Ebola virus infection inversely correlates with the overall expression levels of promyelocytic leukaemia (PML) protein in cultured cells.
BACKGROUND
Ebola virus causes severe, often fatal hemorrhagic fever in humans. The mechanism of escape from cellular anti-viral mechanisms is not yet fully understood. The promyelocytic leukaemia (PML) associated nuclear body is part of the interferon inducible cellular defense system. Several RNA viruses have been found to interfere with the anti-viral function of the PML body. The possible interaction between Ebola virus and the PML bodies has not yet been explored.


RESULTS
We found that two cell lines, Vero E6 and MCF7, support virus production at high and low levels respectively. The expression of viral proteins was visualized and quantified using high resolution immunofluorescence microscopy. Ebola encoded NP and VP35 accumulated in cytoplasmic inclusion bodies whereas VP40 was mainly membrane associated but it was also present diffusely in the cytoplasm as well as in the euchromatic areas of the nucleus. The anti-VP40 antibody also allowed the detection of extracellular virions. Interferon-alpha treatment decreased the production of all three viral proteins and delayed the development of cytopathic effects in both cell lines. Virus infection and interferon-alpha treatment induced high levels of PML protein expression in MCF7 but much less in Vero E6 cells. No disruption of PML bodies, a common phenomenon induced by a variety of different viruses, was observed.


CONCLUSIONS
We have established a simple fixation and immunofluorescence staining procedure that allows specific co-detection and precise sub-cellular localization of the PML nuclear bodies and the Ebola virus encoded proteins NP, VP35 and VP40 in formaldehyde treated cells. Interferon-alpha treatment delays virus production in vitro. Intact PML bodies may play an anti-viral role in Ebola infected cells.
Björndal
Asa Szekely
AS
Centre for Microbiological Preparedness, Swedish Institute for Infectious Disease Control (SMI), Nobels väg 18, 17182 Solna, Sweden. Asa.Bjorndal@mtc.ki.se
Szekely
Laszlo
L
Elgh
Fredrik
F
D000818:Animals; D002460:Cell Line / Q000821:virology*; D002522:Chlorocebus aethiops; D029043:Ebolavirus / Q000737:chemistry / Q000187:drug effects / Q000502:physiology*; D005455:Fluorescent Antibody Technique; D006801:Humans; D016898:Interferon-alpha / Q000494:pharmacology; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000502:physiology; D009687:Nuclear Proteins; D009698:Nucleoproteins / Q000032:analysis; D000071156:Promyelocytic Leukemia Protein; D013347:Subcellular Fractions; D014157:Transcription Factors / Q000096:biosynthesis* / Q000502:physiology; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins; D014709:Vero Cells; D014764:Viral Proteins / Q000032:analysis; D054334:Viral Regulatory and Accessory Proteins
D016898:Interferon-alpha; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D009698:Nucleoproteins; D000071156:Promyelocytic Leukemia Protein; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C080980:VP35 protein, filovirus; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C070576:PML protein, human