title: Nuclear alpha1-adrenergic receptors signal activated ERK localization to caveolae in adult cardiac myocytes.
abstract: We previously identified an alpha1-AR-ERK (alpha1A-adrenergic receptor-extracellular signal-regulated kinase) survival signaling pathway in adult cardiac myocytes. Here, we investigated localization of alpha1-AR subtypes (alpha1A and alpha1B) and how their localization influences alpha1-AR signaling in cardiac myocytes. Using binding assays on myocyte subcellular fractions or a fluorescent alpha1-AR antagonist, we localized endogenous alpha1-ARs to the nucleus in wild-type adult cardiac myocytes. To clarify alpha1 subtype localization, we reconstituted alpha1 signaling in cultured alpha1A- and alpha1B-AR double knockout cardiac myocytes using alpha1-AR-green fluorescent protein (GFP) fusion proteins. Similar to endogenous alpha1-ARs and alpha1A- and alpha1B-GFP colocalized with LAP2 at the nuclear membrane. alpha1-AR nuclear localization was confirmed in vivo using alpha1-AR-GFP transgenic mice. The alpha1-signaling partners Galphaq and phospholipase Cbeta1 also colocalized with alpha1-ARs only at the nuclear membrane. Furthermore, we observed rapid catecholamine uptake mediated by norepinephrine-uptake-2 and found that alpha1-mediated activation of ERK was not inhibited by a membrane impermeant alpha1-blocker, suggesting alpha1 signaling is initiated at the nucleus. Contrary to prior studies, we did not observe alpha1-AR localization to caveolae, but we found that alpha1-AR signaling initiated at the nucleus led to activated ERK localized to caveolae. In summary, our results show that nuclear alpha1-ARs transduce signals to caveolae at the plasma membrane in cardiac myocytes.
authors.affiliation: Cardiovascular Research Center, Sanford Research/USD, Sioux Falls, SD 57105, USA.
authors.forename: Casey D
authors.initials: CD
authors.lastname: Wright
authors.forename: Quanhai
authors.initials: Q
authors.lastname: Chen
authors.forename: Nichole L
authors.initials: NL
authors.lastname: Baye
authors.forename: Yuan
authors.initials: Y
authors.lastname: Huang
authors.forename: Chastity L
authors.initials: CL
authors.lastname: Healy
authors.forename: Sivakanthan
authors.initials: S
authors.lastname: Kasinathan
authors.forename: Timothy D
authors.initials: TD
authors.lastname: O'Connell
mesh_terms: D058668:Adrenergic alpha-1 Receptor Antagonists; D000317:Adrenergic alpha-Antagonists / Q000494:pharmacology; D000818:Animals; D002395:Catecholamines / Q000378:metabolism; D021941:Caveolae / Q000187:drug effects / Q000201:enzymology*; D002458:Cell Fractionation; D002467:Cell Nucleus / Q000187:drug effects / Q000201:enzymology*; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D018613:Microscopy, Confocal; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000201:enzymology*; D027701:Organic Cation Transport Proteins / Q000378:metabolism; D054799:Phospholipase C beta / Q000378:metabolism; D010766:Phosphorylation; D011224:Prazosin / Q000494:pharmacology; D018340:Receptors, Adrenergic, alpha-1 / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D000099254:Organic Cation Transporter 3
chemical_list: D058668:Adrenergic alpha-1 Receptor Antagonists; D000317:Adrenergic alpha-Antagonists; D002395:Catecholamines; D004268:DNA-Binding Proteins; D048049:Extracellular Signal-Regulated MAP Kinases; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D049452:Green Fluorescent Proteins; D008565:Membrane Proteins; D027701:Organic Cation Transport Proteins; D054799:Phospholipase C beta; D011224:Prazosin; D018340:Receptors, Adrenergic, alpha-1; D011993:Recombinant Fusion Proteins; D000099254:Organic Cation Transporter 3; C547714:Adra1a protein, mouse; C547717:Adra1b protein, mouse; C093413:lamina-associated polypeptide 2; C518034:Plcb1 protein, mouse
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