TheXenopushomeobox genepitx3impinges upon somitogenesis and laterality

Abstract

Pitx3 has been identified as the causative locus in a developmental eye mutation associated with mammalian anterior segment dysgenesis, congenital cataracts, and aphakia. In recent studies of frog eye development we discovered that pitx3 expresses symmetrically in the somites and lateral plate mesoderm and asymmetrically during cardiac and gut looping. We report that disruption of pitx3 activity on one side of an embryo relative to the other, either by over- or underexpression of pitx3, elicits a crooked dorsal axis in embryos that is a consequence of a retarded progression through somitogenesis. Unlike in amniotes, Xenopus somites form as cohorts of presomitic cells that rotate perpendicular to the dorsal axis. Since no vertebral anomalies have been reported in mouse and human Pitx3 mutants, we attempt to distinguish whether the segmentation clock is uniquely affected in frog or if the pitx3 perturbation inhibits the cellular changes that are necessary to rotation of presomitic cells. In Xenopus, pitx3 appears to inhibit the rotation of presomitic cell cohorts and to be necessary to the bilaterally symmetric expression of pitx2 in somites.

Xenopus pitx3 target genes lhx1 and xnr5 are identified using a novel three‐fluor flow cytometry–based analysis of promoter activation and repression

“…Finally, xnr5 has been identified as an upstream regulator of xnr1, lefty , and pitx2 (Tadjuidje et al, ). Our study, confirmed by in situ hybridization of the relevant downstream targets, places pitx3 one step further up in the hierarchical control of laterality, and explains laterality phenotypes previously reported (Khosrowshahian et al, ; Smoczer et al, ).…”

“…Our two candidate pitx3 target genes are interesting insofar as they are known to play a role during gastrulation and early dorsal axis development, as well as later during eye development. Previous work has indicated that pitx3 expresses in fish hypoblast (Dutta et al, ), in Xenopus pre‐gastrula (RT‐PCR), and in dorsal mesoderm (in situ) (Khosrowshahian et al, ; Smoczer et al, ). Lhx1 is an LIM ‐class homeodomain transcription factor that is expressed in several waves, the first of which is in Spemann's organizer, where it expresses in the involuting mesodermal cells (Taira et al, ; Hukriede et al, ).…”

“…Either too little or too much protocadherin impairs normal rotation of presomitic mesoderm preliminary to the formation of discrete somites (Kim et al, ). Therefore, pitx3 ‐induced disruption of protocadherin via lhx perturbation would explain why pitx3 over‐ or underexpression leads to identical segmentation phenotypes in Xenopus (Smoczer et al, ). Mutation of the pitx3 binding motif is sufficient to depress lhx1 promoter activity in vitro and the effect in transgenic embryos is more profound— expression appears to be almost completely impaired.…”

“…While Xenopus laevis pitx3 plays a similar role during eye development, it additionally expresses during gastrulation as well as later the in presomitic mesoderm, lateral plate mesoderm, differentiating somites, craniofacial regions, and looping heart and gut (Pommereit et al, 2001;Khosrowshahian et al, 2005;Smoczer et al, 2013). In contrast to its role in mammals, Xenopus pitx3 also affects laterality (left-right organ asymmetry) and somitogenesis (Khosrowshahian et al, 2005;Smoczer et al, 2013). Remarkably, these latter phenotypes are elicited by both gain of function as well as by morpholino-mediated translational knockdown (Smoczer et al, 2013).…”

“…In contrast to its role in mammals, Xenopus pitx3 also affects laterality (left‐right organ asymmetry) and somitogenesis (Khosrowshahian et al, ; Smoczer et al, ). Remarkably, these latter phenotypes are elicited by both gain of function as well as by morpholino‐mediated translational knockdown (Smoczer et al, ). We decided to identify some of the pitx3 target genes that might mediate both eye and novel phenotypes.…”