Investigation of the role of Tbx1, an essential T -box gene, in the mesoderm during embryogenesis

Abstract

In vertebrates, the inner ear is comprised of the cochlea and vestibular system, which develop from the otic vesicle. This process is regulated via inductive interactions from surrounding tissues. Tbx1, the gene responsible for velo-cardio-facial syndrome/DiGeorge syndrome in humans, is required for ear development in mice. Tbx1 is expressed in the otic epithelium and adjacent periotic mesenchyme (POM), and both of these domains are required for inner ear formation. To study the function of Tbx1 in the POM, we conditionally inactivated Tbx1 in the mesoderm while keeping otic vesicle expression intact. Conditional mutants (TCre-KO) displayed malformed inner ears with a severely shortened cochlear duct, indicating that Tbx1 expression in the POM is necessary for proper inner ear formation. Signaling between the POM and the otic epithelium was explored by via analysis of retinoic acid (RA) catabolizing genes. Cyp26a1 expression was altered in the TCre-KO, while Cyp26c1 showed reduced expression in TCre-KO embryos. These results indicate that Tbx1 signaling from the POM regulates local RA activity to control outgrowth of the cochlea.;BRN4 is a Pou-domain transcription factor associated with DFN3, an X-linked form of congenital deafness in humans. Expression of Brn4 was lost in the POM of TCre-KO mutants. To explore the genetic interaction between these two genes, mice mutant for Tbx1 and Brn4 were generated. Brn4 -;Tbxl+/- embryos displayed a reduction of Cyp26c1 expression in the POM and completely penetrant defects in growth of the distal cochlea. In addition, adult Brn4 -;Tbx1+/- mice displayed defects analogous to Mondini dysplasia, a malformation of the cochlea found in humans with mutations in TBX1 or BRN4. These data indicate that Tbx1 and Brn4 genetically interact and have a role in the etiology of congenital deafness in humans.;Finally, analysis of Tbx1 protein function during differentiation of mesodermal cells was performed. Tbx1 was found to translocate from the nucleus to the cytoplasm during differentiation of muscles cells and interact with the shuttling protein Wtip. Tbx1 protein was observed to localization at the Z-discs together with alpha-actininin in mouse skeletal muscle. These data suggest a possible role for Tbx1 in differentiation of muscle cells.