Cooperative Interactions between HOX and PBX Proteins Mediated by a Conserved Peptide Motif

Abstract

Homeoprotein products of theVertebrate embryos are patterned along multiple trunk and limb axes by the Hox gene complex (42). This family encodes homeodomain-containing transcription factors that confer positional identity through the differential regulation of target gene expression. The 38 Hox genes found in mice and humans are arranged in four genomic clusters, Hoxa to Hoxd, related to the HOM complex of Drosophila melanogaster. These complexes can be aligned such that more closely related genes (paralogs) occupy the same position in each complex, suggesting that they arose by duplications of an ancestral complex (42). Recent phylogenetic comparisons indicate that all animals share related Hox/HOM-type gene clusters (76). Expression of the Hox and HOM genes largely follows a colinear pattern, such that the more 3Ј a gene is located in a cluster, the more anteriorly it is expressed. Thus, more closely related HOX proteins from the same paralogous group are expressed in similar fashions. Paralogous genes have both distinct and overlapping functions manifested in their most anterior domain of expression (7,30).DNA binding by HOX and HOM proteins is mediated by the homeodomain, a conserved 60-amino-acid sequence that consists of a flexible N-terminal arm followed by three alpha helices (25). Base-specific contacts are made by the N-terminal arm and helix 3 via the minor and major grooves, respectively. Structural studies have demonstrated that even highly diverged homeodomains interact with DNA in a conserved fashion (83). HOX/HOM homeodomains can be divided into major classes based on sequence homology. This division also serves to classify the proteins according to their DNA-binding preferences. The majority of HOX/HOM homeodomains belong to the Antennapedia (Antp) class (HOX paralogous groups 2 to 8), which includes a number of proteins shown to bind and activate through elements of 6 to 9 bp containing a TAAT core motif. All of the Antp class homeodomains contain arginine at position 3 in the N-terminal arm, which contacts the second position in the TAAT core. Abdominal-B (Abd-B) class homeodomains have a lysine at position 3, and Abd-B exhibits a preference for a TTAT core (17). The members of the labial subfamily (HOX paralogous group 1) are unique since none of their homeodomains encode basic residues at positions 2 and 3. At least for HOXA-1, this results in a decreased affinity for DNA rather than an alteration in DNA-binding specificity (63). This may necessitate cooperation with a cofactor for site-specific DNA-binding by HOXA-1.Three other aspects of DNA-binding by the HOX/HOM family suggest that cofactor interactions may be generally employed. First, inspection of gene regulatory sequences reveals a high proportion of presumptive HOX binding sites, suggesting that not all of these sites are used. Second, most HOX or HOM proteins recognize similar binding sites with only modest preferences, suggesting that the HOX homeodomain alone could not effectively discriminate between targets (8, 13, 18, 61). Th...

HOXB4 homeodomain protein is expressed in developing epidermis and skin disorders and modulates keratinocyte proliferation

“…HOXB4 and other HOX proteins alone exhibit weak, relatively nonspecific DNA binding (Pellerin et al, 1994;Shen et al, 1996). They gain both binding avidity and specificity by forming heterodimeric DNA binding complexes with PBX proteins (Chan et al, 1994;Chang et al, 1995;Phelan et al, 1995;Lu and Kamps, 1996). The PBX1 gene was first identified as an oncogenic fusion between the PBX homeodomain and the activation domain of the E2A gene product (Kamps et al, 1990;Nourse et al, 1990).…”

Cellular proliferation and transformation induced by HOXB4 and HOXB3 proteins involves cooperation with PBX1

“…The conserved adenosines at position 3 and 7 (underlined) were shown to interact with the asparagine (Asn51) residue located in the Cterminal sequence of Pbx and Hox homeodomains (Mann and Chan, 1996). The non-conserved residue 5 and 6 (bold) is believed to contact the N-terminal region of Hox homeodomain, which presumably provides for the DNA-binding speci®city of HOX-PBX heterodimers Lu and Kamps, 1997;Phelan et al, 1995).…”

“…For HOX proteins from paralog groups 1 ± 8, a tryptophan within a conserved tetrapeptide (F/Y)PWM motif located N-terminal to the homeodomain mediates the HOX-PBX interactions (Neuteboom et al, 1995;Phelan et al, 1995;Shen et al, 1996). Although PBX alone is not capable of transforming cells, a region of 20 ± 25 conserved amino acids located C-terminal to the homeodomain, when fused to the E2A-activation domain, was recently found to be su cient for transformation.…”

The highest affinity DNA element bound by Pbx complexes in t(1;19) leukemic cells fails to mediate cooperative DNA-binding or cooperative transactivation by E2a-Pbx1 and Class I Hox proteins – evidence for selective targetting of E2a-Pbx1 to a subset of Pbx-recognition elements

“…Separation of the tetrameric cores by even a single base insertion disrupts complex formation for combinations of all Hox proteins with either E2a-Pbx1 or Pbx1 (Chang et al, 1996;Knoep¯er et al, 1996). Heterodimerization of Hox proteins with either Pbx1 or E2a-Pbx1 requires the Hox HD and a tryptophane-containing N-terminal peptide motif Chang et al, 1995;Phelan et al, 1995;Knoep¯er and Kamps, 1995;Neuteboom et al, 1995;Peers et al, 1995) and requires the Pbx HD and 17 C-terminal residues, which are retained in E2a-Pbx1 and are highly conserved among Pbx family members (Chang et al, 1995;Lu and Kamps, 1996). In these complexes, interaction of the Hox Nterminal peptide motif with either Pbx1 or E2a-Pbx1 alters the DNA-binding speci®city of the Hox protein at position 2 of its four base-pair core, from TAAT to either TGAT or TTAT (Chang et al, 1996;Lu and Kamps, 1997); however, the Hox protein retains inherent DNAbinding speci®city for the dinucleotide GG, GA, TG, or TA 3' to the Hox core (Knoep¯er et al, 1996).…”