title: Mitotic activation of the DISC1-inducible cyclic AMP phosphodiesterase-4D9 (PDE4D9), through multi-site phosphorylation, influences cell cycle progression.
abstract: In Rat-1 cells, the dramatic decrease in the levels of both intracellular cyclic 3'5' adenosine monophosphate (cyclic AMP; cAMP) and in the activity of cAMP-activated protein kinase A (PKA) observed in mitosis was paralleled by a profound increase in cAMP hydrolyzing phosphodiesterase-4 (PDE4) activity. The decrease in PKA activity, which occurs during mitosis, was attributable to PDE4 activation as the PDE4 selective inhibitor, rolipram, but not the phosphodiesterase-3 (PDE3) inhibitor, cilostamide, specifically ablated this cell cycle-dependent effect. PDE4 inhibition caused Rat-1 cells to move from S phase into G2/M more rapidly, to transit through G2/M more quickly and to remain in G1 for a longer period. Inhibition of PDE3 elicited no observable effects on cell cycle dynamics. Selective immunopurification of each of the four PDE4 sub-families identified PDE4D as being selectively activated in mitosis. Subsequent analysis uncovered PDE4D9, an isoform whose expression can be regulated by Disrupted-In-Schizophrenia 1 (DISC1)/activating transcription factor 4 (ATF4) complex, as the sole PDE4 species activated during mitosis in Rat-1 cells. PDE4D9 becomes activated in mitosis through dual phosphorylation at Ser585 and Ser245, involving the combined action of ERK and an unidentified 'switch' kinase that has previously been shown to be activated by H2O2. Additionally, in mitosis, PDE4D9 also becomes phosphorylated at Ser67 and Ser81, through the action of MK2 (MAPKAPK2) and AMP kinase (AMPK), respectively. The multisite phosphorylation of PDE4D9 by all four of these protein kinases leads to decreased mobility (band-shift) of PDE4D9 on SDS-PAGE. PDE4D9 is predominantly concentrated in the perinuclear region of Rat-1 cells but with a fraction distributed asymmetrically at the cell margins. Our investigations demonstrate that the diminished levels of cAMP and PKA activity that characterise mitosis are due to enhanced cAMP degradation by PDE4D9. PDE4D9, was found to locate primarily not only in the perinuclear region of Rat-1 cells but also at the cell margins. We propose that the sequestration of PDE4D9 in a specific complex together with AMPK, ERK, MK2 and the H2O2-activatable 'switch' kinase allows for its selective multi-site phosphorylation, activation and regulation in mitosis.
authors.affiliation: Institute of Neuroscience and Psychology, Wolfson Link and Davidson Buildings, University of Glasgow, University Avenue, Glasgow G12 8QQ, Scotland, UK.
authors.forename: Catherine L
authors.initials: CL
authors.lastname: Sheppard
authors.affiliation: Institute of Neuroscience and Psychology, Wolfson Link and Davidson Buildings, University of Glasgow, University Avenue, Glasgow G12 8QQ, Scotland, UK.
authors.forename: Louisa C Y
authors.initials: LC
authors.lastname: Lee
authors.affiliation: Institute of Neuroscience and Psychology, Wolfson Link and Davidson Buildings, University of Glasgow, University Avenue, Glasgow G12 8QQ, Scotland, UK.
authors.forename: Elaine V
authors.initials: EV
authors.lastname: Hill
authors.affiliation: Institute of Neuroscience and Psychology, Wolfson Link and Davidson Buildings, University of Glasgow, University Avenue, Glasgow G12 8QQ, Scotland, UK.
authors.forename: David J P
authors.initials: DJ
authors.lastname: Henderson
authors.affiliation: Institute of Neuroscience and Psychology, Wolfson Link and Davidson Buildings, University of Glasgow, University Avenue, Glasgow G12 8QQ, Scotland, UK.
authors.forename: Diana F
authors.initials: DF
authors.lastname: Anthony
authors.affiliation: Institute of Neuroscience and Psychology, Wolfson Link and Davidson Buildings, University of Glasgow, University Avenue, Glasgow G12 8QQ, Scotland, UK.
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Houslay
authors.affiliation: Institute of Neuroscience and Psychology, Wolfson Link and Davidson Buildings, University of Glasgow, University Avenue, Glasgow G12 8QQ, Scotland, UK.
authors.forename: Krishna C
authors.initials: KC
authors.lastname: Yalla
authors.affiliation: Institute of Neuroscience and Psychology, Wolfson Link and Davidson Buildings, University of Glasgow, University Avenue, Glasgow G12 8QQ, Scotland, UK.
authors.forename: Lynne S
authors.initials: LS
authors.lastname: Cairns
authors.affiliation: Institute of Neuroscience and Psychology, Wolfson Link and Davidson Buildings, University of Glasgow, University Avenue, Glasgow G12 8QQ, Scotland, UK.
authors.forename: Allan J
authors.initials: AJ
authors.lastname: Dunlop
authors.affiliation: Institute of Cardiovascular and Medical Sciences, Wolfson Link and Davidson Buildings, University of Glasgow, University Avenue, Glasgow G12 8QQ, Scotland, UK.
authors.forename: George S
authors.initials: GS
authors.lastname: Baillie
authors.affiliation: Institute of Pharmaceutical Science, King's College London, 5th Floor, Franklin-Wilkins Building, 150 Stamford Street, London SE1 9NH, UK.
authors.forename: Elaine
authors.initials: E
authors.lastname: Huston
authors.affiliation: Institute of Pharmaceutical Science, King's College London, 5th Floor, Franklin-Wilkins Building, 150 Stamford Street, London SE1 9NH, UK. Electronic address: miles.houslay@kcl.ac.uk.
authors.forename: Miles D
authors.initials: MD
authors.lastname: Houslay
mesh_terms: D051701:Activating Transcription Factor 4 / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4 / Q000378:metabolism*; D004789:Enzyme Activation; D007399:Interphase; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D008938:Mitosis; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D010766:Phosphorylation; D020033:Protein Isoforms / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D051381:Rats; D000099190:MAP-Kinase-Activated Kinase 2
chemical_list: D051701:Activating Transcription Factor 4; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4; D047908:Intracellular Signaling Peptides and Proteins; D009419:Nerve Tissue Proteins; D020033:Protein Isoforms; D017346:Protein Serine-Threonine Kinases; D000099190:MAP-Kinase-Activated Kinase 2; C497110:Atf4 protein, rat; C498795:Disc1 protein, rat; C528468:PDE4D protein, rat
keywords: AMPK; DISC1; MK2; Mitosis; PDE4; PDE4D9