rabbit reticulocyte lysate, the MMTV 5'-UTR was capable of driving translation of the second cistron. In vitro translational activity from the MMTV 5'-UTR was resistant to the addition of m(7)GpppG cap-analog and cleavage of eIF4G by foot-and-mouth disease virus (FMDV) L-protease. IRES activity was also demonstrated in the Xenopus laevis oocyte by micro-injection of capped and polyadenylated bicistronic RNAs harboring the MMTV-5'-UTR. Finally, transfection assays showed that the MMTV-IRES exhibits cell type-dependent translational activity, suggesting a requirement for as yet unidentified cellular factors for its optimal function.
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    The Elav-like protein HuR exerts translational control of viral internal ribosome entry sites
    (ACADEMIC PRESS INC ELSEVIER SCIENCE, 2009) Rivas Aravena, Andrea; Ramdohr, Pablo; Vallejos, Maricarmen; Valiente Echeverria, Fernando; Dormoy Raclet, Virginie; Rodriguez, Felipe; Pino, Karla; Holzmann, Cristian; Huidobro Toro, J. Pablo; Gallouzi, Imed Eddine; Lopez Lastra, Marcelo
    The human embryonic-lethal abnormal vision (ELAV)-like protein. HuR, has been recently found to be involved in the regulation of protein synthesis. In this study we show that HuR participates in the translational control of the HIV-1 and HCV IRES elements. HuR functions as a repressor of HIV-1]RES activity and acts as an activator of the HCV]RES. The effect of HuR was evaluated in three independent experimental systems, rabbit reticulocyte lysate, HeLa cells, and Xenopus laevis oocytes, using both overexpression and knockdown approaches. Furthermore, results suggest that HuR mediated regulation of HIV-1 and HCV IRESes does not require direct binding of the protein to the RNA nor does it need the nuclear translocation of the IRES-containing RNAs. Finally, we show that HuR has a negative impact on post-integration steps of the HIV-1 replication cycle. Thus, our observations yield novel insights into the role of HuR in the post-transcriptional regulation of HCV and HIV-1 gene expression. (C) 2009 Elsevier Inc. All rights reserved.