Friend of GATA (FOG) Interacts with the Nucleosome Remodeling and Deacetylase Complex (NuRD) to Support Primitive Erythropoiesis in Xenopus laevis

Friend of GATA (FOG) plays many diverse roles in adult and embryonic hematopoiesis, however the mechanisms by which it functions and the roles of potential interaction partners are not completely understood. Previous work has shown that overexpression of FOG in Xenopus laevis causes loss of blood suggesting that in contrast to its role in mammals, FOG might normally function to repress erythropoiesis in this species. Using loss-of-function analysis, we demonstrate that FOG is essential to support primitive red blood cell (RBC) development in Xenopus. Moreover, we show that it is specifically required to prevent excess apoptosis of circulating primitive RBCs and that in the absence of FOG, the pro-apoptotic gene Bim-1 is strongly upregulated. To identify domains of FOG that are essential for blood development and, conversely, to begin to understand the mechanism by which overexpressed FOG represses primitive erythropoiesis, we asked whether FOG mutants that are unable to interact with known co-factors retain their ability to rescue blood formation in FOG morphants and whether they repress erythropoiesis when overexpressed in wild type embryos. We find that interaction of FOG with the Nucleosome Remodeling and Deacetylase complex (NuRD), but not with C-terminal Binding Protein, is essential for normal primitive RBC development. In contrast, overexpression of all mutant and wild type constructs causes a comparable repression of primitive erythropoiesis. Together, our data suggest that a requirement for FOG and its interaction with NuRD during primitive erythropoiesis are conserved in Xenopus and that loss of blood upon FOG overexpression is due to a dominant-interfering effect

The Sno Oncogene Antagonizes Wingless Signaling during Wing Development in Drosophila

The Sno oncogene (Snoo or dSno in Drosophila) is a highly conserved protein and a well-established antagonist of Transforming Growth Factor-β signaling in overexpression assays. However, analyses of Sno mutants in flies and mice have proven enigmatic in revealing developmental roles for Sno proteins. Thus, to identify developmental roles for dSno we first reconciled conflicting data on the lethality of dSno mutations. Then we conducted analyses of wing development in dSno loss of function genotypes. These studies revealed ectopic margin bristles and ectopic campaniform sensilla in the anterior compartment of the wing blade suggesting that dSno functions to antagonize Wingless (Wg) signaling. A subsequent series of gain of function analyses yielded the opposite phenotype (loss of bristles and sensilla) and further suggested that dSno antagonizes Wg signal transduction in target cells. To date Sno family proteins have not been reported to influence the Wg pathway during development in any species. Overall our data suggest that dSno functions as a tissue-specific component of the Wg signaling pathway with modest antagonistic activity under normal conditions but capable of blocking significant levels of extraneous Wg, a role that may be conserved in vertebrates

Reconstituting regulation of the canonical Wnt pathway by engineering a minimal β-catenin destruction machine

Negatively regulating key signaling pathways is critical to development and altered in cancer. Wnt signaling is kept off by the destruction complex, which is assembled around the tumor suppressors APC and Axin and targets β-catenin for destruction. Axin and APC are large proteins with many domains and motifs that bind other partners. We hypothesized that if we identified the essential regions required for APC:Axin cooperative function and used these data to design a minimal β-catenin-destruction machine, we would gain new insights into the core mechanisms of destruction complex function. We identified five key domains/motifs in APC or Axin that are essential for their function in reconstituting Wnt regulation. Strikingly, however, certain APC and Axin mutants that are nonfunctional on their own can complement one another in reducing β-catenin, revealing that the APC:Axin complex is a highly robust machine. We used these insights to design a minimal β-catenin-destruction machine, revealing that a minimized chimeric protein covalently linking the five essential regions of APC and Axin reconstitutes destruction complex internal structure, size, and dynamics, restoring efficient β-catenin destruction in colorectal tumor cells. On the basis of our data, we propose a new model of the mechanistic function of the destruction complex as an integrated machine

Deconstructing the ßcatenin destruction complex: mechanistic roles for the tumor suppressor APC in regulating Wnt signaling

APC is a key tumor suppressor and Wnt signaling regulator, but its mechanism of action remains mysterious. We combined parallel assays in Drosophila and cultured human colon cancer cell lines to test hypotheses regarding APC function and to develop novel hypotheses, using mutants altering its structure in specific ways