Proliferation and differentiation control in leukemias by lineage-specific transcription factors
Choe, Kevin S.
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Characteristic features of leukemias and many other cancers include both uncontrolled proliferation and a block to terminal differentiation. Although tremendous progress has been made in elucidating the mechanisms controlling proliferation and differentiation, much less is known about how these two cellular processes are coupled in cancer cells. Using the in vitro differentiation of murine erythroleukemia (MEL) cells, in which the leukemic blasts can be forced to resume their normal terminal differentiation program, we studied the relationships between cell cycle regulators and differentiation-promoting transcription factors.;The hematopoietic-specific transcription factor, PU.1 is required for normal development of myeloid and some lymphoid cells, and it is not expressed in erythroid cells. When PU.1 is abnormally activated in erythroid cells by viral insertion or transgenesis, erythropoiesis is blocked and erythroleukemia ensues. It was found in our laboratory that PU.1 inhibits erythroid differentiation by physically interacting with an erythroid-specific transcription factor, GATA-1 and repressing GATA-1-mediated transcription. We asked whether providing these MEL cells with additional GATA-1 can overcome the repression imposed by PU.1 and resume normal erythroid differentiation. We show here that exogenous expression of GATA-1 was sufficient to induce terminal erythroid differentiation, measured by hemoglobin production and proliferation arrest.;In addition to repressing the differentiation-promoting functions of GATA-1, PU.1 also plays a role in stimulating proliferation of MEL cells. We showed previously that during reentry of MEL cells into terminal differentiation, both PU.1 and cyclin dependent kinase 6 (CDK6) are coordinately down regulated, and that constitutive expression of either protein can block MEL cells from undergoing terminal differentiation. We show by in vitro kinase assays and two-dimensional gel electrophoresis that PU.1 is a novel substrate of CDK6 in vitro and in vivo. We also show that serine 126 of PU.1 is an important site of phosphorylation by CDK6 in vitro, and phosphorylation of the PU.1 protein leads to an increased protein stability and an accumulation in the cells. We also investigated whether the CDK6 gene is a transcriptional target of PU.1. Using chromatin immunoprecipitation assays and reporter transactivation assays, we show that PU.1 binds to the CDK6 promoter and stimulates its activity. (Abstract shortened by UMI.).
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