RESUMEN

BACKGROUND: Pitx3 plays a well understood role in directing development of lens, muscle fiber, and dopaminergic neurons; however, in Xenopus laevis, it may also play a role in early gastrulation and somitogenesis. Potential downstream targets of pitx3 possess multiple binding motifs that would not be readily accessible by conventional promoter analysis. RESULTS: We isolated and characterized pitx3 target genes lhx1 and xnr5 using a novel three-fluor flow cytometry tool that was designed to dissect promoters with multiple binding sites for the same transcription factor. This approach was calibrated using a known pitx3 target gene, Tyrosine hydroxylase. CONCLUSIONS: We demonstrate how flow cytometry can be used to detect gene regulatory changes with exquisite precision on a cell-by-cell basis, and establish that in HEK293 cells, pitx3 directly activates lhx1 and represses xnr5. Developmental Dynamics 246:657-669, 2017. © 2017 Wiley Periodicals, Inc.

Asunto(s)

Regulación del Desarrollo de la Expresión Génica/fisiología , Proteínas de Homeodominio/metabolismo , Proteínas con Homeodominio LIM/metabolismo , Ligandos de Señalización Nodal/metabolismo , Regiones Promotoras Genéticas/genética , Factores de Transcripción/metabolismo , Proteínas de Xenopus/metabolismo , Animales , Citometría de Flujo , Regulación del Desarrollo de la Expresión Génica/genética , Células HEK293 , Proteínas de Homeodominio/genética , Humanos , Proteínas con Homeodominio LIM/genética , Ligandos de Señalización Nodal/genética , Factores de Transcripción/genética , Proteínas de Xenopus/genética , Xenopus laevis