Cellular trafficking of phospholamban and formation of functional sarcoplasmic reticulum during myocyte differentiation.
Phospholamban (PLB) associates with the Ca(2+)-ATPase in sarcoplasmic reticulum (SR) membranes to permit the modulation of contraction in response to beta-adrenergic signaling. To understand how coordinated changes in the abundance and intracellular trafficking of PLB and the Ca(2+)-ATPase contribute to the maturation of functional muscle, we measured changes in abundance, location, and turnover of endogenous and tagged proteins in myoblasts and during their differentiation. We found that PLB is constitutively expressed in both myoblasts and differentiated myotubes, whereas abundance increases of the Ca(2+)-ATPase coincide with the formation of differentiated myotubes. We observed that PLB is primarily present in highly mobile vesicular structures outside the endoplasmic reticulum, irrespective of the expression of the Ca(2+)-ATPase, indicating that PLB targeting is regulated through vesicle trafficking. Moreover, using pulse-chase methods, we observed that in myoblasts, PLB is trafficked through directed transport through the Golgi to the plasma membrane before endosome-mediated internalization. The observed trafficking of PLB to the plasma membrane suggests an important role for PLB during muscle differentiation, which is distinct from its previously recognized role in the regulation of the Ca(2+)-ATPase.
Stenoien
David L
DL
Biological Sciences Division, Pacific Northwest National Laboratory, Richland, WA 99352, USA.
Knyushko
Tatyana V
TV
Londono
Monica P
MP
Opresko
Lee K
LK
Mayer
M Uljana
MU
Brady
Scott T
ST
Squier
Thomas C
TC
Bigelow
Diana J
DJ
D000818:Animals; D015415:Biomarkers; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation; D002460:Cell Line; D005786:Gene Expression Regulation; D049452:Green Fluorescent Proteins; D051379:Mice; D032342:Muscle Cells / Q000166:cytology* / Q000378:metabolism*; D021381:Protein Transport; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000378:metabolism; D000099211:Phospholamban
D015415:Biomarkers; D002135:Calcium-Binding Proteins; D049452:Green Fluorescent Proteins; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000099211:Phospholamban; C506944:Atp2a1 protein, mouse