Transcriptional read through interrupts boundary function in Drosophila

Area of Science:

• Genomics and Molecular Biology
• Epigenetics and Chromosome Organization

Background:

• Enhancer-promoter interactions in eukaryotes are typically confined by chromatin insulators, which define topologically associated domains (TADs).
• Mechanisms allowing enhancer-promoter interactions to bypass these boundaries and span multiple TADs remain largely unknown.
• Long non-coding RNAs (lncRNAs) are abundant in the genome and are hypothesized to have regulatory functions, including chromatin organization.

Approach:

• Utilized the well-characterized Drosophila Bithorax complex (BX-C) and its boundary replacement platforms (Fab-7 and F2) to investigate boundary function.
• Engineered constructs with boundary elements, promoters, and polyadenylation signals in various combinations to assess boundary activity.
• Assayed for the impact of transcriptional readthrough on boundary function and enhancer activity within the BX-C.

Key Points:

• Transcriptional readthrough, even at low levels, was found to impair the function of chromatin boundaries within the BX-C.
• Transcription originating from inserted promoters affected the activity of enhancers located within BX-C regulatory domains.
• Boundary replacement platforms allowed for the systematic study of transcriptional interference with regulatory element function.

Conclusions:

• Transcriptional readthrough can interfere with the activity of chromatin insulators, challenging the integrity of topologically associated domains (TADs).
• This mechanism provides a potential explanation for how enhancer-promoter interactions can occur across TAD boundaries.
• The findings suggest that lncRNAs, as a significant fraction of Pol II transcripts, may actively regulate chromosome structure and gene expression through transcriptional readthrough.