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High-throughput Oligopaint screen identifies druggable 3D genome regulators.

ABSTRACT: The human genome functions as a three-dimensional chromatin polymer, driven by a complex collection of chromosome interactions^1-3. Although the molecular rules governing these interactions are being quickly elucidated, relatively few proteins regulating this process have been identified. Here, to address this gap, we developed high-throughput DNA or RNA labelling with optimized Oligopaints (HiDRO)-an automated imaging pipeline that enables the quantitative measurement of chromatin interactions in single cells across thousands of samples. By screening the human druggable genome, we identified more than 300 factors that influence genome folding during interphase. Among these, 43 genes were validated as either increasing or decreasing interactions between topologically associating domains. Our findings show that genetic or chemical inhibition of the ubiquitous kinase GSK3A leads to increased long-range chromatin looping interactions in a genome-wide and cohesin-dependent manner. These results demonstrate the importance of GSK3A signalling in nuclear architecture and the use of HiDRO for identifying mechanisms of spatial genome organization.

SUBMITTER: Park DS

PROVIDER: S-EPMC11305015 | biostudies-literature | 2023 Aug

REPOSITORIES: biostudies-literature

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High-throughput Oligopaint screen identifies druggable 3D genome regulators.

Park Daniel S DS Nguyen Son C SC Isenhart Randi R Shah Parisha P PP Kim Wonho W Barnett R Jordan RJ Chandra Aditi A Luppino Jennifer M JM Harke Jailynn J Wai May M Walsh Patrick J PJ Abdill Richard J RJ Yang Rachel R Lan Yemin Y Yoon Sora S Yunker Rebecca R Kanemaki Masato T MT Vahedi Golnaz G Phillips-Cremins Jennifer E JE Jain Rajan R Joyce Eric F EF

Nature 20230712 7972

The human genome functions as a three-dimensional chromatin polymer, driven by a complex collection of chromosome interactions<sup>1-3</sup>. Although the molecular rules governing these interactions are being quickly elucidated, relatively few proteins regulating this process have been identified. Here, to address this gap, we developed high-throughput DNA or RNA labelling with optimized Oligopaints (HiDRO)-an automated imaging pipeline that enables the quantitative measurement of chromatin int ...[more]

PMID: 37438531

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Project description:Spinocerebellar Ataxia type 3 (SCA3, also known as Machado-Joseph disease) is a neurodegenerative disorder caused by a CAG repeat expansion encoding an abnormally long polyglutamine (polyQ) tract in the disease protein, ataxin-3 (ATXN3). No preventive treatment is yet available for SCA3. Because SCA3 is likely caused by a toxic gain of ATXN3 function, a rational therapeutic strategy is to reduce mutant ATXN3 levels by targeting pathways that control its production or stability. Here, we sought to identify genes that modulate ATXN3 levels as potential therapeutic targets in this fatal disorder. We screened a collection of siRNAs targeting 2742 druggable human genes using a cell-based assay based on luminescence readout of polyQ-expanded ATXN3. From 317 candidate genes identified in the primary screen, 100 genes were selected for validation. Among the 33 genes confirmed in secondary assays, 15 were validated in an independent cell model as modulators of pathogenic ATXN3 protein levels. Ten of these genes were then assessed in a Drosophila model of SCA3, and one was confirmed as a key modulator of physiological ATXN3 abundance in SCA3 neuronal progenitor cells. Among the 15 genes shown to modulate ATXN3 in mammalian cells, orthologs of CHD4, FBXL3, HR and MC3R regulate mutant ATXN3-mediated toxicity in fly eyes. Further mechanistic studies of one of these genes, FBXL3, encoding a F-box protein that is a component of the SKP1-Cullin-F-box (SCF) ubiquitin ligase complex, showed that it reduces levels of normal and pathogenic ATXN3 in SCA3 neuronal progenitor cells, primarily via a SCF complex-dependent manner. Bioinformatic analysis of the 15 genes revealed a potential molecular network with connections to tumor necrosis factor-α/nuclear factor-kappa B (TNF/NF-kB) and extracellular signal-regulated kinases 1 and 2 (ERK1/2) pathways. Overall, we identified 15 druggable genes with diverse functions to be suppressors or enhancers of pathogenic ATXN3 abundance. Among identified pathways highlighted by this screen, the FBXL3/SCF axis represents a novel molecular pathway that regulates physiological levels of ATXN3 protein.

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High-throughput Oligopaint screen identifies druggable 3D genome regulators.

Publications

High-throughput Oligopaint screen identifies druggable 3D genome regulators.

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