Promoter Repression and 3D-Restructuring Resolves Divergent Developmental Gene Expression in TADs

Ringel, Alessa; Szabo, Quentin; Chiariello, Andrea M.; Chudzik, Konrad; Schöpflin, Robert; Rothe, Patricia; Mattei, Alexandra L.; Zehnder, Tobias; Harnett, Dermot; Laupert, Verena; Bianco, Simona; Hetzel, Sara; Phan, Mai H Q; Schindler, Magdalena; Ibrahim, Daniel M.; Paliou, Christina; Esposito, Andrea; Prada-Medina, César Augusto; Haas, Stefan A.; Giere, Peter; Vingron, Martin; Wittler, Lars; Meissner, Alexander; Nicodemi, Mario; Cavalli, Giacomo; Bantignies, Frédéric; Mundlos, Stefan; Robson, Michael I.

doi:10.2139/ssrn.3947354

Cited by 3 publications

(3 citation statements)

References 104 publications

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“…The ubiquitous pattern was observed for 4/6 lines (Figure S3). For CRE to have induced a ubiquitous YFP pattern, excision of the stop cassette must have occurred at very early stages (one to two cell stage depending on the percentage of recombined cells), indicating that the transgene (CRE and mTomato) is expressed at the 1-2 cells stage, supporting the possibility that Fat1 may be expressed at these stages, consistent with its known expression in embryonic stem cells [35]. The co-occurrence of two YFP patterns in these lines was maintained after several generations, and we did not observe segregation over time of two different patterns of YFP expression, nor a progressive restriction of the pattern from generation to generation.…”

Section: Resultsmentioning

confidence: 86%

“…Under the light of work discussed above, suggesting that the component of FAT1 expression domain relevant to muscle morphogenesis, and possibly to FSHD symptoms, might occur in fibroblast/mesenchymal lineage rather than (or in addition to) in myoblasts [12, 38], it will be interesting to study in future whether the changes in FAT1 regulation occur in mesenchymal/fibroblast lineage rather than in myoblasts. Furthermore, FAT1 is located in a conserved genomic region, between FRG1 and SORBS2 , now identified as a TAD [35, 58], in which long-range interactions allow shaping gene regulation. Thus, the FAT1 locus represents a typical situation in which shadow enhancers with similar tissue-specificity may cumulate their activities to ensure the robustness of the expression pattern [69, 70].…”

Section: Discussionmentioning

confidence: 99%

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An intragenic FAT1 regulatory element deleted in muscular dystrophy patients drives muscle and mesenchyme expression during development

Caruso

Zimmermann

Nigam

et al. 2022

Preprint

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SummaryFat1 is an atypical cadherin playing multiple roles that influence tissue morphogenesis. During mouse development Fat1 is required to modulate muscle morphogenesis through complementary activities in myogenic cells, muscle-associated connective tissue, and motor neurons, ablation of which leads to regionalized muscle phenotypes. We previously identified copy number variants (CNV) deleting an intragenic conserved non-coding element (CNE) in the human FAT1 locus, that were enriched among muscular dystrophy patients with symptoms resembling those of Facioscapulohumeral Dystrophy (FSHD), compared to healthy individuals. Since such deletions of a putative cis-regulatory element had the potential to cause tissue-specific depletion of FAT1, they were postulated to act as symptom modifiers. However, activity of this CNE has not been functionally explored so far. To investigate the possible regulatory activity of this Fat1-CNE, we engineered transgenic mice in which it drives expression of a bi-cistronic reporter comprising the CRE-recombinase (Cre) and a myristilated-tdTomato fluorescent protein. The tissue-specific pattern of cre and tomato expression indicates that this enhancer has bipotential character, and drives expression in skeletal muscle and in muscle-associated mesenchymal cells. We extended our analysis of one of the transgenic lines, which exhibits enhanced expression in mesenchymal cells at extremities of subsets of muscles matching the map of Fat1-dependent muscles. This transgenic line exhibits highly selective CRE-mediated excision in scattered cells within the Tomato-positive territory hotspots. This represents a novel tool to genetically explore the diversity of muscle-associated mesenchymal lineages.

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Section: Resultsmentioning

confidence: 86%

Section: Discussionmentioning

confidence: 99%

An intragenic FAT1 regulatory element deleted in muscular dystrophy patients drives muscle and mesenchyme expression during development

Caruso

Zimmermann

Nigam

et al. 2022

Preprint

View full text Add to dashboard Cite

show abstract

“…We grouped the 22 mutants, all homozygous, into four rough categories (Supplementary Table 1): 1) pleiotropic mutants, representing knockouts of developmental genes expressed in multiple organs (Ttc21b KO, Carm1 KO, Gli2 KO), as well as two mutations of the Sox9 regulatory landscape suspected to have pleiotropic effects, both of which effectively result in the introduction of a boundary element between endogenous Sox9 enhancers and the Sox9 promoter (Sox9 TAD boundary KI; Sox9 regulatory INV) [27][28][29][30] . 2) developmental disorder mutants, intended to model specific human diseases (Scn11a GOF, Ror2 KI, Gorab KO, Cdkl5 -/Y) [31][32][33] , 3) mutations of loci associated with human disease (Scn10a/Scn11a DKO, Atp6v0a2 KO, Atp6v0a2 R755Q, Fat1TAD KO) 34,35 . 4) prospective deletions of cis-regulatory elements, including of TAD boundaries in the vicinity of developmental transcription factors including Smad3, Twist1, Tbx5, Neurog2, Sim1, Smad7, Dmrt1, Tbx3, and Twist1 36 , and, as a positive control, the ZRS distal enhancer (Zone of polarizing activity Regulatory Sequence) which regulates sonic hedgehog (SHH) expression and results in absent distal limb structures 37 .…”

Section: Single-cell Rna-seq Of 101 Mouse Embryosmentioning

confidence: 99%

Single cell, whole embryo phenotyping of pleiotropic disorders of mammalian development

Huang

Henck

Qiu

et al. 2022

Preprint

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Mouse models are a critical tool for studying human diseases, particularly developmental disorders, as well as for advancing our general understanding of mammalian biology. However, it has long been suspected that conventional approaches for phenotyping are insufficiently sensitive to detect subtle defects throughout the developing mouse. Here we set out to establish single cell RNA sequencing (sc-RNA-seq) of the whole embryo as a scalable platform for the systematic molecular and cellular phenotyping of mouse genetic models. We applied combinatorial indexing-based sc-RNA-seq to profile 101 embryos of 26 genotypes at embryonic stage E13.5, altogether profiling gene expression in over 1.6M nuclei. The 26 genotypes include 22 mouse mutants representing a range of anticipated severities, from established multisystem disorders to deletions of individual enhancers, as well as the 4 wildtype backgrounds on which these mutants reside. We developed and applied several analytical frameworks for detecting differences in composition and/or gene expression across 52 cell types or trajectories. Some mutants exhibited changes in dozens of trajectories (e.g., the pleiotropic consequences of altering the Sox9 regulatory landscape) whereas others showed phenotypes affecting specific subsets of cells. We also identify differences between widely used wildtype strains, compare phenotyping of gain vs. loss of function mutants, and characterise deletions of topological associating domain (TAD) boundaries. Intriguingly, even among these 22 mutants, some changes are shared by heretofore unrelated models, suggesting that developmental pleiotropy might be "decomposable" through further scaling of this approach. Overall, our findings show how single cell profiling of whole embryos can enable the systematic molecular and cellular phenotypic characterization of mouse mutants with unprecedented breadth and resolution.

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Enhancer-gene specificity in development and disease

2022

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Enhancers control the establishment of spatiotemporal gene expression patterns throughout development. Over the past decade, the development of new technologies has improved our capacity to link enhancers with their target genes based on their colocalization within the same topological domains. However, the mechanisms that regulate how enhancers specifically activate some genes but not others within a given domain remain unclear. In this Review, we discuss recent insights into the factors controlling enhancer specificity, including the genetic composition of enhancers and promoters, the linear and 3D distance between enhancers and their target genes, and cell-type specific chromatin landscapes. We also discuss how elucidating the molecular principles of enhancer specificity might help us to better understand and predict the pathological consequences of human genetic, epigenetic and structural variants.

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Promoter Repression and 3D-Restructuring Resolves Divergent Developmental Gene Expression in TADs

Cited by 3 publications

References 104 publications

An intragenic FAT1 regulatory element deleted in muscular dystrophy patients drives muscle and mesenchyme expression during development

An intragenic FAT1 regulatory element deleted in muscular dystrophy patients drives muscle and mesenchyme expression during development

Single cell, whole embryo phenotyping of pleiotropic disorders of mammalian development

Enhancer-gene specificity in development and disease

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