The mixed-lineage kinase DLK undergoes Src-dependent tyrosine phosphorylation and activation in cells exposed to vanadate or platelet-derived growth factor (PDGF).
Some data in the literature suggest that serine/threonine phosphorylation is required for activation of the mixed-lineage kinases (MLKs), a subgroup of mitogen-activated protein kinase kinase kinases (MAPKKKs). In this report, we demonstrate that the MLK family member DLK is activated and concurrently tyrosine-phosphorylated in cells exposed to the protein tyrosine phosphatase inhibitor vanadate. Tyrosine phosphorylation appears crucial for activation as incubation of vanadate-activated DLK molecules with a tyrosine phosphatase substantially reduced DLK enzymatic activity. Interestingly, the effects of vanadate on DLK are completely blocked by treatment with a Src family kinase inhibitor, PP2, or the expression of short hairpin RNA (shRNA) directed against Src. DLK also fails to undergo vanadate-stimulated tyrosine phosphorylation and activation in fibroblasts which lack expression of Src, Yes and Fyn, but reintroduction of wild-type Src or Fyn followed by vanadate treatment restores this response. In addition to vanadate, stimulation of cells with platelet-derived growth factor (PDGF) also induces tyrosine phosphorylation and activation of DLK by a Src-dependent mechanism. DLK seems important for PDGF signaling because its depletion by RNA interference substantially reduces PDGF-stimulated ERK and Akt kinase activation. Thus, our findings suggest that Src-dependent tyrosine phosphorylation of DLK may be important for regulation of its activity, and they support a role for DLK in PDGF signaling.
Daviau
Alex
A
Département de biologie, Faculté des sciences, Université de Sherbrooke, Sherbrooke, Québec, Canada J1K 2R1.
Di Fruscio
Marco
M
Blouin
Richard
R
D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism*; D000077214:Becaplermin; D019556:COS Cells / Q000187:drug effects / Q000201:enzymology; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002522:Chlorocebus aethiops; D016572:Cyclosporine / Q000494:pharmacology; D064550:Death-Associated Protein Kinases; D004789:Enzyme Activation / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006801:Humans; D051379:Mice; D041681:NIH 3T3 Cells / Q000187:drug effects / Q000201:enzymology; D019319:Okadaic Acid / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000737:chemistry; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D020574:Proto-Oncogene Proteins c-sis; D034622:RNA Interference; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000502:physiology; D014638:Vanadates / Q000494:pharmacology*; D019061:src-Family Kinases / Q000378:metabolism*
D051017:Apoptosis Regulatory Proteins; D010982:Platelet-Derived Growth Factor; D020574:Proto-Oncogene Proteins c-sis; D011993:Recombinant Fusion Proteins; D000077214:Becaplermin; D019319:Okadaic Acid; D019000:Phosphotyrosine; D014638:Vanadates; D016572:Cyclosporine; D019061:src-Family Kinases; D064550:Death-Associated Protein Kinases; D051057:Proto-Oncogene Proteins c-akt; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D017027:Protein Tyrosine Phosphatases