The broad goal of this application is to understand how embryonic cells are directed to follow particular paths of development. The conserved T-box family of transcriptional regulators specifies the development of the middle germ layer, mesoderm, in triploblastic animals. In mammals, mesoderm includes muscle, heart, skeleton, and blood. Mutations of T-box genes can result in birth defects such as cleft lip and palate. In the model animal C. elegans, many mesodermal cell types, including much of the body muscle and part of the pharynx, are made by descendants of a single embryonic cell called MS. The investigator has identified a T-box gene, tbx-35, that is required to specify the fate of MS. In embryos lacking tbx-35, MS descendants are not made. The conserved Wnt pathway, important in human cancer, transduces a signal that blocks activation of tbx-35 in the sister cell of MS, called E. Surprisingly, this repression does not involve the classical C. elegans Wnt effector TCF/POP-1.
Specific Aim 1 will fully characterize the phenotype of tbx-35(-) embryos by investigating the expression of pharynx and muscle genes using tissue-specific GFP reporters and in situ hybridization to understand in which tissues are the activities absent and identify the aberrant fate MS descendants ultimately adopt.
Specific Aim 2 will assess the ability of the TBX-35 protein to specify mesoderm fates when expressed outside of its normal context by using a heat-shock promoter fusion to overexpress tbx-35 throughout the embryo and assess production of ectopic muscle and pharynx fates by reporter and in situ hybridization.
Specific Aim 3 will test direct interaction of TBX-35 with the promoters of known pharynx (pha-4/FoxA) and muscle (hlh-1/MyoD) regulators, and to identify other putative TBX-35 targets using an in vitro genomic selection strategy to amplify fragments of C. elegans DNA that bind TBX-35.
Specific Aim 4 will characterize the Wnt-dependent repression of tbx-35 that occurs in E by using mutations and RNA interference to deplete function of Wnt/MAPK pathway genes that are known to participate in regulatory changes that make E different from MS, and assess their effect on tbx-35 repression. The investigator will also identify the cis-regulatory sites in tbx-35 that are responsive to this repression. As T-box genes and the Wnt pathway have similar roles in development of all animals, an understanding of how tbx-35 functions in C. elegans may shed light on how development goes awry, a process which can lead to birth defects or cancer. ? ? ?
| Owraghi, Melissa; Broitman-Maduro, Gina; Luu, Thomas et al. (2010) Roles of the Wnt effector POP-1/TCF in the C. elegans endomesoderm specification gene network. Dev Biol 340:209-21 |
| Maduro, Morris F (2009) Structure and evolution of the C. elegans embryonic endomesoderm network. Biochim Biophys Acta 1789:250-60 |
| Broitman-Maduro, Gina; Owraghi, Melissa; Hung, Wendy W K et al. (2009) The NK-2 class homeodomain factor CEH-51 and the T-box factor TBX-35 have overlapping function in C. elegans mesoderm development. Development 136:2735-46 |
| Lin, Katy Tan-Hui; Broitman-Maduro, Gina; Hung, Wendy W K et al. (2009) Knockdown of SKN-1 and the Wnt effector TCF/POP-1 reveals differences in endomesoderm specification in C. briggsae as compared with C. elegans. Dev Biol 325:296-306 |
