Phospholamban and sarcolipin are maintained in the endoplasmic reticulum by retrieval from the ER-Golgi intermediate compartment
Phospholamban and sarcolipin are maintained in the endoplasmic reticulum by retrieval from the ER-Golgi intermediate compartment
Objective
Phospholamban and sarcolipin are small transmembrane proteins that modulate cardiac contractility through their interaction with the sarcoplasmic reticulum (SR) calcium pumps (SERCAs). We have examined the hypothesis that phospholamban and sarcolipin are maintained in the SR by a process of retrieval from post-SR compartments and the role of their transmembrane domains in targeting.
Methods
Antibodies directed against phospholamban and protein markers of the endoplasmic reticulum/Golgi intermediate compartment (ERGIC) and the trans-Golgi were used in fluorescence microscopy studies of cultured human fetal cardiac myocytes. In addition, sarcolipin and phospholamban were tagged at the N-terminus with enhanced-green-fluorescent protein (EGFP) and expressed in COS 7 cells. The EGFP-tagged constructs were localised using fluorescence microscopy and cell fractionation. The length of the transmembrane domains of phospholamban and sarcolipin were extended and the effect on cellular location was also examined.
Results
In fetal cardiac myocytes phospholamban was located in the SR and the ERGIC, but did not migrate to the trans-Golgi network. Tagged-phospholamban and sarcolipin were located in the endoplasmic reticulum (ER) of COS 7 cells indicating that their targeting was unaffected by the EGFP tag. Significant proportions of the tagged phospholamban and sarcolipin were also located in the ERGIC but not in the trans-Golgi. Increasing the length of the transmembranous domains of EGFP-tagged phospholamban and sarcolipin resulted in their mis-targeting to the plasma membrane.
Conclusions
Phospholamban and sarcolipin are maintained in the SR/ER by a process that includes their retrieval from the ERGIC following their passage from the SR/ER into the ERGIC. The transmembrane domains of phospholamban and sarcolipin are involved in the retrieval process.
phospholamban, sarcolipin, targetting, endoplasmic reticulum, sarcoplasmic reticulum, retrieval, ERGIC, transmembrane domain
114-123
Butler, John
5132f0e2-f28d-412a-be9a-e97963b4c0ee
Lee, Anthony G.
0891914c-e0e2-4ee1-b43e-1b70eb072d8e
Wilson, David I.
1500fca1-7082-4271-95f4-691f1d1252a2
Spalluto, Cosma
6802ad50-bc38-404f-9a19-40916425183b
Hanley, Neil A.
bf03f7bb-f377-44fb-8344-0bb1ca8b2ef9
East, J. Malcolm
9fe7f794-1d89-4935-9a99-b831d786056e
2007
Butler, John
5132f0e2-f28d-412a-be9a-e97963b4c0ee
Lee, Anthony G.
0891914c-e0e2-4ee1-b43e-1b70eb072d8e
Wilson, David I.
1500fca1-7082-4271-95f4-691f1d1252a2
Spalluto, Cosma
6802ad50-bc38-404f-9a19-40916425183b
Hanley, Neil A.
bf03f7bb-f377-44fb-8344-0bb1ca8b2ef9
East, J. Malcolm
9fe7f794-1d89-4935-9a99-b831d786056e
Butler, John, Lee, Anthony G., Wilson, David I., Spalluto, Cosma, Hanley, Neil A. and East, J. Malcolm
(2007)
Phospholamban and sarcolipin are maintained in the endoplasmic reticulum by retrieval from the ER-Golgi intermediate compartment.
Cardiovascular Research, 74 (1), .
(doi:10.1016/j.cardiores.2007.01.006).
Abstract
Objective
Phospholamban and sarcolipin are small transmembrane proteins that modulate cardiac contractility through their interaction with the sarcoplasmic reticulum (SR) calcium pumps (SERCAs). We have examined the hypothesis that phospholamban and sarcolipin are maintained in the SR by a process of retrieval from post-SR compartments and the role of their transmembrane domains in targeting.
Methods
Antibodies directed against phospholamban and protein markers of the endoplasmic reticulum/Golgi intermediate compartment (ERGIC) and the trans-Golgi were used in fluorescence microscopy studies of cultured human fetal cardiac myocytes. In addition, sarcolipin and phospholamban were tagged at the N-terminus with enhanced-green-fluorescent protein (EGFP) and expressed in COS 7 cells. The EGFP-tagged constructs were localised using fluorescence microscopy and cell fractionation. The length of the transmembrane domains of phospholamban and sarcolipin were extended and the effect on cellular location was also examined.
Results
In fetal cardiac myocytes phospholamban was located in the SR and the ERGIC, but did not migrate to the trans-Golgi network. Tagged-phospholamban and sarcolipin were located in the endoplasmic reticulum (ER) of COS 7 cells indicating that their targeting was unaffected by the EGFP tag. Significant proportions of the tagged phospholamban and sarcolipin were also located in the ERGIC but not in the trans-Golgi. Increasing the length of the transmembranous domains of EGFP-tagged phospholamban and sarcolipin resulted in their mis-targeting to the plasma membrane.
Conclusions
Phospholamban and sarcolipin are maintained in the SR/ER by a process that includes their retrieval from the ERGIC following their passage from the SR/ER into the ERGIC. The transmembrane domains of phospholamban and sarcolipin are involved in the retrieval process.
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More information
Published date: 2007
Additional Information:
This paper describes the way in which the regulators of endoplasmic reticulum (ER) calcium pumps, phospholamban and sarcolipin, are maintained in the ER and prevented from entering the plasma membrane.
Keywords:
phospholamban, sarcolipin, targetting, endoplasmic reticulum, sarcoplasmic reticulum, retrieval, ERGIC, transmembrane domain
Identifiers
Local EPrints ID: 44691
URI: http://eprints.soton.ac.uk/id/eprint/44691
ISSN: 0008-6363
PURE UUID: 55c2abfd-ee82-4ef0-ac43-4b0765e3363b
Catalogue record
Date deposited: 16 Apr 2007
Last modified: 16 Mar 2024 03:24
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Contributors
Author:
Cosma Spalluto
Author:
Neil A. Hanley
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