EMZF-1--A NOVEL GENE REQUIRED FOR EMBRYONIC HEMATOPOIESIS

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The hematopoietic system in mammals is composed of diverse cell types that perform distinct and specialized functions. These terminally differentiated cells are short-lived, formed from a pool of totipotent hematopoietic stem cells (HSC) whose presumptive origin is the mesodermal cells of the yolk sac blood islands. Although progress has been made in the identification of HSC activity from the adult bone marrow, no such homogeneous cell population has been identified, and no genetic markers exist to identify such cells. We hypothesize that transcripts unique to the earliest HSC will provide genetic lineage markers enabling us to trace and manipulate the development of the primordial HSC. Using a molecular genetic approach we have recently identified a novel cDNA clone from a 9.5 day blood island/yolk sac cDNA library using the human myeloid zinc finger (MZF)-1 cDNAproble. This newly identified cDNA clone, extraembryonic mesoderm zinc finger-1 (EM ZF-1), was shown to be expressed exclusively in the extraembryonic mesoderm of the yolk sac, ina the fetal liver and heart of developing embryos and in the bone marrow, spleen and thymus of adult mice suggesting a role in mammalian hematopoiesis. This gene is located on mouse chromosome 5 and contains 12 C2H2 Kruppel-like zinc finger motifs in two separate regions which are highly homologous to several of the finger domains in MZF-1. EMZF-1 also contains an alanine/arginine-rich region downstream from the zinc finger motif. Such a domain has been found in other zinc finger genes and appears to be important for transcriptional regulation. The EMZF-1 gene maps to human chromosome 7 in an area associated with myeloid and lymphoid leukemias. Interestingly, PLZF, a newly discovered gene identified as a fusion partner with retinoid acid receptor-alpha in a variant t (11;17) translocation associated with acute promyelocytic leukemia, is also homologous to EMZF-1. Thus, EMZF-1 is the third member of a newly formed hematopoietic-specific zinc finger gene family whose function appears to be in the control of hematopoietic differentiation. The functional requirement for the EMZF-1 gene product appears early in mouse development since antisense oligonucleotides specific for EMZF-1 abrogates hematopoiesis in 14.5 day fetal liver and adult bone marrow cultures in vitro. Finally, another cDNA clone demonstrating high cross reactivity to the EMZF-1 was identified. This clone, LM-1D has been partially sequenced and determined to be an independent cDNA. We propose to characterize the lineage-specific expression of EMZF-1, identify its genomic structure and promoter elements and to begin the analysis of its function in vivo. The potential of EMZF-1 and the highly related LM-1D clone as molecular markers of primordial HSCs will be explored in vitro and in vivo and their function in hematopoiesis and leukemigenesis elucidated.