Biology subjects

Viales, R. R.

Publications and source records attributed to Viales, R. R..

7 recordsLinked to original sources

bioRxiv2025

SPT5 regulates Pol II pausing and elongation in different ways at early versus late embryonic stages

Transcription involves a cycle of initiation, pausing, elongation, and termination. SPT5 regulates both promoter proximal pausing and elongation, but how it orchestrates both steps during dynamic changes in gene expression remains unclear. Here, using Drosophila embryogenesis, we show that pausing both precedes and follows gene expression, while active transcription is accompanied by pause release. Optogenetic rapid depletion of SPT5 from the nucleus uncovered different sensitivities at different developmental stages. In early embryogenesis, SPT5 depletion caused a downstream shift in Pol II pausing to the +1 nucleosome, resulting in defective elongation and early termination. In late embryogenesis, it led to both up- and downregulation of expression, depending on the genes transcriptional and pausing state - upregulation is caused by pause release while downregulation is due to defective elongation. These changes are intensified when genes are increasing or decreasing their transcriptional state, indicating that SPT5 contributes to fine-tuning dynamic changes in gene expression.

genomics↗

bioRxiv2025

Chip (Ldb1) is a new cofactor of Zelda forming a functional bridge to CBP during zygotic genome activation

The cofactor Ldb1 (Chip) is linked to many processes in gene regulation, including enhancer-promoter communication, inter-chromosomal interactions and enhanceosome-cofactor-like activity. However, its functional requirement and molecular role during embryogenesis has not been assessed to date. Here, we used optogenetics (iLEXY), to rapidly deplete Drosophila Ldb1 (Chip) from the nucleus at precise time windows. Remarkably, this pinpointed the essential window of Chips function in just one-hour of embryogenesis, overlapping zygotic genome activation (ZGA). We show that Zelda, a pioneer factor essential for ZGA, recruits Chip to chromatin, and both factors regulate concordant changes in gene expression, suggesting that Chip is a cofactor of Zelda. Surprisingly, Chip is not required for chromatin architecture at these stages, instead it recruits CBP, and is essential for the placement of H3K27ac. Taken together, our results identify Chip (Ldb1) as a functional bridge between Zelda and the coactivator CBP to regulate gene expression in early embryogenesis.

genomics↗

bioRxiv2024

Contextualising transcription factor binding during embryogenesis using natural sequence variation

Understanding how genetic variation impacts transcription factor (TF) binding remains a major challenge, limiting our ability to model disease-associated variants. Here, we used a highly controlled system of F1 crosses with extensive genetic diversity to profile allele-specific binding of four TFs at several embryonic time-points, using Drosophila as a model. Using a combined haplotype test, we identified 9-18% of TF bound regions impacted by genetic variation. By expanding WASP (a tool for allele-specific read mapping) to examine INDELs, we increased detection of allele imbalanced (AI) peaks by 30-50%. This fine-grained mutagenesis could reconstruct functionalized binding motifs of all factors. To prioritise potential causal variants, we trained a convolutional neural network (Basenji) to predict TF binding from DNA sequence. The model could accurately predict experimental AI for strong effect variants, providing a mechanistic interpretation for how genetic variation impacted TF binding. This revealed unexpected relationships between TFs, including potential cooperative pairs, and mechanisms of tissue specific recruitment of the ubiquitous factor CTCF.

genetics↗

bioRxiv2024

A function of Spalt proteins in heterochromatin organization and maintenance of genomic DNA integrity

The phylogenetically conserved Spalt proteins regulate gene expression and participate in a variety of cell fate choices during multicellular development, generally acting as transcriptional repressors in different gene regulatory networks. Paradoxically, besides their roles as DNA sequence-specific transcription factors, Spalt proteins show a consistent localization to heterochromatic regions. They can act through interactions with the Nucleosome remodeling and deacetylase complex (NuRD) to promote closing of open chromatin domains, but their activities as epigenetic regulators also rely on interactions with DNA Methyltransferases or with the Lysine-specific histone demethylase LSD1, suggesting that they can participate in multiple regulatory mechanisms. Here we describe several major consequences of loss of spalt function in Drosophila cells, including changes in chromatin accessibility affecting mostly pericentromeric heterochromatin, the generation of DNA damage, alterations in the localization of chromosomes within the nucleus in polyploid cells of the salivary glands and miss-expression of transposable elements. We suggest that most of these effects are related to roles of Spalt proteins in the regulation of heterochromatin formation. We propose that Drosophila Spalt proteins have two complementary functions, acting as sequence-specific transcriptional repressors on specific target genes and regulating more global gene silencing through the generation or maintenance of heterochromatic domains.

developmental biology↗

bioRxiv2022

New enhancer-promoter interactions are gained during tissue differentiation and reflect changes in E/P activity

To regulate gene expression, enhancers must come into proximity with their target gene. At some loci the timing of enhancer-promoter proximity is uncoupled from gene activation, while at others it is tightly linked. Here, we assessed this more globally for 600 characterized enhancers or promoters (E/P) with tissue-specific activity in Drosophila embryos, by performing Capture-C and insulator ChIP in FACS-purified myogenic or neurogenic cells at different stages of embryogenesis. This high-resolution view enabled direct comparison between E/P interactions and activity across 5 developmental conditions. This revealed largely invariant E/P contacts between the blastoderm and cell fate specification stages, despite changes in activity. However, E/P interactions diverge during terminal tissue differentiation when many tissue-specific interactions are gained on top of a pre-existing topology. Changes in E/P proximity reflect changes in enhancer activity and gene activation, and are generally not accompanied by changes in insulator binding. Using transgenes and deletions, we show that many tissue-specific interactions represent functional E-P pairs. Our results reveal a shift in E-P landscapes as embryogenesis proceeds, from largely pre-formed topologies at early stages to more distal tissue-specific loops during differentiation, when E/P proximity appears coupled to activation.

developmental biology↗

bioRxiv2020

The hourglass model of evolutionary conservation during embryogenesis extends to developmental enhancers with signatures of positive selection

Inter-species comparisons of both morphology and gene expression within a phylum have revealed a period in the middle of embryogenesis with more similarity between species compared to earlier and later time-points. This developmental hourglass pattern has been observed in many phyla, yet the evolutionary constraints on gene expression, and underlying mechanisms of how this is regulated, remains elusive. Moreover, the role of positive selection on gene regulation in the more diverged earlier and later stages of embryogenesis remains unknown. Here, using DNase-seq to identify regulatory regions in two distant Drosophila species (D. melanogaster and D. virilis), we assessed the evolutionary conservation and adaptive evolution of enhancers throughout multiple stages of embryogenesis. This revealed a higher proportion of conserved enhancers at the phylotypic period, providing a regulatory basis for the hourglass expression pattern. Using an in silico mutagenesis approach, we detect signatures of positive selection on developmental enhancers at early and late stages of embryogenesis, with a depletion at the phylotypic period, suggesting positive selection as one evolutionary mechanism underlying the hourglass pattern of animal evolution.

evolutionary biology↗

bioRxiv2020

Cis acting variation is common, can propagates across multiple regulatory layers, but is often buffered in developmental programs

Precise patterns of gene expression are driven by interactions between transcription factors, regulatory DNA sequence, and chromatin. How DNA mutations affecting any one of these regulatory layers is buffered or propagated to gene expression remains unclear. To address this, we quantified allele-specific changes in chromatin accessibility, histone modifications, and gene expression in F1 embryos generated from eight Drosophila crosses, at three embryonic stages, yielding a comprehensive dataset of 240 samples spanning multiple regulatory layers. Genetic variation in cis-regulatory elements is common, highly heritable, and surprisingly consistent in its effects across embryonic stages. Much of this variation does not propagate to gene expression. When it does, it acts through H3K4me3 or alternatively through chromatin accessibility and H3K27ac. The magnitude and evolutionary impact of mutations is influenced by a genes regulatory complexity (i.e. enhancer number), with transcription factors being most robust to cis-acting, and most influenced by trans-acting, variation. Overall, the impact of genetic variation on regulatory phenotypes appears context-dependent even within the constraints of embryogenesis.

genomics↗