Both the functional specificity and autoregulative activity of two ascidian T‐box genes HrBra and HrTbx6 are likely to be mediated by the DNA‐binding domain

Abstract

T-box genes encode a family of transcription factors having conserved DNA-binding domains and diverged transcription regulatory domains, and each family member shows a specific expression pattern and plays a specific and crucial role in animal development. Two fundamental questions to be answered are whether the T-box gene functional specificity is located in the DNA-binding domain or in the transcription regulatory domain and how the specific expression of T-box genes is controlled. In the ascidian Halocynthia roretzi, Brachyury (HrBra) is expressed only in notochord cells while Tbx6 (HrTbx6) is expressed in muscle cells. In the present study, we made chimeric constructs of the two genes to determine the above mentioned questions. Our results suggest that the functional specificity of these two ascidian T-box genes is associated with the DNA-binding domain but not with the transcription regulatory domain. The 5' flanking region of both HrBra and HrTbx6 contains T-protein binding motifs near their minimal promoters that are associated with the autoregulative activation of these genes. Using the chimeric constructs, we also determined whether the autoregulative activity is mediated by the DNA-binding domain or by the transcription activation domain of the gene products. Our results suggest that the autoregulative activity of these two ascidian T-box genes is also mediated by the DNA-binding domain, not by the transcription activation domain of the encoded proteins.