Basal endothelial nitric oxide synthase (eNOS) phosphorylation on Ser(1177) occurs in a stable microtubule- and tubulin acetylation-dependent manner.
To better understand the relationship between the subcellular compartmentalization of endothelial nitric oxide synthase (eNOS) and its function in endothelial cells, we addressed the roles of the microtubule network and of its dynamics in organizing Golgi-bound eNOS. We found that part of Golgi-bound eNOS localizes to the trans-Golgi network and/or to trans-Golgi network-derived vesicles and membrane tubules that are organized preferentially by stable microtubules. Also, while most of cellular eNOS was recovered in a detergent-resistant microtubule-enriched subcellular fraction, its recovery was impaired after total microtubule disassembly, but not after selective disassembly of dynamic microtubules or after microtubule stabilization. Basal eNOS phosphorylation on Ser(1177) further required the association of the trans-Golgi network to stable microtubules and was enhanced by microtubule stabilization. We finally show that the involvement of stable microtubules in basal eNOS phosphorylation involved alpha-tubulin acetylation. Microtubule-dependent organization of subcellular eNOS and control over its phosphorylation would thus be essential for endothelial cells to maintain their basal eNOS function.
Giustiniani
J
J
Laboratoire de Biochimie et Biologie Cellulaire, Univ. Paris-Sud 11, JE 2493, IFR141, Faculté de Pharmacie, Châtenay-Malabry, France.
Couloubaly
S
S
Baillet
A
A
Pourci
M-L
ML
Cantaloube
I
I
Fourniat
C
C
Paul
J-L
JL
Poüs
C
C
D000107:Acetylation / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D003902:Detergents / Q000737:chemistry; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D007425:Intracellular Membranes / Q000737:chemistry / Q000378:metabolism; D021962:Membrane Microdomains / Q000378:metabolism; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D015739:Nocodazole / Q000494:pharmacology; D017239:Paclitaxel / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects / Q000502:physiology; D010768:Phosphoserine / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D014404:Tubulin / Q000235:genetics / Q000378:metabolism*; D014471:Umbilical Veins / Q000166:cytology; D021601:trans-Golgi Network / Q000378:metabolism
D003902:Detergents; D018841:HSP90 Heat-Shock Proteins; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D011993:Recombinant Fusion Proteins; D014404:Tubulin; D010768:Phosphoserine; C012589:trichostatin A; D052250:Nitric Oxide Synthase Type III; D017239:Paclitaxel; D015739:Nocodazole