RESUMEN
Wnt ligands are considered classical morphogens, for which the strength of the cellular response is proportional to the concentration of the ligand. Herein, we show an emergent property of bistability arising from feedback among the Wnt destruction complex proteins that target the key transcriptional co-activator ß-catenin for degradation. Using biochemical reconstitution, we identified positive feedback between the scaffold protein Axin and the kinase glycogen synthase kinase 3 (GSK3). Theoretical modeling of this feedback between Axin and GSK3 suggested that the activity of the destruction complex exhibits bistable behavior. We experimentally confirmed these predictions by demonstrating that cellular cytoplasmic ß-catenin concentrations exhibit an "all-or-none" response with sustained memory (hysteresis) of the signaling input. This bistable behavior was transformed into a graded response and memory was lost through inhibition of GSK3. These findings provide a mechanism for establishing decisive, switch-like cellular response and memory upon Wnt pathway stimulation.
Asunto(s)
Complejo de Señalización de la Axina , beta Catenina , Complejo de Señalización de la Axina/metabolismo , beta Catenina/metabolismo , Proteína Axina/genética , Proteína Axina/metabolismo , Glucógeno Sintasa Quinasa 3/metabolismo , Retroalimentación , Fosforilación , Vía de Señalización Wnt/fisiologíaRESUMEN
Immunomodulatory drugs (IMiDs) are important for the treatment of multiple myeloma and myelodysplastic syndrome. Binding of IMiDs to Cereblon (CRBN), the substrate receptor of the CRL4CRBN E3 ubiquitin ligase, induces cancer cell death by targeting key neo-substrates for degradation. Despite this clinical significance, the physiological regulation of CRBN remains largely unknown. Herein we demonstrate that Wnt, the extracellular ligand of an essential signal transduction pathway, promotes the CRBN-dependent degradation of a subset of proteins. These substrates include Casein kinase 1α (CK1α), a negative regulator of Wnt signaling that functions as a key component of the ß-Catenin destruction complex. Wnt stimulation induces the interaction of CRBN with CK1α and its resultant ubiquitination, and in contrast with previous reports does so in the absence of an IMiD. Mechanistically, the destruction complex is critical in maintaining CK1α stability in the absence of Wnt, and in recruiting CRBN to target CK1α for degradation in response to Wnt. CRBN is required for physiological Wnt signaling, as modulation of CRBN in zebrafish and Drosophila yields Wnt-driven phenotypes. These studies demonstrate an IMiD-independent, Wnt-driven mechanism of CRBN regulation and provide a means of controlling Wnt pathway activity by CRBN, with relevance for development and disease.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Péptido Hidrolasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Vía de Señalización Wnt/fisiología , Proteínas de Pez Cebra/genética , Proteínas Adaptadoras Transductoras de Señales/química , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Caseína Quinasa Ialfa/metabolismo , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Embrión no Mamífero , Evolución Molecular , Células HEK293 , Humanos , Factores Inmunológicos/química , Factores Inmunológicos/farmacología , Lenalidomida/química , Lenalidomida/farmacología , Ratones , Organoides , Péptido Hidrolasas/metabolismo , Ubiquitina-Proteína Ligasas/química , Ubiquitina-Proteína Ligasas/genética , Ubiquitinación , Pez Cebra/embriología , Pez Cebra/genética , Proteínas de Pez Cebra/metabolismoRESUMEN
Homeostasis in adult organs involves replacement of cells from a stem cell pool maintained in specialized niches regulated by extracellular signals. This cell-to-cell communication employs signal transduction pathways allowing cells to respond with a variety of behaviors. To study these cellular behaviors, signaling must be perturbed within tissues in precise patterns, a technique recently made possible by the development of optogenetic tools. We developed tools to study signal transduction in vivo in an adult fly midgut stem cell model where signaling was regulated by the application of light. Activation was achieved by clustering of membrane receptors EGFR and Toll, while inactivation was achieved by clustering the downstream activators ERK/Rolled and NFκB/Dorsal in the cytoplasm, preventing nuclear translocation and transcriptional activation. We show that both pathways contribute to stem and transit amplifying cell numbers and affect the lifespan of adult flies. We further present new approaches to overcome overexpression phenotypes and novel methods for the integration of optogenetics into the already-established genetic toolkit of Drosophila.
Asunto(s)
Drosophila melanogaster/crecimiento & desarrollo , Redes Reguladoras de Genes , Mucosa Intestinal/citología , Optogenética/métodos , Animales , Comunicación Celular , Proliferación Celular , Células Cultivadas , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Regulación de la Expresión Génica , Homeostasis , Mucosa Intestinal/metabolismo , Luz , Longevidad , Transducción de Señal , Células Madre/citología , Células Madre/metabolismoRESUMEN
Developmental signaling pathways control a vast array of biological processes during embryogenesis and in adult life. The WNT pathway was discovered simultaneously in cancer and development. Recent advances have expanded the role of WNT to a wide range of pathologies in humans. Here, we discuss the WNT pathway and its role in human disease and some of the advances in WNT-related treatments.