Association of the p85 regulatory subunit of phosphatidylinositol 3-kinase with an essential erythropoietin receptor subdomain
| Autor: | TC He, H Zhuang, N Jiang, MD Waterfield, DM Wojchowski |
|---|---|
| Rok vydání: | 1993 |
| Předmět: |
Macromolecular Substances
Immunology Blotting Western Molecular Sequence Data Restriction Mapping macromolecular substances Moths Transfection Biochemistry Polymerase Chain Reaction Cell Line Epitopes Mice Phosphatidylinositol 3-Kinases Cytosol Receptors Erythropoietin Animals Amino Acid Sequence Erythropoietin DNA Primers Base Sequence Cell Biology Hematology Molecular Weight Kinetics Phosphotransferases (Alcohol Group Acceptor) Mutagenesis biological phenomena cell phenomena and immunity Baculoviridae |
| Zdroj: | Blood. 82(12) |
| ISSN: | 0006-4971 |
| Popis: | Using an active, HAI epitope-tagged form of the murine erythropoietin (EPO) receptor and via direct coimmunoprecipitation, the p85 regulatory subunit of phosphatidyl inositol-3 kinase (p85/PI3-K) is shown to associate with the EPO receptor in transfected FDC-P1 cell lines. Coimmunoprecipitation of p85 with epitope-tagged EPO receptors was observed initially in FDC-HER cells labeled metabolically with [32P]orthophosphate, and association of these factors was confirmed by Western analyses of receptor immunoprecipitates using p85 antiserum. Interestingly, this association occurred in the absence of ligand, and exposure of FDC-HER cells to EPO did not detectably affect levels of receptor-associated p85 or overall levels of p85 phosphorylation. However, EPO was observed to stimulated the rapid formation of phosphatidylinositol 32P-phosphate in FDC-HER and FDC-ER cells. Through baculovirus-mediated expression of epitope-tagged EPO receptor forms in SF9 cells, domains for p85 association were mapped. Analyses of receptor forms with cytosolic truncations and deletions delineated a candidate subdomain for p85 binding to an essential extended box-2 region (P329-E374; including a putative motif for SH2 binding, Y343LVL). These findings extend a mechanistic alignment between the EPO receptor and protein tyrosine kinase-encoding receptors that likewise activate PI3-K, and expand the importance of further defining pathways to PI3-K activation. |
| Databáze: | OpenAIRE |
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