Molecular characterization of HES‐2, a mammalian helix‐loop‐helix factor structurally related to Drosophila hairy and Enhancer of split

Abstract

Drosophila hairy (h) plays a crucial role in early development as a pair‐rule segmentation gene. h and its structurally related gene Enhancer of split [E(spl)] are also required for normal sensory neurogenesis in late development. To analyze the molecular mechanisms of mammalian development, we recently characterized three rat helix‐loop‐helix (HLH) factors that show structural homology to the Drosophila h and E(spl) gene products, and found that rat factors exhibit distinct spatiotemporal expression patterns and act as a negative regulator. Here, we report the molecular characterization of another member of this family, designated HES‐2. Rat HES‐2 protein has a basic HLH domain homologous to h and E(spl) as well as the carboxy‐terminal Trp‐Arg‐Pro‐Trp sequence conserved among this family. The HES‐2 mRNA is present as early as embryonic day 9.5 and is detected in a variety of tissues of both embryos and adults. DNase‐I‐footprinting analyses indicate that HES‐2 binds to all E box sequences (CANNTG) we tested as well as to the N‐box sequences (CACNAG). Further studies of gel‐mobility‐shift assays show that HES‐2 has a higher affinity for the E box than for the N box. Transient transfection analyses suggest that HES‐2 decreases the transcription originating from the promoters containing either the E box or the N box. These results indicate that HES‐2 acts as a negative regulator through interaction with both E‐box and N‐box sequences. Copyright © 1993, Wiley Blackwell. All rights reserved