Bodega, B. | Search bioRxiv

bioRxiv2024

CFC-seq: identification of full-length capped RNAs unveil enhancer-derived transcription

Long-read sequencing has transformed transcriptome profiling, yet capturing full-length, non-polyadenylated transcripts like enhancer RNAs (eRNAs) remains challenging. Here, we introduce CFC-seq, combining cap-trapping and in vitro poly(A)-tailing to sequence poly(A) and non-poly(A) RNAs with precise transcription start site. Paired with our assembler, SALA, we identified 39,425 novel transcriptional units, including [~]24,000 eRNAs. Our data reveal a distinct genomic code governing eRNA fate dictated by core promoter architecture. CpG-island enhancers show high chromatin connectivity but yield short, exosome-sensitive RNAs. Conversely, TATA-box enhancers systematically co-opt LTR retrotransposons to inherit structural motifs that produce long, stable, and spliced RNAs. Mechanistically, the pioneer factor NF-Y activates these viral elements to license transcription, balanced by TEAD4 activity across a dual-gear regulatory axis. Finally, non-poly(A) eRNAs terminate via exosome-associated processing at structural-depleted cleavage zones. This comprehensive annotation links enhancer sequence architecture to RNA fate, providing a new transformative framework for decoding the functional human genome. HighlightsO_LIExpanded genomic architecture: CFC-seq unmasks a hidden layer of human transcriptome, identifying 39,425 novel transcriptional units with high-confidence TSS support, including [~]24,000 eRNAs. C_LIO_LITSS-first assembler: We introduce SALA, a specialized long-read assembler that prioritizes authentic 5 Cap-trapped ends to accurately reconstruct the TSS-resolved transcript models. C_LIO_LIGenomic code of eRNA fate: CGI enhancers drive short and exosome-sensitive transcripts associated with repressive H3K27me3 mark and high chromatin connectivity. TATA-box enhancers produce cell-type-specific, long, stable, and frequently spliced eRNAs. C_LIO_LIEvolutionary co-option of retrotransposons: A major fraction of TATA-box eRNAs originate from LTR retrotransposons, providing a direct mechanism for integration of viral elements into the human regulatory landscape. C_LIO_LIA dual-gear pioneering axis: The pioneer factor NF-Y activates unprimed LTR-TATA enhancers to license transcription independent of histone acetylation cascades, operating in parallel with TEAD4-mediated activation. C_LIO_LIStructural determinants of eRNA termination: Non-poly(A) eRNA TES features a secondary structure depletion zone that coordinates pol II termination and calibrates exosome-mediated turnover. C_LI

genomics↗

bioRxiv2022

IRescue: single cell uncertainty-aware quantification of transposable elements expression

Transposable elements (TEs) are mobile DNA repeats that contribute to the evolution of eukaryotic genomes. In complex organisms, TE expression is tissue specific. However, their contribution to cellular heterogeneity is still unknown and challenging to investigate in single-cell RNA sequencing (scRNA-seq), due to the ubiquity and homology of TEs in the genome. We introduce IRescue (Interspersed Repeats single-cell quantifier), the first software that accurately estimates the expression of TE subfamilies at single-cell level, implementing a UMI deduplication algorithm to allocate reads ambiguously mapped on TEs, while correcting for UMI sequencing errors. Applying IRescue on simulated datasets and real scRNA-seq of colorectal cancers, we could precisely estimate TE subfamilies expression. We show that IRescue improves the definition of cellular heterogeneity, detecting TE expression signatures and specific TE-containing splicing isoforms.

bioinformatics↗

bioRxiv2019

4q-D4Z4 chromatin architecture regulates the transcription of muscle atrophic genes in FSHD

Despite increasing insights in genome structure organization, the role of DNA repetitive elements, accounting for more than two thirds of the human genome, remains elusive. Facioscapulohumeral Dystrophy (FSHD) is associated with deletion of D4Z4 repeat array below 11 units at 4q35.2. It is known that the deletion alters chromatin structure in cis, leading to genes upregulation. Here we show a genome-wide role of 4q-D4Z4 array in modulating gene expression via 3D nuclear contacts. We have developed an integrated strategy of 4q-D4Z4 specific 4C-seq and chromatin segmentation analyses, showing that 4q-D4Z4 3D interactome and chromatin states of interacting genes are impaired in FSHD1 condition; in particular, genes which have lost the 4q-D4Z4 interaction and with a more active chromatin state are enriched for muscle atrophy transcriptional signature. Expression level of these genes is restored by the interaction with an ectopic 4q-D4Z4 array, suggesting that the repeat directly modulates the transcription of contacted targets.\n\nOf note, the upregulation of atrophic genes is a common feature of several FSHD1 and FSHD2 patients, indicating that we have identified a core set of deregulated genes involved in FSHD pathophysiology.

genomics↗