A phosphorylation site in the Na+ channel required for modulation by protein kinase C

In Chinese hamster ovary (CHO) fibroblast cells the protein kinase C activating phorbol ester, phorbol myristate acetate (PMA), stimulates an increase in cell surface transferrin receptor (TR) expression by increasing the exocytic rate of the recycling pathway. The human TR expressed in CHO cells is similarly affected by PMA treatment. A mutant human TR in which the major protein kinase C phosphorylation site, serine 24, has been replaced with the non-phosphorylatable amino acid glycine has been constructed to investigate the role of receptor phosphorylation in the PMA induced up-regulation. The Gly-24-substituted receptor binds, internalizes, and recycles Tf. Furthermore, the altered receptor mediates cellular Fe accumulation from diferric-Tf, thereby fulfilling the receptor's major biological role. The Gly-24 TR behaves identically to the wild-type TR when cells are treated with PMA. Therefore, Ser-24 phosphorylation is not required for the PMA-induced redistribution of the human TR expressed in CHO cells. The increased TR expression on the cell surface after PMA treatment results from an increase in the rate of exocytosis of the recycling receptors. No change in the endocytic rate or the size of the recycling receptor pool was observed. These results indicate that the PMA effect on the TR surface expression may result from a more general perturbation of membrane trafficking rather than a specific modulation of the TR.

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Expression Of BKCa Channels In Pulmonary Arterial Smooth Muscle And The Effect Of Protein Kinase C Phosphorylation Site S1076 On The Response To PKCµ On BKCa Channel Activity

10.1164/ajrccm-conference.2010.181.1_meetingabstracts.a6449 ◽

2010 ◽

Author(s):

Shu Zhu ◽

Richard E. White ◽

Mary L. Meadows ◽

Scott A. Barman

Keyword(s):

Smooth Muscle ◽

Protein Kinase C ◽

Protein Kinase ◽

Phosphorylation Site ◽

Bkca Channel ◽

Channel Activity ◽

Bkca Channels ◽

Kinase C ◽

Pulmonary Arterial ◽

Pulmonary Arterial Smooth Muscle

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A Common Phosphorylation Site for Cyclic AMP-dependent Protein Kinase and Protein Kinase C in Human Placental 6-Phosphofructo-2-kinase/Fructose-2,6-bisphosphatase

Bioscience Biotechnology and Biochemistry ◽

10.1271/bbb.62.2039 ◽

1998 ◽

Vol 62 (10) ◽

pp. 2039-2042 ◽

Cited By ~ 16

Author(s):

Noriko OKAMURA ◽

Ryuzo SAKAKIBARA

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Cyclic Amp ◽

Phosphorylation Site ◽

Dependent Protein Kinase ◽

Kinase C ◽

Dependent Protein

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The Acidic Region and Conserved Putative Protein Kinase C Phosphorylation Site in Nef Are Important for SIV Replication in Rhesus Macaques

Virology ◽

10.1006/viro.1999.9645 ◽

1999 ◽

Vol 257 (1) ◽

pp. 138-155 ◽

Cited By ~ 14

Author(s):

Silke Carl ◽

A.John Iafrate ◽

Sabine M. Lang ◽

Christiane Stahl-Hennig ◽

Eva M. Kuhn ◽

...

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Rhesus Macaques ◽

Phosphorylation Site ◽

Putative Protein ◽

Kinase C ◽

Acidic Region ◽

Putative Protein Kinase

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Testase 1 (ADAM 24) a plasma membrane-anchored sperm protease implicated in sperm function during epididymal maturation or fertilization

Journal of Cell Science ◽

10.1242/jcs.114.9.1787 ◽

2001 ◽

Vol 114 (9) ◽

pp. 1787-1794 ◽

Cited By ~ 1

Author(s):

G.Z. Zhu ◽

D.G. Myles ◽

P. Primakoff

Keyword(s):

Plasma Membrane ◽

Protein Kinase C ◽

Protein Kinase ◽

Acrosome Reaction ◽

Phosphorylation Site ◽

Equatorial Region ◽

Proteolytic Processing ◽

Mature Sperm ◽

Kinase C

Plasma membrane-anchored proteases have key roles in cell signaling, migration and refashioning the cell surface and its surroundings. We report the first example of a plasma membrane-anchored protease on mature sperm, testase 1 (ADAM 24). Unlike other studied sperm ADAMs (fertilin (α) and (β), cyritestin) whose metalloprotease domains are removed during sperm development, we found testase 1 retains an active metalloprotease domain, suggesting it acts as a protease on mature sperm. Testase 1 is a glycoprotein (molecular mass 88 kDa), localized to the equatorial region of the plasma membrane of cauda epididymal sperm. Typically, proteolytic removal of the pro-domain is an initial activation step for ADAM proteases. The pro-domain of the testase 1 precursor (108 kDa) is proteolytically removed as sperm transit the caput epididymis to produce processed (mature) testase 1 (88 kDa). Testase 1 is unique among all studied ADAMs in that its proteolytic processing occurs on the sperm plasma membrane instead of at an intracellular site (the Golgi). Using GST-fusion proteins and a synthetic testase 1 C-terminal peptide, we found that the cytoplasmic tail of testase 1 could be phosphorylated in vitro by protein kinase C (PKC). Thus testase 1 apparently has a cytoplasmic PKC phosphorylation site(s). Protein kinase C is known to stimulate other ADAMs' protease activity. Because events of the acrosome reaction include PKC activation, we speculate that testase 1 protease function could be important in sperm penetration of the zona pellucida after sperm PKC is activated during the acrosome reaction.

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Down-regulation of Na channel α-subunit mRNA by protein kinase C e in adrenal chromaffin cells

The Japanese Journal of Pharmacology ◽

10.1016/s0021-5198(19)45128-7 ◽

1997 ◽

Vol 73 ◽

pp. 157

Author(s):

Toshihiko Yanagita ◽

Hideyuki Kobayashi ◽

Keizou Masumoto ◽

Ryuichi Yamamoto ◽

Tomoaki Yuhi ◽

...

Keyword(s):

Protein Kinase C ◽

Protein Kinase ◽

Chromaffin Cells ◽

Na Channel ◽

Adrenal Chromaffin Cells ◽

Down Regulation ◽

Α Subunit ◽

Subunit Mrna ◽

Kinase C ◽

Adrenal Chromaffin

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Invariant Leu Preceding Turn Motif Phosphorylation Site Controls the Interaction of Protein Kinase C with Hsp70

Journal of Biological Chemistry ◽

10.1074/jbc.m604076200 ◽

2006 ◽

Vol 281 (43) ◽

pp. 32461-32468 ◽

Cited By ~ 26

Author(s):

Tianyan Gao ◽

Alexandra C. Newton

Keyword(s):

Protein Kinase C ◽

Heat Shock ◽

Protein Kinase ◽

Phosphorylation Site ◽

Insoluble Fraction ◽

Down Regulation ◽

Kinase C ◽

Shock Proteins ◽

Invariant Residue ◽

Autophosphorylation Site

Heat shock proteins play important roles in regulating signal transduction in cells by associating with, and stabilizing, diverse signaling molecules, including protein kinases. Previously, we have shown that heat shock protein Hsp70 associates with protein kinase C (PKC) via an interaction that is triggered by dephosphorylation at the turn phosphorylation motif. Here we have identified an invariant residue in the carboxyl terminus of PKC that mediates the binding to Hsp70. Specifically, we show that Hsp70 binds to Leu (Leu-640) immediately preceding the conserved turn motif autophosphorylation site (Thr-641) in PKC βII. Co-immunoprecipitation experiments reveal that mutation of Leu-640 to Gly decreases the interaction of Hsp70 with PKC βII. This weakened interaction between Hsp70 and the mutant PKCs results in accumulation of dephosphorylated PKC in the detergent-insoluble fraction of cells. In addition, the Hsp70-binding mutant is considerably more sensitive to down-regulation compared with WT PKC: disruption of Hsp70 binding leads to accelerated dephosphorylation and enhanced ubiquitination of mutant PKC upon phorbol ester treatment. Last, pulse-chase experiments demonstrate that Hsp70 preferentially binds the species of mature PKC that has become dephosphorylated compared with the newly synthesized protein that has yet to be phosphorylated. Thus, Hsp70 binds a hydrophobic residue preceding the turn motif, protecting PKC from down-regulation and sustaining the signaling lifetime of the kinase.

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