title: Low affinity binding site clusters confer hox specificity and regulatory robustness.
abstract: In animals, Hox transcription factors define regional identity in distinct anatomical domains. How Hox genes encode this specificity is a paradox, because different Hox proteins bind with high affinity in vitro to similar DNA sequences. Here, we demonstrate that the Hox protein Ultrabithorax (Ubx) in complex with its cofactor Extradenticle (Exd) bound specifically to clusters of very low affinity sites in enhancers of the shavenbaby gene of Drosophila. These low affinity sites conferred specificity for Ubx binding in vivo, but multiple clustered sites were required for robust expression when embryos developed in variable environments. Although most individual Ubx binding sites are not evolutionarily conserved, the overall enhancer architecture-clusters of low affinity binding sites-is maintained and required for enhancer function. Natural selection therefore works at the level of the enhancer, requiring a particular density of low affinity Ubx sites to confer both specific and robust expression.
authors.affiliation: Janelia Research Campus, Howard Hughes Medical Institute, 19700 Helix Drive, Ashburn, VA 20147, USA.
authors.forename: Justin
authors.initials: J
authors.lastname: Crocker
authors.affiliation: Columbia University Medical Center, 701 West 168(th) Street, HHSC 1104, New York, NY 10032, USA.
authors.forename: Namiko
authors.initials: N
authors.lastname: Abe
authors.affiliation: Columbia University Medical Center, 701 West 168(th) Street, HHSC 1104, New York, NY 10032, USA.
authors.forename: Lucrezia
authors.initials: L
authors.lastname: Rinaldi
authors.affiliation: Department of Biological and Medical Sciences, Oxford Brookes University, Gipsy Lane, Oxford OX3 0BP, UK.
authors.forename: Alistair P
authors.initials: AP
authors.lastname: McGregor
authors.affiliation: Departamento de Ecología, Genética y Evolución, IEGEBA-CONICET, Facultad, de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad, Universitaria, Pabellón 2, 1428 Buenos Aires, Argentina.
authors.forename: Nicolás
authors.initials: N
authors.lastname: Frankel
authors.affiliation: New Jersey Neuroscience Institute, 65 James Street, Edison, NJ 08820, USA.
authors.forename: Shu
authors.initials: S
authors.lastname: Wang
authors.affiliation: Centre de Biologie du Développement, Université de Toulouse, UPS, 31062 Cedex 9, France; CNRS, UMR5547, Centre de Biologie du Développement, Toulouse, 31062 Cedex 9, France.
authors.forename: Ahmad
authors.initials: A
authors.lastname: Alsawadi
authors.affiliation: Centre de Biologie du Développement, Université de Toulouse, UPS, 31062 Cedex 9, France; CNRS, UMR5547, Centre de Biologie du Développement, Toulouse, 31062 Cedex 9, France.
authors.forename: Philippe
authors.initials: P
authors.lastname: Valenti
authors.affiliation: Centre de Biologie du Développement, Université de Toulouse, UPS, 31062 Cedex 9, France; CNRS, UMR5547, Centre de Biologie du Développement, Toulouse, 31062 Cedex 9, France.
authors.forename: Serge
authors.initials: S
authors.lastname: Plaza
authors.affiliation: Centre de Biologie du Développement, Université de Toulouse, UPS, 31062 Cedex 9, France; CNRS, UMR5547, Centre de Biologie du Développement, Toulouse, 31062 Cedex 9, France.
authors.forename: François
authors.initials: F
authors.lastname: Payre
authors.affiliation: Columbia University Medical Center, 701 West 168(th) Street, HHSC 1104, New York, NY 10032, USA. Electronic address: rsm10@columbia.edu.
authors.forename: Richard S
authors.initials: RS
authors.lastname: Mann
authors.affiliation: Janelia Research Campus, Howard Hughes Medical Institute, 19700 Helix Drive, Ashburn, VA 20147, USA. Electronic address: sternd@janelia.hhmi.org.
authors.forename: David L
authors.initials: DL
authors.lastname: Stern
mesh_terms: D000818:Animals; D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000235:genetics*; D029721:Drosophila Proteins / Q000235:genetics* / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism*; D004625:Embryo, Nonmammalian / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation; D018398:Homeodomain Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D011485:Protein Binding; D016415:Sequence Alignment; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism*
chemical_list: D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D018398:Homeodomain Proteins; D014157:Transcription Factors; C066177:Ubx protein, Drosophila; C083405:exd protein, Drosophila; C089512:ovo protein, Drosophila