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The embryonic nephric mesenchyme contains pluripotent progenitor cells. Six2, a homeodomain transcription factor, is expressed in a subset of the nephric mesenchyme, and it functions to maintain a progenitor state by suppressing nephrogenesis. Despite the functional significance of Six2 in nephric development, its regulatory mechanisms remain unclear. To identify the cis-regulatory elements for Six2, we focused on the evolutionarily conserved sequences known as conserved noncoding sequences (CNSs) associated with the Six2 locus. Transgenic experiments using Xenopus laevis embryos revealed that three of the eight CNSs located within a 317-kb segment of the Six2 genomic locus were nephric enhancers. Motif analysis of transcription factors combined with phylogenetic footprinting revealed the enrichment of putative T-cell factor (Tcf)-, Hox-, and SWI/SNF complex helicase-like transcription factor (Hltf)- and AT-rich interactive domain 3A (Arid3a)-binding motif sequences in these enhancers.

Identification of distal enhancers for Six2 expression in pronephros