Xenopus pitx3 target genes lhx1 and xnr5 are identified using a novel three-fluor flow cytometry-based analysis of promoter activation and repression
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Updated: Feb 28, 2026
Using Confocal Analysis of Xenopus laevis to Investigate Modulators of Wnt and Shh Morphogen Gradients
Published on: December 14, 2015
Lara N Hooker1, Cristine Smoczer2, Samuel Abbott3
1Validation Technologies, Inc, San Diego, California.
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.
