A human phosphatidylinositol 3-kinase complex related to the yeast Vps34p-Vps15p protein sorting system.

Abstract

Phosphoinositide (PI) 3‐kinases have been characterized as enzymes involved in receptor signal transduction in mammalian cells and in a complex which mediates protein trafficking in yeast. PI 3‐kinases linked to receptors with intrinsic or associated tyrosine kinase activity are heterodimeric proteins, consisting of p85 adaptor and p110 catalytic subunits, which can generate the 3‐phosphorylated forms of phosphatidylinositol (PtdIns), PtdIns4P and PtdIns(4,5)P2 as potential second messengers. Yeast Vps34p kinase, however, has a substrate specificity restricted to PtdIns and is a PtdIns 3‐kinase. Here the molecular characterization of a new human PtdIns 3‐kinase with extensive sequence homology to Vps34p is described. PtdIns 3‐kinase does not associate with p85 and phosphorylates PtdIns, but not PtdIns4P or PtdIns(4,5)P2. In vivo PtdIns 3‐kinase is in a complex with a cellular protein of 150 kDa, as detected by immunoprecipitation from human cells. Protein sequence analysis and cDNA cloning show that this 150 kDa protein is highly homologous to Vps15p, a 160 kDa protein serine/threonine kinase associated with yeast Vps34p. These results suggest that the major components of the yeast Vps intracellular trafficking complex are conserved in humans.

Endosomal Localization and Receptor Dynamics Determine Tyrosine Phosphorylation of Hepatocyte Growth Factor-Regulated Tyrosine Kinase Substrate

“…PI 3-kinase activity is required for Hrs phosphorylation. FYVE domains in proteins such as EEA1 and Hrs bind to PI 3-P, a product of some PI 3-kinase enzymes, most notably hVPS34 (36). Treatment of HeLa or BHK cells with the PI 3-kinase inhibitor, wortmannin, leads to an almost complete redistribution of EEA1 from membranes to the cytosol (16,20).…”

“…One consequence of treating cells with wortmannin is to inhibit the hVPS34 PI 3-kinase enzyme (36), which solely and constitutively produces PI 3-P, the lipid that accumulates on endosomes and binds to FYVE domains. If this interaction is necessary for phosphorylation, then removal or This phosphorylation is also greatly reduced by wortmannin treatment (73% Ϯ 8% inhibition; n ϭ 4 ( Fig.…”

Rapamycin and mTOR kinase inhibitors

“…The active site lysine (Lys802 in p110α and Lys833 in p110γ) is critical for catalysis and is highly conserved in lipid and protein kinases. Wortmannin also targets class II PI3K C2β (IC 50 =1.6 nM), the human class III PI3K hVps34p (IC 50 = 2.5 nM), type III PI4Ks (IC 50 =50-300 nM) and polo-like protein kinases that regulate mitosis (IC 50 =24-48 nM) [60][61][62][63][64]. Not surprisingly, considering the similarity between the PI3K and PIKK catalytic domains, wortmannin also irreversibly inhibits PIKK family members.…”

Inhibition of Calcium-independent Mannose 6-Phosphate Receptor Incorporation into trans-Golgi Network-derived Clathrin-coated Vesicles by Wortmannin

“…These heterodimeric PI3Ks exhibit an in vivo substrate preference for PIP 2 (17). The second class, specific for phosphatidylinositol (PtdIns), is exemplified by the wortmannin-resistant Vps34p from Saccharomyces cerevisiae (18) and a recently identified wortmannin-sensitive mammalian PtdIns 3-kinase (19).…”