The anterior heart field (AHF) mediates formation of the outflow tract (OFT) and right ventricle (RV) during looping morphogenesis of the heart. Foxh1 is a forkhead DNA binding transcription factor in the TGFbeta-Smad pathway. Here we demonstrate that Foxh1-/- mutant mouse embryos form a primitive heart tube, but fail to form OFT and RV and display loss of outer curvature markers of the future working myocardium, similar to the phenotype of Mef2c-/- mutant hearts. Further, we show that Mef2c is a direct target of Foxh1, which physically and functionally interacts with Nkx2-5 to mediate strong Smad-dependent activation of a TGFbeta response element in the Mef2c gene. This element directs transgene expression to the presumptive AHF, as well as the RV and OFT, a pattern that closely parallels endogenous Mef2c expression in the heart. Thus, Foxh1 and Nkx2-5 functionally interact and are essential for development of the AHF and its derivatives, the RV and OFT, in response to TGFbeta-like signals.

The role of polarity signaling in cancer metastasis is ill defined. Using two three-dimensional culture models of mammary epithelial cells and an orthotopic mouse model of breast cancer, we reveal that Par6 signaling, which is regulated directly by TGF␤, plays a role in breast cancer metastasis. Interference with Par6 signaling blocked TGF␤-dependent loss of polarity in acini-like structures formed by non-transformed mammary cells grown in three-dimensional structures and suppressed the protrusive morphology of mesenchymal-like invasive mammary tumor cells without rescuing E-cadherin expression. Moreover, blockade of Par6 signaling in an in vivo orthotopic model of metastatic breast cancer induced the formation of ZO-1-positive epithelium-like structures in the primary tumor and suppressed metastasis to the lungs. Analysis of the pathway in tissue microarrays of human breast tumors further revealed that Par6 activation correlated with markers of the basal carcinoma subtype in BRCA1-associated tumors. These studies thus reveal a key role for polarity signaling and the control of morphologic transformation in breast cancer metastasis.epithelial-to-mesenchymal transition ͉ cell polarity ͉ metastasis ͉ tumor invasion ͉ epithelial plasticity M etastasis, the spread of cancer cells from the primary tumor site to distant organs, accounts for over 90% of deaths in breast cancer patients (1). Metastasis has been associated with epithelial-to-mesenchymal transition (EMT), which is a complex manifestation of epithelial plasticity, in which polarized epithelial cells embedded in organized, stratified, or single cell layers convert into single fibroblastoid cells capable of locomotion (2). Cellular changes necessary for EMT include both morphological changes, as well as alterations in gene expression. While the role of the gene expression program associated with EMT has been well-described (3), it is unclear how the morphological changes associated with EMT specifically contribute to cancer progression and metastasis in vivo. The Par6 polarity complex localizes to the tight junction (TJ) and is an important regulator of the morphological transitions associated with epithelial cell plasticity (4). The complex is comprised of three highly conserved proteins, including Par3, Par6, and aPKC. Par6 is a core component that was initially identified as one of the six Par (for ''partitioning''-defective) proteins essential for asymmetric cell division in the C. elegans zygote, and was subsequently found to be required for asymmetric division of neuroblasts and the differentiation of oocytes in Drosophila, as well as the establishment/maintenance of apical-basal polarity and polarized migration in both Drosophila and mammalian cells. Par6-dependent control of apical-basal polarity is mediated by its interaction with Par3 and aPKC, as well as the Crumbs complex (4). Par6 is regulated directly by TGF␤ (5) and ErbB-2 receptors (6) to control epithelial cell plasticity and misregulation in expression of polarity proteins, including Scribb...

SummaryC1D is a gamma-irradiation inducible nuclear matrix protein that interacts with and activates the DNAdependent protein kinase (DNA-PK) that is essential for the repair of the DNA double-strand breaks and V(D)J recombination. Recently, it was demonstrated that C1D can also interact with TRAX and prevent the association of TRAX with Translin, a factor known to bind DNA break-point junctions, and that over expression of C1D can induce p53-dependent apoptosis. Taken together, these findings suggest that mammalian C1D could be involved in maintenance of genome integrity by regulating the activity of proteins involved in DNA repair and recombination. To obtain direct evidence for the biological function of C1D that we show is highly conserved between diverse species, we have analysed the Saccharomyces cerevisiae C1D homologue. We report that the disruption of the YC1D gene results in a temperature sensitivity and that yc1d mutant strains exhibit defects in nonhomologous DNA end joining (NHEJ) and accurate DNA repair. In addition, using a novel plasmid-based in vivo recombination assay, we show that yc1d mutant strains are also defective in homologous recombination. These results indicate that YC1D is implicated in both homologous recombination and NHEJ pathways for the repair of DNA double-strand breaks.

The nuclear matrix protein C1D is an activator of the DNA-dependent protein kinase (DNA-PK), which is essential for the repair of DNA double-strand breaks (DSBs) and V(D)J recombination. C1D is phosphorylated very efficiently by DNA-PK, and its mRNA and protein levels are induced upon γ-irradiation, suggesting that C1D may play a role in repair of DSBs in vivo. In an attempt to identify the biological function of C1D, we have employed the yeast two-hybrid system and found that C1D interacts specifically with Translin-associated factor X, TRAX. Although the biological function of TRAX remains unknown, its bipartite nuclear targeting sequences suggest a role for TRAX in the movement of associated proteins, including Translin, into the nucleus. We show that C1D and TRAX interact specifically in both yeast and mammalian cells. Interestingly, however, interaction of these two proteins in mammalian cells only occur following γ-irradiation, raising the possibility of involvement of TRAX in DNA double-strand break repair and providing evidence for biological functions of the nuclear matrix protein C1D and TRAX. Moreover, we show, using fluorescently tagged proteins, that the relative expression levels of TRAX and Translin affect their subcellular localization. These results suggest that one role for C1D may be to regulate TRAX/Translin complex formation.