tbx-9 is one of T-box family genes that share a DNA binding motif, T-box. Genes of the family are found widely in metazoans, and many of them are involved in important events for development. Three T-box genes, Brachyury (T) , VegT and
tbx-2 , have been shown to encode transcription factors. In C. elegans , the whole genome sequence predicts that this organism has about 20 T-box genes, and cDNA clones corresponding to the predicted genes are being isolated by the cDNA project. As I am interested in the regulation of gene expression, I have been studying one of the T-box genes, CELK02736 =
tbx-9 , which is the first one among T-box genes whose cDNA clones have been isolated. I previously showed that
tbx-9 is expressed in only a few cells of early-stage embryos and
tbx-9 deletion mutants generated by gene disruption have defects in morphogenesis of the posterior body region of the embryo. To test whether the
tbx-9 protein has a sequence-specific DNA binding activity, the
tbx-9 DNA binding sequence was determined as consensus among sequences of DNA fragments that were selected from a mixture of DNA fragments of random sequence by forming complex with the
tbx-9 protein. The determined
tbx-9 binding sequence was 18 bases in length and had a palindromic structure. The T binding sequence determined by a similar method has been reported to be palindromic, and the binding sequences of T and
tbx-9 are almost identical. To know whether the
tbx-9 protein is a transcription factor, transactivation of a reporter gene depending on the
tbx-9 binding sequence was examined. To this end, the following plasmids were constructed. For forced expression of
tbx-9 , the
tbx-9 gene was ligated downstream of the
hsp16-41 heat shock promoter. For assay of transactivation, the
tbx-9 binding sequence was inserted upstream of the
pes-10 minimal promoter and the reporter lacZ gene. Transgenic lines containing the both plasmids were produced by co-injection. Ectopic expression of
tbx-9 by subjecting transgenic embryos to heat shock caused activation of the lacZ gene, indicating that the
tbx-9 protein functions as a transcription activator. As the next goal, I am aiming at identifying target genes of
tbx-9 .