ATRA Resolves the Differentiation Block in t(15;17) Myeloid Leukemia by Restoring PU.1 Expression.

The transcription factor PU.1 is crucial for hematopoiesis. pu.1 knockout mice lack mature neutrophils, macrophages, B- and T-cells. PU.1 regulates important taget genes, like the M-CSF-, the G-CSF-, and the GM-CSF receptor promoter. Heterozygous mutations of the PU.1 gene have been found in some patients with AML, but not in AML with the t(15;17) encoding the PML-RARA fusion protein. These facts let us to hypothesize that decreased expression of PU.1 might contribute to the block in myeloid differentiation in t(15;17) AML. We therefore screened 141 AML patients for PU.1 mRNA expression using real-time PCR technology. We detected that t(15;17) AML patients (n=18) had significantly less PU.1 mRNA than AML-M2 patients (n=35), and 16fold less PU.1 mRNA than mature neutrophils. Moreover, the amount of PML-RARA transcripts in APL patients was inversely correlated to PU.1 mRNA levels. Conditional expression of PML-RARA in U937 cells suppressed PU.1 mRNA and PU.1 protein. HT93A cells derived from a t(15;17) AML-M3 patient nearly lack PU.1 mRNA and protein expression, and binding activity to the PU.1 site in the M-CSF-Receptor promoter is suppressed. Treatment of HT93A cells with all-trans retinoic acid (ATRA) powerfully restores PU.1 mRNA and protein and induces neutrophilic differentiation. We identified a region of extending −122 bp upstream of the translation start site in the PU.1 promoter mediates ATRA-induced activation of the PU.1 promoter in HT93A cells. Finally, conditional expression of PU.1 in HT93A cells is sufficient to trigger neutrophilic differentiation. This is the first report demonstrating that PU.1 is a critical target of the PML-RARA protein and an early ATRA-responsive gene.