Defects of protein phosphatase 2A causes corticosteroid insensitivity in severe asthma.
BACKGROUND
Corticosteroid insensitivity is a major barrier of treatment for some chronic inflammatory diseases, such as severe asthma, but the molecular mechanism of the insensitivity has not been fully elucidated. The object of this study is to investigate the role of protein phosphate 2A (PP2A), a serine/threonine phosphatase, on corticosteroid sensitivity in severe asthma.


RESULTS
Corticosteroid sensitivity was determined by the dexamethasone ability to inhibit TNFα-induced IL-8 or LPS-induced TNFα production. PP2A expression, glucocorticoid receptor (GR) nuclear translocation defined as the nuclear/cytoplasmic GR ratio and phosphorylation of GR-Ser²²⁶, c-Jun N-terminal kinase 1 (JNK1) and PP2A were analysed by Western-blotting. Phosphatase activity was measured by fluorescence-based assay. Okadaic acid (OA), a PP2A inhibitor, reduced corticosteroid sensitivity with reduced GR nuclear translocation and increased GR phosphorylation in U937 monocytic cells. PP2A knockdown by RNA interference showed similar effects. IL-2/IL-4 treatment to U937 reduced corticosteroid sensitivity, and PP2A expression/activity. In peripheral blood mononuclear cells (PBMCs) from severe asthma, the PP2A expression and activity were significantly reduced with concomitant enhancement of PP2A(C)-Tyr³⁰⁷ phosphorylation compared with those in healthy volunteers. As the results, GR-Ser²²⁶ and JNK1 phosphorylation were increased. The expression and activity of PP2A were negatively correlated with phosphorylation levels of GR-Ser²²⁶. Furthermore, co-immunoprecipitation assay in U937 cells revealed that PP2A associated with GR and JNK1 and IL-2/IL-4 exposure caused dissociation of each molecule. Lastly, PP2A overexpression increased corticosteroid sensitivity in U937 cells.


CONCLUSIONS
PP2A regulates GR nuclear translocation and corticosteroid sensitivity possibly by dephosphorylation of GR-Ser²²⁶ via dephosphorylation of upstream JNK1. This novel mechanism will provide new insight for the development of new therapy for severe asthma.
Airway Disease Section, National Heart and Lung Institute, Imperial College London, London, United Kingdom.
Yoshiki
Y
Kobayashi
Nicolas
N
Mercado
Peter J
PJ
Barnes
Kazuhiro
K
Ito
D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000305:Adrenal Cortex Hormones / Q000494:pharmacology* / Q000627:therapeutic use; D001249:Asthma / Q000188:drug therapy* / Q000201:enzymology* / Q000378:metabolism / Q000473:pathology; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology; D004351:Drug Resistance; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D007376:Interleukin-2 / Q000494:pharmacology; D015847:Interleukin-4 / Q000494:pharmacology; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D048055:Mitogen-Activated Protein Kinase 8 / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D054648:Protein Phosphatase 2 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011965:Receptors, Glucocorticoid / Q000737:chemistry / Q000378:metabolism; D012694:Serine / Q000378:metabolism
D000305:Adrenal Cortex Hormones; D004791:Enzyme Inhibitors; D007376:Interleukin-2; D011965:Receptors, Glucocorticoid; D015847:Interleukin-4; D012694:Serine; D003907:Dexamethasone; D048055:Mitogen-Activated Protein Kinase 8; D054648:Protein Phosphatase 2