The phosphoinositide (PI) 3-kinase family

Abstract

Phosphoinositide (PI) 3-kinase was first observed in 1984 as a minor inositol lipid kinase activity associated with immunoprecipitated oncogene products (e.g. Src, Abl and polyoma mT antigen) and present in activated growth factor receptor complexes (e.g. PDGF receptor). In 1988, the enzyme associated with this activity was found to have the novel ability to phosphorylate the 3 position hydroxyl group of the inositol ring (see poster)of phosphatidylinositol (PtdIns). PI 3-kinase activities have been subsequently found in all eukaryotic cell types examined(Fry, 1994; Katso et al., 2001) and are linked to an incredibly diverse set of key cellular functions, including cell growth, proliferation, motility, differentiation, survival and intracellular trafficking (Fry, 1994; Rameh and Cantley, 1999; Fry, 2001; Katso et al., 2001). The emerging links between PI 3-kinase activity and many human maladies, including allergy, inflammation, heart disease and cancer, has made them the focus of intense study, and inhibitors of these enzymes are considered potential therapeutic agents (Stein and Waterfield,2000).Although the majority of published studies have focused on the classical p110-p85 (now known as class I) PI 3-kinases, it has emerged over the past 10 years that the PI 3-kinase superfamily (EC 2.7.1.137) is made up of a large family of structurally related enzymes, with differing PI substrate requirements and modes of regulation, which probably accounts for the reported diversity of function (Rameh and Cantley,1999; Fry, 2001; Katso et al., 2001). PCR cloning strategies and data mining of genome sequencing projects would seem to set the family limit at eight distinct PI 3-kinase catalytic subunits that are capable of phosphorylating inositol lipids. These eight isoforms have been divided into three functional classes on the basis of their protein domain structure, lipid substrate specificity and associated regulatory subunits:namely, the class I enzymes, p110α, p110β, p110δ and p110γ; the class II enzymes, PI3K-C2α, PI3K-C2β and PI3K-C2γ; and the sole class III enzyme, Vps34(Fry, 2001; Katso et al., 2001).The relationships between the kinase domains of the different enzymes identified in the human, fly, worm and yeast genomes are indicated in the poster by the non-rooted phylogenetic tree, which was prepared using ClustalX(hs, Homo sapien; dm, Drosophila melanogaster; ce, Caenorhabditis elegans; sp, Schizosaccharomyces pombe; sc, Saccharomyces cerivisiae). Simple eukaryotes, such as yeasts, and all plant species investigated to date seem to possess only a single class III PI 3-kinase. Multicellular invertebrate organisms, exemplified by C. elegans and D. melanogaster, have a single representative member of each of the three functional classes. Vertebrate genomes (human, mouse)contain eight distinct PI 3-kinase genes, with some of the family members being widely or ubiquitously expressed (e.g. p110α, p110β,PI3K-C2α, PI3K-C2β and Vps34), while others are more restricted to specific cell and tissue types (e.g. p110δ, p110γ and PI3K-C2γ). To add to this complexity there is a class IV group of PI-3-kinase-related protein serine/threonine kinases found in all eukaryotes(Kastan and Lim, 2000). Mammals have four such protein kinases: TOR (the target of the drug rapamycin), ATM (Ataxia telangiectasia mutated), ATR (Ataxia telangiectasia mutated related) and DNA-PK (DNA-dependent protein kinase). Here, we focus solely on the three classes of true PI 3-kinase.The various 3-phosphorylated lipid products that are produced by these enzymes [PtdIns(3)P, PtdIns(3,4)P2,PtdIns(3,5)P2 and PtdIns(3,4,5)P3]function as part of the mechanism by which a diverse set of signalling molecules, containing pleckstrin homology (PH), FYVE, Phox (PX) and other lipid-binding domains, are recruited to various cellular membranes(Rameh and Cantley, 1999; Wurmser et al., 1999; Ellson et al., 2002). Cellular PI 3-kinase activities are balanced by phosphoinositide 3-phosphatase activities, found in the tumour suppressor protein PTEN and in members of the myotubularin (MTM) family (Maehama et al.,2001).Two selective, chemically unrelated PI 3-kinase inhibitors are in wide use:namely, wortmannin and LY294002. Unfortunately, the use of these inhibitors is limited by the fact that they inhibit all known PI 3-kinase isoforms,including the class IV PI-3-kinase-like enzymes. This has led to many cellular processes being linked with PI 3-kinase activity, but considerable confusion as to which PI 3-kinase isoforms control specific cellular functions. In mammalian cells, wortmannin suppresses class I, class II PI3K-C2β and PI3K-C2γ, and class III PI 3-kinase activity with an IC50 in the 1-10 nM range, while it inhibits the class II PI3K-C2α isoform with an approximate IC50 of 400 nM, and class IV PI-3-kinase-related enzymes in the 100-300 nM range (data summarising many sources). Similarly,LY294002 inhibits all PI 3-kinases with an IC50 in the 1-50 μM range. Thu