Subtle distinct regulations of late erythroid molecular events by PI3K/AKT-mediated activation of Spi-1/PU.1 oncogene autoregulation loop.
Spi-1/PU.1 oncogene is downregulated as proerythroblasts undergo terminal differentiation. Insertion of the Friend virus upstream of the Spi-1/PU.1 locus leads to the constitutive upregulation of Spi-1/PU.1, and a subsequent block in the differentiation of the affected erythroblasts. We have shown that sustained overexpression of Spi-1/PU.1 also inhibits the erythroid splicing of protein 4.1R exon 16, irrespective of chemical induction of differentiation. Here, we show a positive feedback loop that couples constitutive phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling to high expression of Spi-1/PU.1 in Friend erythroleukemia cells. Inhibition of PI3K/AKT results in Spi-1/PU.1 downregulation in a stepwise manner and induces cell differentiation. Chromatin immunoprecipitation assays further supported the positive autoregulatory effect of Spi-1/PU.1. Mutational analysis indicated that Ser41, but not Ser148, is necessary for Spi-1/PU.1-mediated repression of hemoglobin expression, whereas both Ser residues are required for Spi-1/PU.1 inhibition of the erythroid splicing event. We further show that inhibition of the erythroid transcriptional and splicing events are strictly dependent on distinct Spi-1/PU.1 phosphorylation modifications rather than Spi-1/PU.1 expression level per se. Our data further support the fact that Spi-1/PU.1 inhibits 4.1R erythroid splicing through two different pathways, and bring new insights into the extracellular signal impact triggered by erythropoietin on late erythroid regulatory program, including pre-mRNA splicing.
Breig
O
O
mRNA Metabolism in Normal and Pathological Cells, CGMC, CNRS, Université Lyon 1, Villeurbanne, France.
Théoleyre
O
O
Douablin
A
A
Baklouti
F
F
D000818:Animals; D001798:Blood Proteins / Q000235:genetics; D002454:Cell Differentiation / Q000187:drug effects; D045744:Cell Line, Tumor; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D041905:Erythroid Cells / Q000166:cytology / Q000378:metabolism*; D004921:Erythropoietin / Q000378:metabolism; D005091:Exons / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D005914:Globins / Q000235:genetics; D006706:Homeostasis / Q000187:drug effects; D051379:Mice; D008840:Microfilament Proteins; D009857:Oncogenes; D009928:Organ Specificity; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012326:RNA Splicing / Q000187:drug effects; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects; D000099232:Proto-Oncogene Protein Spi-1
D001798:Blood Proteins; D004121:Dimethyl Sulfoxide; D004921:Erythropoietin; D005914:Globins; D008840:Microfilament Proteins; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D012694:Serine; D015534:Trans-Activators; D000099232:Proto-Oncogene Protein Spi-1; C514956:Epb41 protein, mouse