title: Rough endoplasmic reticulum to junctional sarcoplasmic reticulum trafficking of calsequestrin in adult cardiomyocytes.
abstract: Cardiac calsequestrin (CSQ) is synthesized on rough endoplasmic reticulum (ER), but concentrates within the junctional sarcoplasmic reticulum (SR) lumen where it becomes part of the Ca(2+)-release protein complex. To investigate CSQ trafficking through biosynthetic/secretory compartments of adult cardiomyocytes, CSQ-DsRed was overexpressed in cultured cells and examined using confocal fluorescence microscopy. By 48h of adenovirus treatment, CSQ-DsRed fluorescence had specifically accumulated in perinuclear cisternae, where it co-localized with markers of rough ER. From rough ER, CSQ-DsRed appeared to traffic directly to junctional SR along a transverse (Z-line) pathway along which sec 23-positive (ER-exit) sites were enriched. In contrast to DsRed direct fluorescence that presumably reflected DsRed tetramer formation, both anti-DsRed and anti-CSQ immunofluorescence did not detect the perinuclear CSQ-DsRed protein, but labeled only junctional SR puncta. These putative CSQ-DsRed monomers, but not the fluorescent tetramers, were observed to traffic anterogradely over the course of a 48h overexpression from rough ER towards the cell periphery. We propose a new model of CSQ and junctional SR protein traffic in the adult cardiomyocyte, wherein CSQ traffics from perinuclear cisternae, along contiguous ER/SR lumens in cardiomyocytes as a mobile monomer, but is retained in junctional SR as a polymer.
authors.affiliation: Department of Physiology, Wayne State University, Detroit, MI 48201, USA.
authors.forename: Timothy P
authors.initials: TP
authors.lastname: McFarland
authors.forename: Michelle L
authors.initials: ML
authors.lastname: Milstein
authors.forename: Steven E
authors.initials: SE
authors.lastname: Cala
mesh_terms: D000818:Animals; D001145:Arrhythmias, Cardiac / Q000378:metabolism*; D002155:Calsequestrin / Q000378:metabolism*; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D018870:Endoplasmic Reticulum, Rough / Q000378:metabolism*; D015151:Immunoblotting; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D032383:Myocytes, Cardiac / Q000378:metabolism; D051381:Rats; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*
chemical_list: D002155:Calsequestrin