RESUMO
The Hedgehog signaling pathway is part of the ancient developmental-evolutionary animal toolkit. Frequently co-opted to pattern new structures, the pathway is conserved among eumetazoans yet flexible and pleiotropic in its effects. The Hedgehog receptor, Patched, is transcriptionally activated by Hedgehog, providing essential negative feedback in all tissues. Our locus-wide dissections of the cis-regulatory landscapes of fly patched and mouse Ptch1 reveal abundant, diverse enhancers with stage- and tissue-specific expression patterns. The seemingly simple, constitutive Hedgehog response of patched/Ptch1 is driven by a complex regulatory architecture, with batteries of context-specific enhancers engaged in promoter-specific interactions to tune signaling individually in each tissue, without disturbing patterning elsewhere. This structure-one of the oldest cis-regulatory features discovered in animal genomes-explains how patched/Ptch1 can drive dramatic adaptations in animal morphology while maintaining its essential core function. It may also suggest a general model for the evolutionary flexibility of conserved regulators and pathways.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Proteínas Hedgehog/metabolismo , Receptor Patched-1/metabolismo , Transdução de Sinais , Animais , Linhagem Celular , Drosophila , CamundongosRESUMO
In the era of functional genomics, the role of transcription factor (TF)-DNA binding affinity is of increasing interest: for example, it has recently been proposed that low-affinity genomic binding events, though frequent, are functionally irrelevant. Here, we investigate the role of binding site affinity in the transcriptional interpretation of Hedgehog (Hh) morphogen gradients. We noted that enhancers of several Hh-responsive Drosophila genes have low predicted affinity for Ci, the Gli family TF that transduces Hh signalling in the fly. Contrary to our initial hypothesis, improving the affinity of Ci/Gli sites in enhancers of dpp, wingless and stripe, by transplanting optimal sites from the patched gene, did not result in ectopic responses to Hh signalling. Instead, we found that these enhancers require low-affinity binding sites for normal activation in regions of relatively low signalling. When Ci/Gli sites in these enhancers were altered to improve their binding affinity, we observed patterning defects in the transcriptional response that are consistent with a switch from Ci-mediated activation to Ci-mediated repression. Synthetic transgenic reporters containing isolated Ci/Gli sites confirmed this finding in imaginal discs. We propose that the requirement for gene activation by Ci in the regions of low-to-moderate Hh signalling results in evolutionary pressure favouring weak binding sites in enhancers of certain Hh target genes.
Assuntos
Sítios de Ligação/genética , Proteínas de Ligação a DNA/genética , Proteínas de Drosophila/genética , Drosophila/crescimento & desenvolvimento , Drosophila/genética , Evolução Molecular , Regulação da Expressão Gênica no Desenvolvimento/genética , Fatores de Transcrição/genética , Asas de Animais/crescimento & desenvolvimento , Animais , Sequência de Bases , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Discos Imaginais/metabolismo , Imuno-Histoquímica , Modelos Genéticos , Dados de Sequência Molecular , Ligação Proteica/genética , Seleção Genética , Alinhamento de Sequência , Asas de Animais/metabolismoRESUMO
How do gene regulatory networks evolve? A new study in yeasts shows that cis- and trans-regulatory changes resulted in a hybrid state of coexisting ancestral and derived regulatory circuits. This hybrid state then diversified into a variety of modern networks.
Assuntos
Evolução Molecular , Redes Reguladoras de Genes , Leveduras/genética , AnimaisRESUMO
Gradients of diffusible signaling proteins control precise spatial patterns of gene expression in the developing embryo. Here, we use quantitative expression measurements and thermodynamic modeling to uncover the cis-regulatory logic underlying spatially restricted gene expression in a Hedgehog (Hh) gradient in Drosophila. When Hh signaling is low, the Hh effector Gli, known as Cubitus interruptus (Ci) in Drosophila, acts as a transcriptional repressor; when Hh signaling is high, Gli acts as a transcriptional activator. Counterintuitively and in contrast to previous models of Gli-regulated gene expression, we found that low-affinity binding sites for Ci were required for proper spatial expression of the Hh target gene decapentaplegic (dpp) in regions of low Hh signal. Three low-affinity Ci sites enabled expression of dpp in response to low signal; increasing the affinity of these sites restricted dpp expression to regions of maximal signaling. A model incorporating cooperative repression by Ci correctly predicted the in vivo expression of a reporter gene controlled by a single Ci site. Our work clarifies how transcriptional activators and repressors, competing for common binding sites, can transmit positional information to the genome. It also provides an explanation for the widespread presence of conserved, nonconsensus Gli binding sites in Hh target genes.