Role of the protein tyrosine phosphatase SHP-1 (Src homology phosphatase-1) in the regulation of interleukin-3-induced survival, proliferation and signalling.
The tyrosine phosphatase SHP-1 (Src homology phosphatase-1) has been widely implicated as a negative regulator of signalling in immune cells. We have investigated in detail the role of SHP-1 in interleukin-3 (IL-3) signal transduction by inducibly expressing wild-type (WT), C453S (substrate-trapping) and R459M (catalytically inactive) forms of SHP-1 in the IL-3-dependent cell line BaF/3. Expression of WT SHP-1 had little impact on IL-3-induced proliferation, but enhanced apoptosis following IL-3 withdrawal. Expression of R459M SHP-1 increased the proliferative response of BaF/3 cells to IL-3 and increased cell survival at low doses of IL-3 and following IL-3 withdrawal. Investigation into the biochemical consequences resulting from expression of these SHP-1 variants demonstrated that the beta chain of the IL-3 receptor (Aic2A) was hypo-phosphorylated in cells expressing WT SHP-1 and hyper-phosphorylated in those expressing R459M SHP-1. Further, ectopic expression of the trapping mutant, C453S SHP-1, protected Aic2A from dephosphorylation, suggesting that Aic2A is a SHP-1 substrate in BaF/3 cells. Examination of overall levels of tyrosine phosphorylation demonstrated that they were not perturbed in these transfectants. Activation-specific phosphorylation of STAT (signal transducer and activator of transcription) 5a/b, protein kinase B and ERK (extracellular-signal-regulated kinase)-1 and -2 was also unaffected by expression of WT or R459M SHP-1. However, overall levels of IL-3-induced tyrosine phosphorylation of STAT5 were reduced upon expression of WT SHP-1 and increased when R459M SHP-1 was expressed, consistent with STAT5 being a potential SHP-1 substrate. These results demonstrate that SHP-1 acts to negatively regulate IL-3-driven survival and proliferation, potentially via regulation of tyrosine phosphorylation of Aic2A and STAT5.
Paling
Nicholas R D
NR
Department of Pharmacy & Pharmacology, University of Bath, Bath BA2 7AY, UK.
Welham
Melanie J
MJ
D000818:Animals; D002384:Catalysis; D002455:Cell Division; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D005786:Gene Expression Regulation; D015151:Immunoblotting; D007377:Interleukin-3 / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008894:Milk Proteins; D008928:Mitochondria / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010766:Phosphorylation; D011233:Precipitin Tests; D054645:Protein Phosphatase 1; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism* / Q000502:physiology*; D050799:STAT5 Transcription Factor; D015398:Signal Transduction; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism; D014162:Transfection; D014343:Trypan Blue / Q000494:pharmacology; D014443:Tyrosine / Q000378:metabolism
D004268:DNA-Binding Proteins; D007377:Interleukin-3; D047908:Intracellular Signaling Peptides and Proteins; D008894:Milk Proteins; D050799:STAT5 Transcription Factor; D015534:Trans-Activators; D014443:Tyrosine; D020928:Mitogen-Activated Protein Kinases; D054645:Protein Phosphatase 1; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; C509248:Ptpn6 protein, mouse; D014343:Trypan Blue