Direct Interaction of Major Histocompatibility Complex Class II-derived Peptides with Class Ia Phosphoinositide 3-Kinase Results in Dose-dependent Stimulatory Effects

Abstract

Peptides corresponding to residues 65-79 of human lymphocyte antigen class II sequence (DQA*03011) are cell-permeable and at high concentrations block activation of protein kinase B/Akt and p70-S6 kinase in T-cells, effects attributed to inhibition of phosphoinositide (PI) 3-kinase activity. To understand the molecular basis of this, we analyzed the effect this peptide had on activity of class I PI 3-kinases. Although there was no effect on the activity of class Ib PI 3-kinase or on the protein kinase activity of class I PI 3-kinases, there was a biphasic effect on lipid kinase activity of the class Ia enzymes. There was an inhibition of activity at higher peptide concentrations because of a formation of insoluble complexes between peptide and enzyme. Conversely, at lower peptide concentrations there was a profound activation of PI 3-kinase activity of class Ia PI 3-kinases. Studies of peptide variants revealed that all active peptides conform to heptad repeat motifs characteristic of coiled-coil helices. Surface plasmon resonance studies confirmed direct sequence-specific binding of active peptide to the p85␣ adapter subunit of class Ia PI 3-kinase. Active peptides also activated protein kinase B and extracellular signal-regulated kinase (ERK) in vivo in a wortmannin-sensitive manner while reducing recoverable cellular p85 levels. These results indicate that the human lymphocyte antigen class II-derived peptides regulate PI 3-kinase by direct interaction, probably via the coiled-coil domain. These peptides define a novel mechanism of regulating PI 3-kinase and will provide a useful tool for specifically dissecting the function of class Ia PI 3-kinase in cells and for probing structurefunction relationships in the class Ia PI 3-kinase heterodimers. Phosphoinositide (PI)1 3-kinases play a pivotal role in signaling pathways regulating a wide range of cellular processes including apoptosis, metabolism, cell growth, cell proliferation, and cytoskeletal rearrangements (1-3). Several classes of PI 3-kinase exist, but tyrosine kinase-linked receptors signal mainly through class Ia PI 3-kinases, which are bifunctional kinases, possessing both lipid and protein kinase activities (4). The class Ia PI 3-kinases can be regulated in a number of ways. These include interactions via the SH2, SH3, and Bcr homology domains of the adapter subunit and direct interaction of Ras through the Ras binding domain of the catalytic subunit (1-3).A potentially novel mechanism for regulating PI 3-kinase activity has recently emerged from studies of the properties of biologically active peptides derived from class II MHC molecules (5). These studies identified a peptide fragment of DQA*03011 ␣ chain allele of MHC class II that was capable of blocking IL2-induced T-cell proliferation and thus offered a possibly novel means of achieving immunosuppression. Further work found that treatment of T-cells with one of these peptides, corresponding to the residues 65-79 of DQA*03011, blocked IL2-induced activation of protein kinase B/Akt and p70-S6K a...

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