Date of Award

Spring 2011

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Cell & Molecular Biology

First Advisor

Daniel S. Kessler

Abstract

Establishment of the vertebrate body axis requires the formation of the organizer domain during early embryogenesis. In amphibians, this domain is referred to as the Spemann Organizer and is essential for germ layer patterning and formation of the embryonic body axes. The Wnt and Nodal signaling pathways are both essential for organizer formation, but how these signals are integrated to influence gene expression in the organizer is largely unknown. The Wnt pathway activates expression of two transactivators, Siamois (Sia) and Twin (Twn), which mediate organizer formation downstream of Wnt. Expression of Sia or Twn is sufficient to induce an ectopic axis, suggesting they play an essential role in organizer formation. However, whether Sia/Twn play equivalent roles in organizer formation, or whether both are required for all aspects of organizer formation in not clear. Here, we report that knockdown of Sia/Twn together, but not individually, disrupts organizer gene expression and axis formation. We identify the Sia/Twn binding site within the promoter of the organizer gene Goosecoid (Gsc). Sia/Twn form homodimers and heterodimers through direct homeodomain interaction and both dimer forms are found at the endogenous Gsc promoter. The Gsc promoter also contains a Nodal responsive distal element, suggesting that Sia/Twn cooperate with Nodal signals in the transcription of Gsc. We find that Wnt and Nodal effectors synergize to activate transcription of three organizer genes, Gsc, Cerberus (Cer), and Chordin (Chd). Sia/Twn and the Nodal effectors FoxH1 and Smad2/3 occupy the promoters of these genes in vivo and their occupancy increases with active signaling from both pathways. This suggests that a complex consisting of Sia/Twn and Nodal effectors forms at organizer gene promoters. Consistent with this, p300 is recruited to organizer gene promoters in response to Sia/Twn or Nodal signals. Thus, Sia/Twn interact with Nodal effectors to regulate the spatial and temporal expression of organizer genes, suggesting a general mechanism for the regulation of organizer gene expression in the early embryo.

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