Function of the TAD/PEST Domains in the Mouse Notchl and Notch 4 Genes

Abstract

The Notch gene, cell-surface protein, is necessary for cell-fate decisions in many developing organisms. There are multiple homologs in mammals, including four Notch proteins in mice. Improper signaling has been linked to neoplasia and possibly schizophrenia. After Notch has interacted with ligand, the intracellular portion of Notch is released from the cell surface and enters the nucleus. In the nucleus, Notch interacts with CBFl, Su(H), and Lag-l proteins (CSL), upregulating downstream targets. This association is also modified via interactions with other activating transcriptional proteins. Many of Notch's specific interactions with CSL, however, are unknown. Previous experiments show that Notch4 exhibits greater activation of known-CSL dependent pathways when the T ADIPEST domain, a large portion of the C-terminal domain, is removed. Therefore, we made a construct that placed the Notch4 T ADIPEST domain onto the corresponding region of Notch 1; this chimera was named N1//N4. The signaling efficacy of these constructs was tested using three different reporters. Contrasting the ability of N1/N4 to activate of the reporter constructs, we were able to determine possible functions for the Notch1 and Notch4 TAD/PEST domains. We found that Notch activity is promoter dependent and that the function of the Notch4 T ADIPEST domain, although inhibitory in certain environments, was not transferable to Notch1. We believe that the TAD/PEST domain of Notch1 and Notch4, as well as the promoter, dictates which interactions with different transcriptional proteins will occur. Further testing is necessary to elucidate these unknown proteins and the specific functions of the TAD/PEST domains of Notch1 and Notch4.