Evidence that the respiratory syncytial virus polymerase complex associates with lipid rafts in virus-infected cells: a proteomic analysis.
The interaction between the respiratory syncytial virus (RSV) polymerase complex and lipid rafts was examined in HEp2 cells. Lipid-raft membranes were prepared from virus-infected cells and their protein content was analysed by Western blotting and mass spectrometry. This analysis revealed the presence of the N, P, L, M2-1 and M proteins. However, these proteins appeared to differ from one another in their association with these structures, with the M2-1 protein showing a greater partitioning into raft membranes compared to that of the N, P or M proteins. Determination of the polymerase activity profile of the gradient fractions revealed that 95% of the detectable viral enzyme activity was associated with lipid-raft membranes. Furthermore, analysis of virus-infected cells by confocal microscopy suggested an association between these proteins and the raft-lipid, GM1. Together, these results provide evidence that the RSV polymerase complex is able to associate with lipid rafts in virus-infected cells.
McDonald
Terence P
TP
MRC Virology Unit, Institute of Virology, Glasgow G11 5JR, UK.
Pitt
Andrew R
AR
Brown
Gaie
G
Rixon
Helen W McL
HW
Sugrue
Richard J
RJ
D000595:Amino Acid Sequence; D045744:Cell Line, Tumor; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D013058:Mass Spectrometry; D021962:Membrane Microdomains / Q000821:virology*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D018113:Respiratory Syncytial Virus, Human / Q000201:enzymology*
D010446:Peptide Fragments; D012321:DNA-Directed RNA Polymerases