Structure and Mechanism of Notch Transcription Complex Dimerization in Gene Regulation

Abstract

Notch signaling triggers cell growth, differentiation, and death. Deficiency in Notch genes is embryonic lethal in mice, and aberrant Notch signals have been implicated in many human diseases. During activation, Notch is cleaved from the membrane and enters the nucleus where it cooperates with the DNA-binding transcription factor CSL and the transcriptional coactivator MAML, to form the Notch transciption complex (NTC). Recently, our lab observed dimerization of NTCs on a segment of DNA from the HES1 promoter region. This head-to-head pairing of CSL sites (called an SPS site for Su(H) paired site or sequence paired site) is found in the promoters of a number of well-characterized Notch targets in Drosophila, Xenopus, and in the Hes1 gene in mammals. The goal of this work is to determine the structural and energetic foundations underlying Notch mediated dimerization, and ultimately to understand how NTC dimerization influences the expression of different Notch targets. Toward this goal, the structure of a dimer of Notch transcription complex trimers on DNA has been determined to 3.5A resolution. Each of the two NTC complexes superimposes well on an NTC bound to a single site and undergo little conformational change, but the two NTC are twisted relative to each other and the DNA untwists and bends away from the protein binding interface. Although HES1 is one of the most well characterized mammalian Notch target genes, the role of Notch dimerization in other contexts is unexplored. To begin to address this question, work has begun to identify other functional paired sites and to characterize the range of DNA duplexes able to cooperatively bind dimers of Notch Transcriptions complexes.