Erythropoietin induces sustained phosphorylation of STAT5 in primitive but not definitive erythrocytes generated from mouse embryonic stem cells.
OBJECTIVE
During embryonic development murine erythropoiesis occurs in two waves by producing first primitive erythroid cells (EryPs) and then definitive erythroid cells (EryDs). Erythropoietin (EPO) signaling is compared between EryPs and EryDs.


METHODS
We studied the EPO signaling in EryPs and EryDs using an embryonic stem-derived culture system, which can recapitulate this in vivo development process and has thus been used as a convenient in vitro model system of erythropoiesis.


RESULTS
We found that EPO induced sustained phosphorylation and nuclear translocation of signal transducer and activator of transcription 5 (STAT5) in EryPs but not EryDs. EryPs expressed dramatically higher amounts of EPO receptor compared with EryDs, indicating there was excessive signaling from the receptor upon EPO stimulation. In addition, reduced expression of tyrosine phosphatase, Src homology region 2 domain-containing phosphatase-1, and decreased total phosphatase activity in EryPs partly explain the persistent activation of STAT5. Nevertheless, Janus kinase 2 (JAK2) phosphorylation, which is essential for transduction of EPO signaling from the EPO receptor to STAT5, was observed in a transient but not a persistent manner. Inhibition of JAK activity resulted in partial suppression of transient phosphorylation of STAT5 and no suppression of sustained phosphorylation of STAT5.


CONCLUSIONS
This study presents a unique feature of EryPs, as this is the first known example of sustained activation of STAT5 in normal cells. Our results also imply the existence of a JAK2-independent pathway of EPO signaling to induce STAT5 activation.
Tsuji-Takayama
Kazue
K
Cell Biology Institute, Research Center, Hayashibara Biochemical Laboratories, Inc., Fujisaki, Okayama, Japan.
Otani
Takeshi
T
Inoue
Toshiya
T
Nakamura
Shuji
S
Motoda
Ryuichi
R
Kibata
Masayoshi
M
Orita
Kunzo
K
D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D004622:Embryo, Mammalian / Q000166:cytology / Q000502:physiology*; D004789:Enzyme Activation / Q000187:drug effects; D004912:Erythrocytes / Q000166:cytology / Q000502:physiology*; D004920:Erythropoiesis / Q000187:drug effects / Q000502:physiology; D004921:Erythropoietin / Q000378:metabolism* / Q000494:pharmacology; D053614:Janus Kinase 2; D051379:Mice; D008954:Models, Biological; D010766:Phosphorylation / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D050799:STAT5 Transcription Factor / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013234:Stem Cells / Q000166:cytology / Q000502:physiology*
D011518:Proto-Oncogene Proteins; D050799:STAT5 Transcription Factor; D004921:Erythropoietin; D011505:Protein-Tyrosine Kinases; C507926:Jak2 protein, mouse; D053614:Janus Kinase 2