Hematopoietic cell phosphatase negatively regulates erythropoietin-induced hemoglobinization in erythroleukemic SKT6 cells.
In an increasing number of hematopoietic cytokine receptor systems (T-cell receptor, B-cell receptor, and macrophage colony-stimulating factor, stem cell factor, interleukin-3, and erythropoietin [EPO] receptors), inhibitory roles for the protein tyrosine phosphatase hematopoietic cell phosphatase (HCP; SHPTP1, PTP1C, and SHP1) have been defined in proliferative signaling. However, evidence exists to suggest that HCP also may exert important effects on blood cell differentiation. To investigate possible roles for HCP during late erythroid differentiation, effects of manipulating HCP expression or recruitment on EPO-induced hemoglobinization in erythroleukemic SKT6 cells have been investigated. No effects of EPO on levels of HCP, Syp, Stat5, the EPO receptor, or GATA-1 expression were observed during induced differentiation. However, the tyrosine phosphorylation of JAK2, the EPO receptor, and Stat5 was efficiently activated, and HCP was observed to associate constitutively with the EPO receptor in this differentiation-specific system. In studies of HCP function, inhibition of HCP expression by antisense oligonucleotides enhanced hemoglobinization, whereas the enforced ectopic expression of wild-type (wt) HCP markedly inhibited EPO-induced globin expression and Stat5 activation. Based on these findings, epidermal growth factor (EGF) receptor/EPO receptor chimeras containing either the wt EPO receptor cytoplasmic domain (EECA) or a derived HCP binding site mutant (EECA-Y429,431F) were expressed in SKT6 cells, and their abilities to mediate differentiation were assayed. Each chimera supported EGF-induced hemoglobinization, but efficiencies for EECA-Y429,431F were enhanced 400% to 500%. Thus, these studies show a novel role for HCP as a negative regulator of EPO-induced erythroid differentiation. In normal erythroid progenitor cells, HCP may act to prevent premature commitment to terminal differentiation. In erythroleukemic SKT6 cells, this action also may enforce mitogenesis.
Sharlow
E R
ER
Graduate Program in Genetics, Department of Veterinary Science, Amgen, Inc, Thousand Oaks, CA, USA.
Pacifici
R
R
Crouse
J
J
Batac
J
J
Todokoro
K
K
Wojchowski
D M
DM
D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D066246:ErbB Receptors / Q000737:chemistry; D050981:Erythroid-Specific DNA-Binding Factors; D004920:Erythropoiesis; D004921:Erythropoietin / Q000494:pharmacology*; D050982:GATA1 Transcription Factor; D005914:Globins / Q000235:genetics; D006454:Hemoglobins / Q000096:biosynthesis*; D047908:Intracellular Signaling Peptides and Proteins; D053614:Janus Kinase 2; D004915:Leukemia, Erythroblastic, Acute / Q000503:physiopathology; D051379:Mice; D008894:Milk Proteins; D016376:Oligonucleotides, Antisense; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D011518:Proto-Oncogene Proteins; D017467:Receptors, Erythropoietin / Q000235:genetics; D011993:Recombinant Fusion Proteins; D050799:STAT5 Transcription Factor; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics; D014162:Transfection; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D050982:GATA1 Transcription Factor; C494689:Gata1 protein, mouse; D006454:Hemoglobins; D047908:Intracellular Signaling Peptides and Proteins; D008894:Milk Proteins; D016376:Oligonucleotides, Antisense; D011518:Proto-Oncogene Proteins; D017467:Receptors, Erythropoietin; D011993:Recombinant Fusion Proteins; D050799:STAT5 Transcription Factor; D015534:Trans-Activators; D014157:Transcription Factors; D004921:Erythropoietin; D019000:Phosphotyrosine; D005914:Globins; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; C507926:Jak2 protein, mouse; D053614:Janus Kinase 2; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; C516725:Ptpn11 protein, mouse; C509248:Ptpn6 protein, mouse