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Identification of sequences which regulate the expression of Drosophila melanogaster Doc elements
C Contursi1, G Minchiotti, P P Di Nocera
1Dipartimento di Biologia e Patologie Cellulare e Molecolare L. Califano, Università degli Studi di Napoli Federico II, Italy.
The Journal of Biological Chemistry
|November 3, 1995
Summary
Doc elements, a type of mobile DNA (Long Interspersed Nuclear Elements), show transcription initiation regulated by specific DNA sequences near their start site. Their expression is also repressed by downstream elements, impacting gene translation.
Area of Science:
- Molecular Biology
- Genetics
- Genomics
Background:
- Long Interspersed Nuclear Elements (LINEs) are mobile DNA elements that replicate via RNA intermediates.
- LINEs lack long terminal repeats, and their expression is regulated by internal promoters.
- Doc elements are one of seven LINE families identified in Drosophila melanogaster.
Purpose of the Study:
- To investigate the transcriptional regulation of Doc elements in Drosophila melanogaster.
- To identify specific DNA sequences controlling Doc element transcription initiation and expression.
- To understand the mechanisms of transcriptional repression mediated by downstream Doc sequences.
Main Methods:
- Transient expression assays in Drosophila S2 cells using plasmids containing the CAT gene linked to Doc DNA segments.
- Analysis of transcription initiation sites within Doc elements.
- Site-directed mutagenesis to assess the impact of DNA region positioning on transcription.
Main Results:
- Transcription of Doc elements initiates at specific hexamer sequences ((C/G)AYTCG) within their 5' region.
- A downstream DNA region, approximately 20 bp from the start site, containing DE1, DE2 (RGACGTGY motif), and DE3, is crucial for regulating transcription.
- Altering the position of this downstream region significantly inhibited transcription.
- Sequences ~200 bp downstream repressed CAT expression in an orientation- and position-dependent manner, potentially via impaired translation.
Conclusions:
- Doc element transcription initiation is precisely controlled by internal promoter elements and a closely located downstream regulatory region.
- Downstream sequences play a complex role in regulating Doc element expression, including potential translational repression.
- These findings provide insights into the regulatory mechanisms of mobile DNA elements in Drosophila.