scholarly journals Palmitoylation of multiple Src-family kinases at a homologous N-terminal motif

1994 ◽  
Vol 303 (3) ◽  
pp. 749-753 ◽  
Author(s):  
M Koegl ◽  
P Zlatkine ◽  
S C Ley ◽  
S A Courtneidge ◽  
A I Magee
Keyword(s):  
Tyrosine Kinases ◽  
Recombinant Baculovirus ◽  
Fatty Acid Analysis ◽  
Site Directed Mutagenesis ◽  
Protein Tyrosine Kinases ◽  
Protein Tyrosine ◽  
Cos Cells ◽  
Cysteine Motif ◽  
Lipid Fatty Acid ◽  
And Function

We have recently identified a novel N-terminal cysteine-containing motif which specifies the palmitoylation of several G-protein alpha-subunits [Parenti, Viganó, Newman, Milligan and Magee (1993) Biochem. J. 291, 349-353]. A related motif occurs at the N-terminus of members of the Src family of protein tyrosine kinases except for Src itself and Blk. We have investigated whether the Src, Fyn, Yes and Lck gene products are palmitoylated. Src was not labelled with [3H]palmitate when endogenously expressed in COS cells. In contrast, endogenous Yes immunoprecipitated from COS cells was palmitoylated. Fyn was palmitoylated in insect cells infected with a recombinant baculovirus and the palmitoylation was independent of protein synthesis, suggesting a dynamic turnover of this lipid. Fatty acid analysis indicated that most of the label was incorporated as palmitate. Lck was palmitoylated when expressed by transfection in COS cells. All of these protein tyrosine kinases were also detectably myristoylated in each of the systems tested. Experiments performed with mutants of Lck expressed by transfection in COS cells indicated that cysteines at positions 3 and 5 were both palmitoylation sites and that myristoylation was required for palmitoylation. To confirm that palmitoylation was occurring on cysteines in the N-terminal region of Fyn, site-directed mutagenesis was used to replace the cysteines at positions 3 and 6 with alanine. The resulting protein was not palmitoylated but was still myristoylated when expressed in COS cells. A glycine to alanine mutant at position 2 was also not palmitoylated, showing that myristoylation is a prerequisite for palmitoylation. Our data indicate that Src family members containing the N-terminal cysteine motif are indeed palmitoylated. By analogy with Ras, it is possible that palmitoylation may play an important role in the localization and function of Src family protein tyrosine kinases.

scholarly journals Palmitylation of an amino-terminal cysteine motif of protein tyrosine kinases p56lck and p59fyn mediates interaction with glycosyl-phosphatidylinositol-anchored proteins.

1993 ◽  
Vol 13 (10) ◽  
pp. 6385-6392 ◽  
Author(s):  
A M Shenoy-Scaria ◽  
L K Gauen ◽  
J Kwong ◽  
A S Shaw ◽  
D M Lublin
Keyword(s):  
Amino Acids ◽  
Cell Line ◽  
Tyrosine Kinases ◽  
Cell Activation ◽  
Mutational Analysis ◽  
Protein Tyrosine Kinases ◽  
Protein Tyrosine ◽  
Glycosyl Phosphatidylinositol ◽  
Amino Terminal ◽  
Cysteine Motif

Cross-linking of glycosyl-phosphatidylinositol (GPI)-anchored membrane proteins on T cells can trigger cell activation. We and others have shown an association between GPI-anchored proteins and the protein tyrosine kinases (PTKs) p56lck and p59fyn, suggesting a pathway for signaling through GPI-anchored proteins. Studies of decay-accelerating factor (DAF) or CD59 in either the C32 cell line or the HeLa cell line transfected with PTK cDNA demonstrated that the GPI-anchored proteins associated noncovalently with p56lck and p59fyn but not with p60src. Nonmyristylated versions of p56lck and p59fyn also failed to associate with the GPI-anchored proteins. Mutational analysis of the PTK demonstrated that the association with the GPI-anchored proteins mapped to the unique amino-terminal domains of the PTK. A chimeric PTK consisting of the 10 amino-terminal residues of p56lck or p59fyn replacing the corresponding amino acids in p60src was sufficient for association with DAF, but the converse constructs containing the first 10 amino acids of p60src plus the remainder of p56lck or p59fyn did not associate with DAF. Mutation of cysteine to serine at positions 3 and 6 in p59fyn or positions 3 and 5 in p56lck abolished the association of these kinases with DAF. Mutation of serine to cysteine at positions 3 and 6 in p60src conferred on p60src the ability to associate with DAF. Direct labeling with [3H]palmitate demonstrated palmitylation of this amino-terminal cysteine motif in p56lck. Thus, palmitylation of the amino-terminal cysteine residue(s) together with myristylation of the amino-terminal glycine residue defines important motifs for the association of PTKs with GPI-anchored proteins.

Regulation of ion channels by protein tyrosine phosphorylation

2001 ◽  
Vol 281 (5) ◽  
pp. H1835-H1862 ◽  
Author(s):  
Michael J. Davis ◽  
Xin Wu ◽  
Timothy R. Nurkiewicz ◽  
Junya Kawasaki ◽  
Peichun Gui ◽  
...  
Keyword(s):  
Ion Channels ◽  
Tyrosine Phosphorylation ◽  
Tyrosine Kinases ◽  
Protein Tyrosine Kinases ◽  
Scaffolding Proteins ◽  
Ion Channel Regulation ◽  
Protein Tyrosine ◽  
Channel Regulation ◽  
Nonselective Cation Channels ◽  
And Function

Ion channels are regulated by protein phosphorylation and dephosphorylation of serine, threonine, and tyrosine residues. Evidence for the latter process, tyrosine phosphorylation, has increased substantially since this topic was last reviewed. In this review, we present a comprehensive summary and synthesis of the literature regarding the mechanism and function of ion channel regulation by protein tyrosine kinases and phosphatases. Coverage includes the majority of voltage-gated, ligand-gated, and second messenger-gated channels as well as several types of channels that have not yet been cloned, including store-operated Ca2+ channels, nonselective cation channels, and epithelial Na+ and Cl− channels. Additionally, we discuss the critical roles that channel-associated scaffolding proteins may play in localizing protein tyrosine kinases and phosphatases to the vicinity of ion channels.

scholarly journals Palmitylation of an amino-terminal cysteine motif of protein tyrosine kinases p56lck and p59fyn mediates interaction with glycosyl-phosphatidylinositol-anchored proteins

1993 ◽  
Vol 13 (10) ◽  
pp. 6385-6392
Author(s):  
A M Shenoy-Scaria ◽  
L K Gauen ◽  
J Kwong ◽  
A S Shaw ◽  
D M Lublin
Keyword(s):  
Amino Acids ◽  
Cell Line ◽  
Tyrosine Kinases ◽  
Cell Activation ◽  
Mutational Analysis ◽  
Protein Tyrosine Kinases ◽  
Protein Tyrosine ◽  
Glycosyl Phosphatidylinositol ◽  
Amino Terminal ◽  
Cysteine Motif

Cross-linking of glycosyl-phosphatidylinositol (GPI)-anchored membrane proteins on T cells can trigger cell activation. We and others have shown an association between GPI-anchored proteins and the protein tyrosine kinases (PTKs) p56lck and p59fyn, suggesting a pathway for signaling through GPI-anchored proteins. Studies of decay-accelerating factor (DAF) or CD59 in either the C32 cell line or the HeLa cell line transfected with PTK cDNA demonstrated that the GPI-anchored proteins associated noncovalently with p56lck and p59fyn but not with p60src. Nonmyristylated versions of p56lck and p59fyn also failed to associate with the GPI-anchored proteins. Mutational analysis of the PTK demonstrated that the association with the GPI-anchored proteins mapped to the unique amino-terminal domains of the PTK. A chimeric PTK consisting of the 10 amino-terminal residues of p56lck or p59fyn replacing the corresponding amino acids in p60src was sufficient for association with DAF, but the converse constructs containing the first 10 amino acids of p60src plus the remainder of p56lck or p59fyn did not associate with DAF. Mutation of cysteine to serine at positions 3 and 6 in p59fyn or positions 3 and 5 in p56lck abolished the association of these kinases with DAF. Mutation of serine to cysteine at positions 3 and 6 in p60src conferred on p60src the ability to associate with DAF. Direct labeling with [3H]palmitate demonstrated palmitylation of this amino-terminal cysteine motif in p56lck. Thus, palmitylation of the amino-terminal cysteine residue(s) together with myristylation of the amino-terminal glycine residue defines important motifs for the association of PTKs with GPI-anchored proteins.

scholarly journals Haemophilus ducreyi LspA Proteins Are Tyrosine Phosphorylated by Macrophage-Encoded Protein Tyrosine Kinases

2008 ◽  
Vol 76 (10) ◽  
pp. 4692-4702 ◽  
Author(s):  
Kaiping Deng ◽  
Jason R. Mock ◽  
Steven Greenberg ◽  
Nicolai S. C. van Oers ◽  
Eric J. Hansen
Keyword(s):  
Tyrosine Phosphorylation ◽  
Tyrosine Kinases ◽  
Immune Cell ◽  
Kinase Inhibitor ◽  
Divalent Cations ◽  
Site Directed Mutagenesis ◽  
Haemophilus Ducreyi ◽  
Protein Tyrosine Kinases ◽  
Protein Tyrosine

ABSTRACTThe LspA proteins (LspA1 and LspA2) ofHaemophilus ducreyiare necessary for this pathogen to inhibit the phagocytic activity of macrophage cell lines, an event that can be correlated with a reduction in the level of active Src family protein tyrosine kinases (PTKs) in these eukaryotic cells. During studies investigating this inhibitory mechanism, it was discovered that the LspA proteins themselves were tyrosine phosphorylated after wild-typeH. ducreyicells were incubated with macrophages. LspA proteins in cell-free concentratedH. ducreyiculture supernatant fluid could also be tyrosine phosphorylated by macrophages. This ability to tyrosine phosphorylate the LspA proteins was not limited to immune cell lineages but could be accomplished by both HeLa and COS-7 cells. Kinase inhibitor studies with macrophages demonstrated that the Src family PTKs were required for this tyrosine phosphorylation activity. In silico methods and site-directed mutagenesis were used to identify EPIYG and EPVYA motifs in LspA1 that contained tyrosines that were targets for phosphorylation. A total of four tyrosines could be phosphorylated in LspA1, with LspA2 containing eight predicted tyrosine phosphorylation motifs. Purified LspA1 fusion proteins containing either the EPIYG or EPVYA motifs were shown to be phosphorylated by purified Src PTK in vitro. Macrophage lysates could also tyrosine phosphorylate the LspA proteins and an LspA1 fusion protein via a mechanism that was dependent on the presence of both divalent cations and ATP. Several motifs known to interact with or otherwise affect eukaryotic kinases were identified in the LspA proteins.

Naturally Occurring Homoisoflavonoids as Potent Protein Tyrosine Kinases Inhibitors

Planta Medica ◽  
2008 ◽  
Vol 74 (03) ◽  
Author(s):  
Y Ye ◽  
LG Lin ◽  
H Xie ◽  
HL Li ◽  
HL Jiang ◽  
...  
Keyword(s):  
Tyrosine Kinases ◽  
Protein Tyrosine Kinases ◽  
Protein Tyrosine ◽  
Naturally Occurring
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