title: Regulation of Hoxb2 by APL-associated PLZF protein.
abstract: The PLZF gene is translocated in a subset of all-trans-retinoic acid resistant acute promyelocytic leukaemia (APL) cases, encodes a DNA binding transcription factor and is expressed highly in haematopoietic progenitor cells as well-developing central nervous system (CNS). The spatially restricted and temporally dynamic pattern of PLZF expression in the developing CNS suggested that it might play a role in the circuitry regulating hindbrain segmentation. We have now identified a PLZF binding site (PLZF-RE) in an enhancer region of Hoxb2 that itself is required for directing high-level expression in rhombomers 3 and 5 of the developing hindbrain. The wild-type r3/r5 enhancer linked to a heterologous promoter was responsive to regulation by PLZF, and this activity was lost in variants containing a mutated PLZF-RE. Compared with the wild-type protein, the binding of the APL-associated reciprocal RARalpha-PLZF fusion to PLZF-RE was much stronger, suggesting that the N-terminal PLZF sequences missing from the fusion may play a role in the regulation of DNA binding. Consistent with this, the N-terminal POZ domain was required for cooperative binding of PLZF to a multimerized PLZF-RE. In the context of the r3/r5 enhancer, the PLZF-RE cooperated for PLZF binding with an additional A/T-rich motif positioned downstream of the PLZF-RE. This A/T motif was previously shown to be essential for the regulation of Hoxb2 expression in r3 and r5 in cooperation with another Krüppel-like zinc finger protein Krox 20. The presence of both the PLZF-RE and the A/T-rich motif was required for a maximal effect of PLZF on a heterologous promoter and was essential in vivo to direct the expression of a lacZ reporter in the chick neural tube. Hence, both PLZF and Krox20 cooperate with a common A/T motif in mediating in vivo activity of the Hoxb2 enhancer. Our findings indicate that Hoxb2 is a direct target for regulation by PLZF in the developing CNS and suggest that deregulation of Hox gene expression may contribute to APL pathogenesis.
authors.affiliation: Leukaemia Research Fund Centre at the Institute of Cancer Research, Chester Beatty Laboratories, Fulham Road, London SW3 6JB, UK.
authors.forename: Sarah
authors.initials: S
authors.lastname: Ivins
authors.forename: Kieran
authors.initials: K
authors.lastname: Pemberton
authors.forename: Fabien
authors.initials: F
authors.lastname: Guidez
authors.forename: Louise
authors.initials: L
authors.lastname: Howell
authors.forename: Robb
authors.initials: R
authors.lastname: Krumlauf
authors.forename: Arthur
authors.initials: A
authors.lastname: Zelent
mesh_terms: D000818:Animals; D001665:Binding Sites; D002642:Chick Embryo; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000502:physiology*; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation; D006410:Hematopoiesis; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D051741:Kruppel-Like Transcription Factors; D015473:Leukemia, Promyelocytic, Acute / Q000209:etiology; D000075722:Promyelocytic Leukemia Zinc Finger Protein; D018168:Receptors, Retinoic Acid / Q000502:physiology; D012097:Repressor Proteins / Q000502:physiology; D000072482:Retinoic Acid Receptor alpha; D012249:Rhombencephalon / Q000196:embryology; D014157:Transcription Factors / Q000235:genetics* / Q000502:physiology*
chemical_list: D004268:DNA-Binding Proteins; C086061:HOXB2 protein, human; D018398:Homeodomain Proteins; D051741:Kruppel-Like Transcription Factors; D000075722:Promyelocytic Leukemia Zinc Finger Protein; C000607071:RARA protein, human; D018168:Receptors, Retinoic Acid; D012097:Repressor Proteins; D000072482:Retinoic Acid Receptor alpha; D014157:Transcription Factors; C092037:ZBTB16 protein, human; D004247:DNA