Cooperative dimerization of paired class homeo domains on DNA.

Abstract

Homeo domain-containing proteins mediate many transcriptional processes in eukaryotes. Because nearly all animal homeo proteins are believed to bind to short, highly related DNA sequences, the basis for their high specificity of action is not understood. We show that cooperative dimerization on palindromic DNA sequences can provide increased specificity to one of the three major classes of homeo domains, the Paired/Pax class. The 60-amino-acid homeo domains from this class contain sufficient information to bind cooperatively as homo-and heterodimers to palindromic DNA sequences; that is, the binding of one homeo domain molecule can increase the affinity of a second molecule by up to 300-fold. Different members of the Paired (Prd) class of homeo domains prefer different spacings between half-sites, as determined by the ninth amino acid residue of the recognition helix. In addition, this residue determines the identity of the base pairs at the center of the palindromic sites, as well as the magnitude of the cooperative interaction. The cooperative dimerization of homeo domains in the Prd class distinguishes them from other classes, whereas binding-site configuration and sequence specificity allow for distinctions within this class.

Transcriptional modulation of the anti-apoptotic protein BCL-XL by the paired box transcription factors PAX3 and PAX3/FKHR

“…Indeed, at positions 7458 to 7454, and +108 to +112 we detected the sequences CCTTG and GGAAC respectively (see Figure 3c), which are core sequences for recognition by the paired box of PAX3 (Goulding et al, 1991). Additionally, at positions 742 to 739, +578 to +581 and +719 to +722 we identi®ed several ATTA boxes which might mediate binding by the homeodomain of PAX3 (Wilson et al, 1993). Next, a series of deletion constructs from the BCL-X promoter were generated in front of a luciferase reporter gene and used in cotransfection assays.…”

Gsh‐1: A novel murine homeobox gene expressed in the central nervous system

“…The random oligonucleotide selected binding and PCR amplification procedure described by Blackwell and Weintraub (1990), and modified by Wilson et al (1993), was used to define the binding sequence. A 74 base oligonucleotide with a 26 base random sequence core was synthesized and made double stranded, using a primer complementary to one end and Klenow fragment DNA polymerase.…”

“…Moreover, the non-homeodomain components of these proteins are quite variable. These regions can determine transcription activator or repressor function as well as dimerization reactions with other proteins and cooperative binding to target sequences (Wilson et al, 1993). In Drosophila the extradenticle encoded divergent homeodomain protein undergoes cooperative interactions with other homeoproteins to alter their target site specificity (Chan et al, 1994;van Dijk and Murre, 1994).…”

“…1) and 13 (bp 620-848) resulted in the most efficient protein production in E. coli and therefore column purified and used for selection. Four rounds of selection were performed as previously described (Wilson et al, 1993). Individual selected oligonucleotides were cloned and sequenced using either Sequenase (USB) or dsDNA Cycle Sequencing (BRL, Gaithersburg, MD).…”

Control of gastrula cell motility by the Goosecoid/Mix.1/ Siamois network: Basic patterns and paradoxical effects

“…Gsc belongs to a subgroup of pairedclass homeodomain transcription factors, which are characterized by a lysine at position 9 of the DNA recognition helix, and thus by specific binding site requirements (Wilson et al, 1993;Mailhos et al, 1998). It represses transcription by competing with activators for the same binding sites, or as an active DNA-binding repressor that physically interacts with other paired-class factors (Smith and Jaynes, 1996;Mailhos et al, 1998).…”