Biology subjects

Giuliani, S.

Publications and source records attributed to Giuliani, S..

4 recordsLinked to original sources

bioRxiv2025

Pervasive binding of the stem cell transcription factor SALL4 shapes the chromatin landscape

Mechanistic understanding of how gene activity is regulated has focussed on the roles of transcription factors at promoters and enhancers, whereas mechanisms capable of globally fine-tuning gene expression through dispersed binding across large genomic regions have received less attention. Here we provide evidence that the essential stem cell transcription factor SALL4 modulates gene expression according to DNA base composition by reading the frequency of its AT-rich target motifs. Using an acute depletion strategy, we establish that SALL4-repressed genes localise to AT-rich genomic domains with high levels of dispersed SALL4 occupancy. While SALL4 is localised within peaks and distributed broadly across the genome, explainable machine learning revealed that its occupancy across the gene body is a strong predictor of transcriptional output. We observed rapid increases in chromatin accessibility and histone acetylation independent of transcriptional activity, suggesting that SALL4 primarily acts upon chromatin, while transcriptional changes are secondary. Accordingly, preventing SALL4 from recruiting the histone deacetylase and nucleosome remodelling corepressor NuRD mimicked a Sall4-null phenotype in stem cells and animal models. Our findings reveal that SALL4s interpretation of DNA sequence optimises the global epigenome and transcriptome, a process integral to maintaining the stem cell gene expression programme.

genomics↗

bioRxiv2025

Tetramerisation governs SALL transcription factor function in development and disease

Spalt-like (SALL) proteins are C2H2 zinc-finger transcription factors important for embryogenesis, with mutations in SALL1 and SALL4 causing rare congenital disorders Townes-Brocks and Okihiro syndromes, respectively. While SALL proteins are known to associate with one another, the biological significance of the resulting complexes is unknown. Here we define a conserved glutamine-rich region that mediates SALL1/4 homo- and heterotetramerisation and find that complex formation is indispensable for DNA binding. Modelling a patient mutation that abolishes SALL4 multimerisation led to gene misregulation and, in mice, embryonic lethality, therefore phenocopying a complete Sall4 knockout. Furthermore, a common disease-causing SALL1 truncation, which retains multimerisation but lacks DNA-binding domains, sequesters SALL4 into heterotetramers that are defective in DNA binding, thereby providing a mechanistic explanation for the dominant-negative effects of many Townes-Brocks mutations. Together, our findings establish tetramerisation as a prerequisite for SALL function, linking complex formation to developmental gene regulation and human disease.

molecular biology↗

bioRxiv2025

A modular toolbox for in cellulo screening of small molecule inhibitors targeting chromatin reader domains

The dysregulation of bromodomain proteins, a family of "reader" proteins that recognize the critical post-translational modification of acylation, is implicated in diseases like cancer, making them important therapeutic targets. However, the development of specific small-molecule inhibitors is hindered by the lack of robust, high-throughput cellular assays to measure target engagement and off-target binding in living cells. To address this gap, we developed a modular platform of cell lines that stably express synthetic chromatin reader constructs, termed Acyl-eCRs, containing various bromodomains fused to eGFP. We demonstrate that these Acyl-eCRs recapitulate the same response to bromodomain inhibitors and PROTACs as endogenous proteins, allowing for the quantitative assessment of drug effects. We introduce two complementary flow cytometry-based assays to evaluate inhibitor-target engagement: a competitive binding assay leveraging PROTAC-induced degradation, and a nuclear retention assay that directly measures the displacement of bromodomains from chromatin. Our approach circumvents the need for laborious protein purification and in vitro characterization, providing a scalable and physiologically relevant method for assessing inhibitor potency and specificity. This platform represents a versatile tool for chemical biology, enabling the functional evaluation of chromatin-targeting drugs in a native cellular context.

molecular biology↗

bioRxiv2018

Common loci underlie natural variation in diverse toxin responses

Phenotypic complexity is caused by the contributions of environmental factors and multiple genetic loci, interacting or acting independently. Studies of yeast and Arabidopsis found that the majority of natural variation across phenotypes is attributable to independent additive quantitative trait loci (QTL). Detected loci in these organisms explain most of the estimated heritable variation. By contrast, many heritable components underlying phenotypic variation in metazoan models remain undetected. Before the relative impacts of additive and interactive variance components on metazoan phenotypic variation can be dissected, high replication and precise phenotypic measurements are required to obtain sufficient statistical power to detect loci contributing to this missing heritability. Here, we used a panel of 296 recombinant inbred advanced intercross lines of Caenorhabditis elegans and a high-throughput fitness assay to detect loci underlying responses to 16 different toxins, including heavy metals, chemotherapeutic drugs, pesticides, and neuropharmaceuticals. Using linkage mapping, we identified 82 QTL that underlie variation in responses to these toxins and predicted the relative contributions of additive loci and genetic interactions across various growth parameters. Additionally, we identified three genomic regions that impact responses to multiple classes of toxins. These QTL hotspots could represent common factors impacting toxin responses. We went further to generate near-isogenic lines and chromosome-substitution strains and then experimentally validated these QTL hotspots, implicating additive and interactive loci that underlie toxin-response variation.

genetics↗