|
|
|
|
|
|
|
||
Article |
Department of Cell Biology and Physiology, University of Pittsburgh, Pittsburgh, Pennsylvania 15261
PDZ-domain–containing proteins such as PSD-95 have been implicated in the targeting and clustering of membrane proteins. Biochemical and immunohistochemical studies indicate that PSD-95 recognizes COOH-terminal S/TXV sequences present in Kv1 K+ channels. However, the effect of binding a PDZ domain on a target protein has not been studied in live cells. In the present study, a green fluorescent protein–Kv1.4 fusion protein is used to study the effect of PSD-95 on channel movement. Fluorescence recovery after photobleaching showed that PSD-95 can immobilize K+ channels in the plasma membrane in an all-or-none manner. Furthermore, time lapse imaging showed that channel clusters formed in the presence of PSD-95 are stable in size, shape, and position. As expected from previous reports, two green fluorescent protein–tagged COOH-terminal variants of Kv1.4,
15 and V655A, are not clustered by PSD-95. However, coexpression of PSD-95 with V655A, but not 15, leads to the appearance of PSD-95 immunoreactivity in the plasma membrane. Furthermore, fluorescence recovery after photobleaching studies show that V655A channels are immobilized by PSD-95. Thus, V655A channels can interact with PSD-95 in a manner that leads to channel immobilization, but not clustering. These experiments document for the first time that PSD-95 immobilizes target proteins. Additionally, the data presented here demonstrate that the structural requirements for protein clustering and immobilization by PSD-95 are distinct.
Key Words: green fluorescent protein • Kv1.4 • fluorescence recovery after photobleaching • channel immobilization
Abbreviations: EGFP, enhanced green fluorescent protein; FRAP, fluorescence recovery after photobleaching; wt, wild type
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Facebook 
Reddit 
Technorati 
Twitter What's this?
This article has been cited by other articles:
|
|
 |
|
  S. A. L. Correa, J. Muller, G. L. Collingridge, and N. V. Marrion Rapid endocytosis provides restricted somatic expression of a K+ channel in central neurons J. Cell Sci., November 15, 2009; 122(22): 4186 - 4194. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. Mckeown, M. P. Burnham, C. Hodson, and O. T. Jones Identification of an Evolutionarily Conserved Extracellular Threonine Residue Critical for Surface Expression and Its Potential Coupling of Adjacent Voltage-sensing and Gating Domains in Voltage-gated Potassium Channels J. Biol. Chem., October 31, 2008; 283(44): 30421 - 30432. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Abi-Char, S. El-Haou, E. Balse, N. Neyroud, R. Vranckx, A. Coulombe, and S. N. Hatem The anchoring protein SAP97 retains Kv1.5 channels in the plasma membrane of cardiac myocytes Am J Physiol Heart Circ Physiol, April 1, 2008; 294(4): H1851 - H1861. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. M. S. O'Connell, J. D. Whitesell, and M. M. Tamkun Localization and mobility of the delayed-rectifer K+ channel Kv2.1 in adult cardiomyocytes Am J Physiol Heart Circ Physiol, January 1, 2008; 294(1): H229 - H237. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Maguy, T. E. Hebert, and S. Nattel Involvement of lipid rafts and caveolae in cardiac ion channel function Cardiovasc Res, March 1, 2006; 69(4): 798 - 807. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. M. S. O'Connell and M. M. Tamkun Targeting of voltage-gated potassium channel isoforms to distinct cell surface microdomains J. Cell Sci., May 15, 2005; 118(10): 2155 - 2166. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Nesti, B. Everill, and A. D. Morielli Endocytosis as a Mechanism for Tyrosine Kinase-dependent Suppression of a Voltage-gated Potassium Channel Mol. Biol. Cell, September 1, 2004; 15(9): 4073 - 4088. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Cha, A. Kenworthy, R. Murtazina, and M. Donowitz The lateral mobility of NHE3 on the apical membrane of renal epithelial OK cells is limited by the PDZ domain proteins NHERF1/2, but is dependent on an intact actin cytoskeleton as determined by FRAP J. Cell Sci., July 1, 2004; 117(15): 3353 - 3365. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. M. Haggie, B. A. Stanton, and A. S. Verkman Increased Diffusional Mobility of CFTR at the Plasma Membrane after Deletion of Its C-terminal PDZ Binding Motif J. Biol. Chem., February 13, 2004; 279(7): 5494 - 5500. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
W. Wong and L. C. Schlichter Differential Recruitment of Kv1.4 and Kv4.2 to Lipid Rafts by PSD-95 J. Biol. Chem., January 2, 2004; 279(1): 444 - 452. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Pal, K. A. Hartnett, J. M. Nerbonne, E. S. Levitan, and E. Aizenman Mediation of Neuronal Apoptosis by Kv2.1-Encoded Potassium Channels J. Neurosci., June 15, 2003; 23(12): 4798 - 4802. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. I. Rokitskaya, E. A. Kotova, and Y. N. Antonenko Tandem Gramicidin Channels Cross-linked by Streptavidin J. Gen. Physiol., April 28, 2003; 121(5): 463 - 476. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. Panyi, M. Bagdany, A. Bodnar, G. Vamosi, G. Szentesi, A. Jenei, L. Matyus, S. Varga, T. A. Waldmann, R. Gaspar, et al. Colocalization and nonrandom distribution of Kv1.3 potassium channels and CD3 molecules in the plasma membrane of human T lymphocytes PNAS, March 4, 2003; 100(5): 2592 - 2597. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. Bowie, E. P Garcia, J. Marshall, S. F Traynelis, and G D. Lange Allosteric Regulation and Spatial Distribution of Kainate Receptors Bound to Ancillary Proteins J. Physiol., March 1, 2003; 547(2): 373 - 385. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Hattan, E. Nesti, T. G. Cachero, and A. D. Morielli Tyrosine Phosphorylation of Kv1.2 Modulates Its Interaction with the Actin-binding Protein Cortactin J. Biol. Chem., October 4, 2002; 277(41): 38596 - 38606. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Ottschytsch, A. Raes, D. Van Hoorick, and D. J. Snyders Obligatory heterotetramerization of three previously uncharacterized Kv channel alpha -subunits identified in the human genome PNAS, June 11, 2002; 99(12): 7986 - 7991. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
W. Wong, E. W. Newell, D. G. M. Jugloff, O. T. Jones, and L. C. Schlichter Cell Surface Targeting and Clustering Interactions between Heterologously Expressed PSD-95 and the Shal Voltage-gated Potassium Channel, Kv4.2 J. Biol. Chem., May 31, 2002; 277(23): 20423 - 20430. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Murata, P. D. Buckett, J. Zhou, M. Brunner, E. Folco, and G. Koren SAP97 interacts with Kv1.5 in heterologous expression systems Am J Physiol Heart Circ Physiol, December 1, 2001; 281(6): H2575 - H2584. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Li, K. Takimoto, and E. S. Levitan Surface Expression of Kv1 Channels Is Governed by a C-terminal Motif J. Biol. Chem., April 14, 2000; 275(16): 11597 - 11602. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. Brown Targeting of membrane transporters in renal epithelia: when cell biology meets physiology Am J Physiol Renal Physiol, February 1, 2000; 278(2): F192 - F201. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. G. M. Jugloff, R. Khanna, L. C. Schlichter, and O. T. Jones Internalization of the Kv1.4 Potassium Channel Is Suppressed by Clustering Interactions with PSD-95 J. Biol. Chem., January 14, 2000; 275(2): 1357 - 1364. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. B. Nehring, E. Wischmeyer, F. Doring, R. W. Veh, M. Sheng, and A. Karschin Neuronal Inwardly Rectifying K+ Channels Differentially Couple to PDZ Proteins of the PSD-95/SAP90 Family J. Neurosci., January 1, 2000; 20(1): 156 - 162. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E.-K. Yang, M. R. Alvira, E. S. Levitan, and K. Takimoto Kvbeta Subunits Increase Expression of Kv4.3 Channels by Interacting with Their C Termini J. Biol. Chem., February 9, 2001; 276(7): 4839 - 4844. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Breton, T. Wiederhold, V. Marshansky, N. N. Nsumu, V. Ramesh, and D. Brown The B1 Subunit of the H+ATPase Is a PDZ Domain-binding Protein. COLOCALIZATION WITH NHE-RF IN RENAL B-INTERCALATED CELLS J. Biol. Chem., June 9, 2000; 275(24): 18219 - 18224. [Abstract] [Full Text] [PDF]
|
|
|
|
