title: Signaling via class IA Phosphoinositide 3-kinases (PI3K) in human, breast-derived cell lines.
abstract: 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.
authors.affiliation: The Babraham Institute, Babraham, Cambridge, United Kingdom.
authors.forename: Veronique
authors.initials: V
authors.lastname: Juvin
authors.forename: Mouhannad
authors.initials: M
authors.lastname: Malek
authors.forename: Karen E
authors.initials: KE
authors.lastname: Anderson
authors.forename: Carine
authors.initials: C
authors.lastname: Dion
authors.forename: Tamara
authors.initials: T
authors.lastname: Chessa
authors.forename: Charlotte
authors.initials: C
authors.lastname: Lecureuil
authors.forename: G John
authors.initials: GJ
authors.lastname: Ferguson
authors.forename: Sabina
authors.initials: S
authors.lastname: Cosulich
authors.forename: Phillip T
authors.initials: PT
authors.lastname: Hawkins
authors.forename: Len R
authors.initials: LR
authors.lastname: Stephens
mesh_terms: D002460:Cell Line; D045744:Cell Line, Tumor; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005260:Female; D006801:Humans; D051059:PTEN Phosphohydrolase / Q000235:genetics / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology
chemical_list: D004815:Epidermal Growth Factor; D019869:Phosphatidylinositol 3-Kinases; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human