Structural and Functional Dissection of the 5’ Region of the Notch Gene in Drosophila melanogaster
| Autor: | Sergey A. Demakov, Darya S. Sidorenko, Igor F. Zhimulev, Natalya G. Andreyenkova, Oleg V. Andreyenkov, E. I. Volkova |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
polytene chromosomes 030102 biochemistry & molecular biology biology Cellular differentiation CRISPR/Cas9 system biology.organism_classification Cell biology interbands 03 medical and health sciences Exon 030104 developmental biology Notch gene Notch proteins CTCF gene activity regulation open chromatin state Genetics CRISPR Drosophila melanogaster Homologous recombination Gene insulator proteins Genetics (clinical) |
| Zdroj: | Genes Volume 10 Issue 12 |
| ISSN: | 2073-4425 |
| DOI: | 10.3390/genes10121037 |
| Popis: | Notch is a key factor of a signaling cascade which regulates cell differentiation in all multicellular organisms. Numerous investigations have been directed mainly at studying the mechanism of Notch protein action however, very little is known about the regulation of activity of the gene itself. Here, we provide the results of targeted 5&rsquo end editing of the Drosophila Notch gene in its native environment and genetic and cytological effects of these changes. Using the Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated protein 9 (CRISPR/Cas9) system in combination with homologous recombination, we obtained a founder fly stock in which a 4-kb fragment, including the 5&rsquo nontranscribed region, the first exon, and a part of the first intron of Notch, was replaced by an attachment Phage (attP) site. Then, fly lines carrying a set of six deletions within the 5&rsquo untranscribed region of the gene were obtained by &Phi C31-mediated integration of transgenic constructs. Part of these deletions does not affect gene activity, but their combinations with transgenic construct in the first intron of the gene cause defects in the Notch target tissues. At the polytene chromosome level we defined a DNA segment (~250 bp) in the Notch5&rsquo nontranscribed region which when deleted leads to disappearance of the 3C6/C7 interband and elimination of CTC-Factor (CTCF) and Chromator (CHRIZ) insulator proteins in this region. |
| Databáze: | OpenAIRE |
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