Role of Nuclear Transport in Acute Leukemia

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DESCRIPTION (provided by applicant): The candidate is a physician-scientist recently appointed as a tenure-track Assistant Professor in the Department of Pathology at Northwestern University Medical School. His major focus (80 percent) will be basic research into nuclear transport and its role in leukemia; with the rest dedicated to clinical hematopathology and teaching. The nuclear pore protein (nucleoporin) Nup98 gene is a frequent target of chromosomal rearrangements in acute myelogenous leukemia (AML), a disease characterized by failure of myeloid differentiation. These rearrangements produce chimeric proteins, the best characterized of which is Nup98-HOXA9. T h e mechanism by which Nup98 chimeras cause leukemia is not known. Nucleoporins, along with karyopherins (importins), are part of the machinery that mediates nucleocytoplasmic transport. Myeloid differentiation is regulated by transcription factors (TFs) that need the nuclear transport machinery to reach their destinations inside the nucleus. Of these TFs, AML1 and RAR-alpha play a central role in both normal myeloid differentiation and AML. There is substantial evidence that a decrease and/or abnormal distribution of AML1 and RAR-alpha within the nucleus contribute to the pathogenesis of AML. Our hypothesis is that Nup98 gene rearrangements cause AML by disrupting the nuclear import of the transcription factors that mediate myeloid differentiation. Focusing on the effect of Nup98-HOXA9 on the nuclear trafficking of AML1 and RAR-alpha and myeloid differentiation, we will: (I) Delineate the role of Nup98 and karyopherins in the nuclear import of AML1 and RAR-alpha. (ii) Determine whether Nup98-HOXA9 disrupts this process by monitoring the nuclear import, localization, and activity of AML1 and RAR- alpha in cells stably transfected with an inducible Nup98-HOXA9 construct. Our hypothesis predicts reduced nuclear import of AML1 and RAR-alpha and/or alteration in their intranuclear distribution and transcriptional activity upon induction of Nup98-HOXA9. (iii) Determine the effect of Nup98-HOXA9 on myeloid-specific gene transcription during myeloid differentiation of HL60 cells. Myeloid differentiation is accompanied by increased transcription of some genes such as CD11b and CD18 and decreased transcription of others such as myeloperoxidase (MPO). Our hypothesis predicts that Nup98-HOXA9 would counteract these tendencies. We will monitor the effect of Nup98-HOXA9 during differentiation on endogenous and transiently transfected CD11b, CD18, and MPO promoters. Looking at entire promoters will provide a global assessment of the effects of Nup98-HOXA9 on myeloid transcription.