The human respiratory syncytial virus nonstructural protein 1 regulates type I and type II interferon pathways.
Respiratory syncytial viruses encode a nonstructural protein (NS1) that interferes with type I and III interferon and other antiviral responses. Proteomic studies were conducted on human A549 type II alveolar epithelial cells and type I interferon-deficient Vero cells (African green monkey kidney cells) infected with wild-type and NS1-deficient clones of human respiratory syncytial virus to identify other potential pathway and molecular targets of NS1 interference. These analyses included two-dimensional differential gel electrophoresis and quantitative Western blotting. Surprisingly, NS1 was found to suppress the induction of manganese superoxide dismutase (SOD2) expression in A549 cells and to a much lesser degree Vero cells in response to infection. Because SOD2 is not directly inducible by type I interferons, it served as a marker to probe the impact of NS1 on signaling of other cytokines known to induce SOD2 expression and/or indirect effects of type I interferon signaling. Deductive analysis of results obtained from cell infection and cytokine stimulation studies indicated that interferon-γ signaling was a potential target of NS1, possibly as a result of modulation of STAT1 levels. However, this was not sufficient to explain the magnitude of the impact of NS1 on SOD2 induction in A549 cells. Vero cell infection experiments indicated that NS1 targeted a component of the type I interferon response that does not directly induce SOD2 expression but is required to induce another initiator of SOD2 expression. STAT2 was ruled out as a target of NS1 interference using quantitative Western blot analysis of infected A549 cells, but data were obtained to indicate that STAT1 was one of a number of potential targets of NS1. A label-free mass spectrometry-based quantitative approach is proposed as a means of more definitive identification of NS1 targets.
Protein Discovery Centre, Queensland Institute of Medical Research, Herston, Queensland 4029, Australia.
Marcus L
ML
Hastie
Madeleine J
MJ
Headlam
Nirav B
NB
Patel
Alexander A
AA
Bukreyev
Ursula J
UJ
Buchholz
Keyur A
KA
Dave
Emma L
EL
Norris
Cassandra L
CL
Wright
Kirsten M
KM
Spann
Peter L
PL
Collins
Jeffrey J
JJ
Gorman
D000818:Animals; D002374:Catalase / Q000235:genetics / Q000378:metabolism; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D016000:Cluster Analysis; D005786:Gene Expression Regulation; D054884:Host-Pathogen Interactions; D006801:Humans; D007370:Interferon Type I / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D007371:Interferon-gamma / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D018384:Oxidative Stress; D020543:Proteome / Q000235:genetics / Q000378:metabolism; D018113:Respiratory Syncytial Virus, Human / Q000502:physiology*; D050794:STAT1 Transcription Factor / Q000378:metabolism; D050795:STAT2 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction; D013482:Superoxide Dismutase / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D057825:Two-Dimensional Difference Gel Electrophoresis; D014709:Vero Cells; D017361:Viral Nonstructural Proteins / Q000502:physiology*; D000099156:Superoxide Dismutase 2
D002374:Catalase; D007370:Interferon Type I; D007371:Interferon-gamma; D047908:Intracellular Signaling Peptides and Proteins; D020543:Proteome; D050794:STAT1 Transcription Factor; D050795:STAT2 Transcription Factor; D013482:Superoxide Dismutase; D017361:Viral Nonstructural Proteins; D000099156:Superoxide Dismutase 2; C116468:IFIT3 protein, human; C494079:STAT1 protein, human; C494084:STAT2 protein, human