Date of Award

2012

Publication Type

Doctoral Thesis

Degree Name

Ph.D.

Department

Biological Sciences

Keywords

Health and environmental sciences, Gastrulation, Laterality, Morpholino, Somitogenesis, Embryogenesis

Supervisor

Michael J. Crawford

Rights

info:eu-repo/semantics/openAccess

Abstract

Pitx3 encodes a homeodomain transcription factor that represents the causative locus for the aphakia phenotype in mouse as well as congenital cataracts and anterior segment mesenchymal dysgenesis in humans. Mutations in Pitx3 can also lead to the development of Parkinson's disease. A conserved role for Pitx3 has been established in the terminal differentiation and maintenance of lens fibres and dopaminergic neurons within the midbrain. Pitx3 has also been reported to contribute to skeletal muscle differentiation in mice. Through the use of morpholino-mediated knockdown, the Xenopus model system has allowed further refining of putative roles for pitx3 during embryogenesis. We report a novel role for pitx3 in regulating somitogenesis and laterality pathways and also influences upon retinoic acid signalling. Micorarray analysis identified gene networks affected by pitx3 knockdown within the eye, brain, segmentation patterning, and tailbud region. Early expression of pitx3 reveals unique involvement in early signaling pathways and subsequent effects on gastrulation. Novel transcripts were also identified and characterized for Rbp4l, GalectinIX, Rdh16, and Baz2b. Through the development of a novel reporter assay that utilizes flow cytometry, bicistronic vectors, and a three-fluor system, we determined nodal5, lhx1, and crybb1 to be direct targets of pitx3 regulation. This unique assay allows us to report that pitx3 operates in an all-or-none mechanism as both an activator and repressor protein.

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