The T-box genes constitute an evolutionarily conserved family of putative transcription factors which are expressed in discrete domains during embryogenesis, suggesting that they may play roles in inductive interactions. Members have been identified by virtue of their homology to the prototypical T-box gene, T or Brachyury, which is required for mesoderm formation and axial elongation during embryogenesis. We have previously reported the discovery of six new mouse T-box genes, Tbx1-Tbx6, and described the expression patterns of Tbx1-Tbx5 (Bollag et al., 1994; Agulnik et al., 1996; Chapman et al., 1996; Gibson-Brown et al., 1996). We have obtained cDNA clones encoding the full-length Tbx6 protein from screens of gastrulation-stage mouse cDNA libraries and determined the spatial and temporal distribution of Tbx6 transcripts during embryogenesis. The gene codes for a 1.9-kb transcript with an open reading frame coding for a 540-amino acid protein, with a predicted molecular weight of 59 kDa. Tbx6 maps to chromosome 7 and does not appear to be linked to any known mutation. Unlike other members of the mouse T-box gene family which are expressed in a wide variety of tissues derived from all germ layers, Tbx6 expression is quite restricted. Tbx6 transcripts are first detected in the gastrulation stage embryo in the primitive streak and newly recruited paraxial mesoderm. Later in development, Tbx6 expression is restricted to presomitic, paraxial mesoderm and to the tail bud, which replaces the streak as the source of mesoderm. Expression in the tail bud persists until 12. 5 days postcoitus. Tbx6 expression thus overlaps that of Brachyury in the primitive streak and tail bud, although Brachyury is expressed earlier in the primitive streak. Brachyury is also expressed in a second domain, the node and notochord, that is not shared with Tbx6. The onset of Tbx6 expression is not affected in homozygous null Brachyury mutant embryos at 7.5 days postcoitus. However, Tbx6 expression is extinguished in mutant embryos as soon as the Brachyury phenotype becomes evident at 8.5 days postcoitus, indicating that the continued expression of Tbx6 is directly or indirectly dependent upon Brachyury expression.

The T-box genes comprise an ancient family of putative transcription factors conserved across species as divergent as Mus musculus and Caenorhabditis elegans. All T-box gene products are characterized by a novel 174-186amino acid DNA binding domain called the T-box that was first discovered in the polypeptide products of the mouse T locus and the Drosophila melanogaster optomotor-blind gene. Earlier studies allowed the identification of five mouse T-box genes, T, Tbx1-3, and Tbr1, that all map to different chromosomal locations and are expressed in unique temporal and spatial patterns during embryogenesis. Here, we report the discovery of three new members of the mouse T-box gene family, named Tbx4, Tbx5, and Tbx6. Two of these newly discovered genes, Tbx4 and Tbx5, were found to be tightly linked to previously identified T-box genes. Combined results from phylogenetic, linkage, and physical mapping studies provide a picture for the evolution of a T-box subfamily by unequal crossing over to form a two-gene cluster that was duplicated and dispersed to two chromosomal locations. This analysis suggests that Tbx4 and Tbx5 are cognate genes that diverged apart from a common ancestral gene during early vertebrate evolution.

The brachyury, or T, locus encodes a transcription factor that plays a crucial role in the early development of all animals. In the mouse, animals heterozygous for a null mutation at this locus are born with a characteristic short tail. Expressivity of the short tail phenotype is greatly affected by genetic background. As a genetic entry into the identification of genes that interact with the Brachyury locus, we have performed a QTL analysis for modifiers of this phenotype. Surprisingly, we discovered that the major modifiers uncovered all act in a sex-limited manner. We have identified two QTLs--Brm1 on Chr 9 and Brm2 on Chr 15--that act only in female offspring(N2) from female T/+ parents(F1) and are responsible together for most, or all, of the genetic variance in phenotypic expression observed between C57BL/10 and C3H/HeJ animals.