Signaling via Class IA Phosphoinositide 3-Kinases (PI3K) in Human, Breast-Derived Cell Lines

• Published in: PloS one Vol. 8; no. 10; p. e75045
• Main Authors: Juvin, Veronique, Malek, Mouhannad, Anderson, Karen E., Dion, Carine, Chessa, Tamara, Lecureuil, Charlotte, Ferguson, G. John, Cosulich, Sabina, Hawkins, Phillip T., Stephens, Len R.
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 04.10.2013; Public Library of Science (PLoS)
• Subjects: 1-Phosphatidylinositol 3-kinase; AKT protein; Alleles; Cancer; Catalysis; Cell growth; Cell Line; Cell Line, Tumor; Cell lines; Cell migration; Chemokinesis; Chemotaxis; Epidermal growth factor; Epidermal Growth Factor - pharmacology; Epidermal growth factors; Female; Gene mutation; Humans; Inhibitors; Kinases; Leukocyte migration; Life Sciences; Motility; mRNA; Mutants; Mutation; Phosphatase; Phosphatases; Phosphatidylinositol; Phosphatidylinositol 3-Kinases - genetics; Phosphatidylinositol 3-Kinases - metabolism; Phosphorylation; Phosphorylation - drug effects; Phytopathology and phytopharmacy; Prostate; Protein kinases; Proteins; PTEN Phosphohydrolase - genetics; PTEN Phosphohydrolase - metabolism; PTEN protein; Regulation; Regulatory subunits; Signal Transduction - genetics; Signal Transduction - physiology; Signaling; Tumors; Vegetal Biology; France
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0075045

We have addressed the differential roles of class I Phosphoinositide 3-kinases (PI3K) in human breast-derived MCF10a (and iso-genetic derivatives) and MDA-MB 231 and 468 cells. Class I PI3Ks are heterodimers of p110 catalytic (α, β, δ and γ) and p50-101 regulatory subunits and make the signaling lipid, phosphatidylinositol (3,4,5)-trisphosphate (PtdIns(3,4,5)P3) that can activate effectors, eg protein kinase B (PKB), and responses, eg migration. The PtdIns(3,4,5)P3-3-phosphatase and tumour-suppressor, PTEN inhibits this pathway. p110α, but not other p110s, has a number of onco-mutant variants that are commonly found in cancers. mRNA-seq data shows that MCF10a cells express p110β>>α>δ with undetectable p110γ. Despite this, EGF-stimulated phosphorylation of PKB depended upon p110α-, but not β- or δ- activity. EGF-stimulated chemokinesis, but not chemotaxis, was also dependent upon p110α, but not β- or δ- activity. In the presence of single, endogenous alleles of onco-mutant p110α (H1047R or E545K), basal, but not EGF-stimulated, phosphorylation of PKB was increased and the effect of EGF was fully reversed by p110α inhibitors. Cells expressing either onco-mutant displayed higher basal motility and EGF-stimulated chemokinesis.This latter effect was, however, only partially-sensitive to PI3K inhibitors. In PTEN(-/-) cells, basal and EGF-stimulated phosphorylation of PKB was substantially increased, but the p110-dependency was variable between cell types. In MDA-MB 468s phosphorylation of PKB was significantly dependent on p110β, but not α- or δ- activity; in PTEN(-/-) MCF10a it remained, like the parental cells, p110α-dependent. Surprisingly, loss of PTEN suppressed basal motility and EGF-stimulated chemokinesis. These results indicate that; p110α is required for EGF signaling to PKB and chemokinesis, but not chemotaxis; onco-mutant alleles of p110α augment signaling in the absence of EGF and may increase motility, in part, via acutely modulating PI3K-activity-independent mechanisms. Finally, we demonstrate that there is not a universal mechanism that up-regulates p110β function in the absence of PTEN.