Membrane Localization of Phosphatidylinositol 3-Kinase Is Sufficient To Activate Multiple Signal-Transducing Kinase Pathways

Abstract

Phosphatidylinositol (PI) 3-kinase is a cytoplasmic signaling molecule recruited to the membrane by activated growth factor receptors. The p85 subunit of PI 3-kinase links the catalytic p110 subunit to activated growth factor receptors and is required for enzymatic activity of p110. In this report, we describe the effects of expressing novel forms of p110 that are targeted to the membrane by either N-terminal myristoylation or C-terminal farnesylation. The expression of membrane-localized p110 is sufficient to trigger downstream responses characteristic of growth factor action, including the stimulation of pp70 S6 kinase, Akt/Rac, and Jun N-terminal kinase (JNK). These responses can also be triggered by expression of a form of p110 (p110*) that is cytosolic but exhibits a high specific activity. Finally, targeting of p110* to the membrane results in maximal activation of downstream responses. Our data demonstrate that either membrane-targeted forms of p110 or a form of p110 with high specific activity can act as constitutively active PI 3-kinases and induce PI 3-kinasedependent responses in the absence of growth factor stimulation. The results also show that PI 3-kinase activation is sufficient to stimulate several kinases that appear to function in different signaling pathways.Phosphatidylinositol (PI) 3-kinase activity has been implicated in the regulation of a number of different cellular responses, including the regulation of cell growth. Oncogenic transformation or the stimulation of cells with growth factors results in an increased level in the phospholipid products of PI 3-kinase (for reviews, see references 4, 23, and 50). Mutants of platelet-derived growth factor (PDGF) receptor or certain oncogenes which fail to activate PI 3-kinase are deficient in triggering either mitogenic responses or oncogenic transformation, respectively. These data suggest that PI 3-kinase is an important mediator of signaling events that regulate cell growth and cellular transformation.PI 3-kinase is a heterodimeric complex consisting of 85-and 110-kDa subunits (p85 and p110) (23). The p85 subunit consists of multiple domains including a Src homology 3 (SH3) domain, a breakpoint cluster region domain, and two SH2 domains. The two SH2 domains bind tyrosine-phosphorylated receptors and in this manner recruit the p85-p110 complex to activated receptors. The two SH2 domains are separated by the inter-SH2 (iSH2) region. The iSH2 domain mediates the interaction of p85 with p110, and this interaction is required for the enzymatic activity of p110 (28) (Fig. 1). It is possible that the role of p85 is to target p110 to the membrane, where its lipid substrates reside (23).PI 3-kinase has been implicated in the regulation of many other cellular processes, including the reorganization of the actin cytoskeleton (24,30,40,59), receptor internalization (21), histamine secretion (63), neutrophil activation (56), platelet activation (64), cell migration (31), glucose transport (41), and vesicular sorting (48). PI 3-kinase was implicated in the...

Concomitant activation of MEK-1 and Rac-1 increases the proliferative potential of thyroid epithelial cells, without affecting their differentiation

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