| Autor: |
Monastirioti M; Institute of Molecular Biology and Biotechnology (IMBB), Foundation for Research and Technology-Hellas (FORTH), 70013 Heraklion, Greece., Koltsaki I; Institute of Molecular Biology and Biotechnology (IMBB), Foundation for Research and Technology-Hellas (FORTH), 70013 Heraklion, Greece.; Department of Biology, University of Crete, 70013 Heraklion, Greece.; Division of Tumor Metabolism and Microenvironment, German Cancer Research Center (DKFZ), 69120 Heidelberg, Germany., Pitsidianaki I; Institute of Molecular Biology and Biotechnology (IMBB), Foundation for Research and Technology-Hellas (FORTH), 70013 Heraklion, Greece.; Department of Biology, University of Crete, 70013 Heraklion, Greece.; Department of Cell and Developmental Biology, University College London (UCL), London WC1E 6BT, UK., Skafida E; Institute of Molecular Biology and Biotechnology (IMBB), Foundation for Research and Technology-Hellas (FORTH), 70013 Heraklion, Greece.; Department of Biology, University of Crete, 70013 Heraklion, Greece.; Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS), Foundation Saint Lucia, Rome and School of Medicine and Surgery, University of Milano-Bicocca (UniMiB), 20900 Monza, Italy., Batsiotos N; Institute of Molecular Biology and Biotechnology (IMBB), Foundation for Research and Technology-Hellas (FORTH), 70013 Heraklion, Greece.; Department of Biology, University of Crete, 70013 Heraklion, Greece.; Evotec SE, 22419 Hamburg, Germany., Delidakis C; Institute of Molecular Biology and Biotechnology (IMBB), Foundation for Research and Technology-Hellas (FORTH), 70013 Heraklion, Greece.; Department of Biology, University of Crete, 70013 Heraklion, Greece. |
| Abstrakt: |
Drosophila Hey is a basic helix-loop-helix-orange (bHLH-O) protein with an important role in the establishment of distinct identities of postmitotic cells. We have previously identified Hey as a transcriptional target and effector of Notch signalling during the asymmetric division of neuronal progenitors, generating neurons of two types, and we have shown that Notch-dependent expression of Hey also marks a subpopulation of the newborn enteroendocrine (EE) cells in the midgut primordium of the embryo. Here, we investigate the transcriptional regulation of Hey in neuronal and intestinal tissues. We isolated two genomic regions upstream of the promoter (HeyUP) and in the second intron (HeyIN2) of the Hey gene, based on the presence of binding motifs for Su(H), the transcription factor that mediates Notch activity. We found that both regions can direct the overlapping expression patterns of reporter transgenes recapitulating endogenous Hey expression. Moreover, we showed that while HeyIN2 represents a Notch-dependent enhancer, HeyUP confers both Notch-dependent and independent transcriptional regulation. We induced mutations that removed the Su(H) binding motifs in either region and then studied the enhancer functionality in the respective Hey mutant lines. Our results provide direct evidence that although both enhancers support Notch-dependent regulation of the Hey gene, their role is redundant, as a Hey loss-of-function lethal phenotype is observed only after deletion of all their Su(H) binding motifs by CRISPR/Cas9. |
