Dissertations, Theses, and Capstone Projects

Date of Degree

9-2021

Document Type

Dissertation

Degree Name

Ph.D.

Program

Biology

Advisor

Daniel Weinstein

Committee Members

Nathalia Holtzman

Cathy Savage-Dunn

Mark Emerson

Jean-Pierre Saint-Jeannet

Subject Categories

Cell and Developmental Biology

Keywords

Tbx2, development, Xenopus laevis, TGF-beta signaling, Histone 2A, R-Smads

Abstract

The differentiation of the three primary germ layers is precisely regulated by inductive cues, the intracellular networks through which these signals are transduced, and a broad array of nuclear proteins, such as transcription factors and epigenetic modifiers. Precise regulation of these various factors is crucial to proper development. Members of the T-box family of DNA-binding proteins play a prominent role in the differentiation of the three primary germ layers. VegT, Brachyury, and Eomesodermin function as transcriptional activators, are expressed in the presumptive mesendoderm and, in addition to directly activating the transcription of endoderm- and mesoderm-specific genes, serve variously as regulators of growth factor signaling during induction of these germ layers. In contrast, expression of the T-box gene tbx2 is present in the embryonic ectoderm, where Tbx2 functions as a transcriptional repressor and inhibits mesodermal and endodermal differentiation by the TGFβ ligand Activin. Notably, Tbx2 misexpression also promotes dorsal ectodermal fate via inhibition of the BMP branch of the TGFβ signaling network. We report here that Tbx2 physically associates with both Smad1 and Smad2, mediators of BMP and Activin/Nodal signaling, respectively. We perform structure function analysis of Tbx2 to elucidate the roles of both Tbx2-Smad interaction and Tbx2-DNA binding in germ layer suppression. Our studies demonstrate that Tbx2 associates with intracellular mediators of the Activin/Nodal and BMP/GDF pathways, crucial factors in germ layer patterning and differentiation. We have identified a novel repressor domain within Tbx2, and have determined that Tbx2 DNA-binding activity is required for repression of TGFβ signaling. Protein- protein interaction assays suggest that Tbx2-mediated repression involves an epigenetic mechanism. Finally, our data also point to overlapping yet distinct mechanisms for Tbx2-mediated repression of Activin/Nodal and BMP/GDF signaling.

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