Functional cloning and characterization of a novel nonhomeodomain protein that inhibits the binding of PBX1-HOX complexes to DNA

PBX1 is a homeodomain protein that functions in complexes with other homeod omain-containing proteins to regulate gene expression during developmental and/or differentiation processes. A yeast two hybrid screen of a fetal live r-hematopoietic cDNA library using PBX1a as bait led to the discovery of a novel non-homeodomain-containing protein that interacts with PBX1 as well a s PBX2 and PBX3. RNA analysis revealed it to be expressed in CD34(+) hemato poietic cell populations enriched in primitive progenitors, as is PBX1; sea rch of the expressed sequence tag data base indicated that it is also expre ssed in other early embryonic as well as adult tissues. The full-length cDN A encodes a 731-amino acid protein that has no significant homology to know n proteins. This protein that we have termed hematopoietic PBX-interacting protein (HPIP) is mainly localized in the cytosol and in small amounts in t he nucleus. The region of PBX that interacts with HPIP includes both the ho meodomain and immediate N-terminal flanking sequences. Strikingly, electrop horetic mobility shift assays revealed that HPIP inhibits the ability of PB X-HOX heterodimers to bind to target sequences. Moreover, HPIP strongly inh ibits the transactivation activity of ESA-PBX. Together these findings sugg est that HPIP is a new regulator of PBX function.