Binding of influenza A virus NS1 protein to the inter-SH2 domain of p85 suggests a novel mechanism for phosphoinositide 3-kinase activation.
Influenza A virus NS1 protein stimulates host-cell phosphoinositide 3-kinase (PI3K) signaling by binding to the p85beta regulatory subunit of PI3K. Here, in an attempt to establish a mechanism for this activation, we report further on the functional interaction between NS1 and p85beta. Complex formation was found to be independent of NS1 RNA binding activity and is mediated by the C-terminal effector domain of NS1. Intriguingly, the primary direct binding site for NS1 on p85beta is the inter-SH2 domain, a coiled-coil structure that acts as a scaffold for the p110 catalytic subunit of PI3K. In vitro kinase activity assays, together with protein binding competition studies, reveal that NS1 does not displace p110 from the inter-SH2 domain, and indicate that NS1 can form an active heterotrimeric complex with PI3K. In addition, it was established that residues at the C terminus of the inter-SH2 domain are essential for mediating the interaction between p85beta and NS1. Equivalent residues in p85alpha have previously been implicated in the basal inhibition of p110. However, such p85alpha residues were unable to substitute for those in p85beta with regards NS1 binding. Overall, these data suggest a model by which NS1 activates PI3K catalytic activity by masking a normal regulatory element specific to the p85beta inter-SH2 domain.
Hale
Benjamin G
BG
Centre for Biomolecular Sciences, University of St. Andrews, St. Andrews, Fife KY16 9ST, United Kingdom. Electronic address: bgh1@st-andrews.ac.uk.
Batty
Ian H
IH
Division of Molecular Physiology, Faculty of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom.
Downes
C Peter
CP
Division of Molecular Physiology, Faculty of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom.
Randall
Richard E
RE
Centre for Biomolecular Sciences, University of St. Andrews, St. Andrews, Fife KY16 9ST, United Kingdom.
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002460:Cell Line; D004789:Enzyme Activation; D006801:Humans; D008954:Models, Biological; D008969:Molecular Sequence Data; D019869:Phosphatidylinositol 3-Kinases / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D012367:RNA, Viral / Q000378:metabolism; D016415:Sequence Alignment; D013329:Structure-Activity Relationship; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D018909:src Homology Domains
C085820:INS1 protein, influenza virus; D021122:Protein Subunits; D012367:RNA, Viral; D017361:Viral Nonstructural Proteins; D019869:Phosphatidylinositol 3-Kinases