Biology subjects

Blobel, G.

Publications and source records attributed to Blobel, G..

11 recordsLinked to original sources

bioRxiv2026

LDB1-dependent enhancer connectivity defines T-cell leukemia identities and masks metabolic vulnerabilities

Spatial enhancer connectivity is fundamental to proper gene regulation. Enhancer dysregulation has emerged as a hallmark of cancers, including T-cell acute lymphoblastic leukemias (T-ALL). T-ALL are aggressive malignancies characterized by marked transcriptional heterogeneity driven by distinct stages of developmental arrest and diverse noncoding alterations. How these cancers co-opt nuclear architecture to rewire enhancer connectivity remains poorly understood. Here, we report that the LDB1 chromatin architectural complex is an essential mediator of enhancer-oncogene looping that sustains oncogenic transcriptional programs across multiple T-ALL subtypes. Integrating bulk and single-cell transcriptomic data from patients with T-ALL and healthy hematopoietic controls, we show that the LDB1-dependent regulatory circuitry defines the molecular identities of distinct T-ALL subtypes while restricting plasticity toward alternative cell states. LDB1 loss dismantles chromatin looping among cell state-defining enhancers liberating them to form promiscuous interactions with nearby genes. This enhancer rewiring stimulates expression of key metabolic genes, creating a mevalonate pathway dependency exploitable with statin treatment. Our study establishes LDB1 as a central executor of T-ALL regulatory circuitry and more broadly illustrates chromatin rewiring as a source of targetable dependencies in cancer.

Cancer Biology↗

bioRxiv2025

Cis-regulatory chromatin contacts form de novo in the absence of loop extrusion

NIPBL promotes chromatin loop extrusion by the cohesin complex until it stalls at convergently oriented CTCF sites, leading to the formation of structural loops. However, to what extent loop extrusion contributes to the establishment vs maintenance of cis-regulatory element (CRE) connectivity is poorly understood. Here, we explored the de novo establishment of chromatin folding patterns at the mitosis-to-G1-phase transition upon acute NIPBL loss. NIPBL depletion primarily impaired the formation of cohesin-mediated structural loops with NIPBL dependence being proportional to loop length. In contrast, the majority of CRE loops were established independently of loop extrusion regardless of length. However, NIPBL depletion slowed the re-formation of CRE loops with weak enhancers. Transcription of genes at NIPBL-independent loop anchors was activated normally in the absence of NIPBL. In sum, establishment of most regulatory contacts and gene transcription following mitotic exit is independent of loop extrusion.

molecular biology↗

bioRxiv2024

Dynamics of microcompartment formation at the mitosis-to-G1 transition

As cells exit mitosis and enter G1, mitotic chromosomes decompact and transcription is reestablished. Previously, Hi-C studies showed that essentially all interphase 3D genome features including A/B-compartments, TADs, and CTCF loops, are lost during mitosis. However, Hi-C remains insensitive to features such as microcompartments, nested focal interactions between cis-regulatory elements (CREs). We therefore applied Region Capture Micro-C to cells from mitosis to G1. Unexpectedly, we observe microcompartments in prometaphase, which further strengthen in ana/telophase before gradually weakening in G1. Loss of loop extrusion through condensin depletion differentially impacts microcompartments and large A/B-compartments, suggesting that they are partially distinct. Using polymer modeling, we show that microcompartment formation is favored by chromatin compaction and disfavored by loop extrusion activity, explaining why ana/telophase likely provides a particularly favorable environment. Our results suggest that CREs exhibit intrinsic homotypic affinity leading to microcompartment formation, which may explain transient transcriptional spiking observed upon mitotic exit.

biophysics↗

bioRxiv2024

LDB1 establishes multi-enhancer networks to regulate gene expression

How specific enhancer-promoter pairing is established is still mostly unclear. Besides the CTCF/cohesin machinery, only a few nuclear factors have been studied for a direct role in physically connecting regulatory elements. Here, we show via acute degradation experiments that LDB1 directly and broadly promotes enhancer-promoter loops. Most LDB1-mediated contacts, even those spanning hundreds of kb, can form in the absence of CTCF, cohesin, or YY1 as determined via the use of multiple degron systems. Moreover, an engineered LDB1-driven chromatin loop is cohesin independent. Cohesin-driven loop extrusion does not stall at LDB1 occupied sites but may aid the formation of a subset of LDB1 anchored loops. Leveraging the dynamic reorganization of nuclear architecture during the transition from mitosis to G1-phase, we establish a relationship between LDB1-dependent interactions in the context of TAD organization and gene activation. Lastly, Tri-C and Region Capture Micro-C reveal that LDB1 organizes multi-enhancer networks to activate transcription. This establishes LDB1 as a direct driver of regulatory network inter-connectivity.

molecular biology↗

bioRxiv2024

Extensive mutual influences of SMC complexes shape 3D genome folding

Mammalian genomes are folded by the distinct actions of SMC complexes which include the chromatin loop-extruding cohesin, the sister-chromatid cohesive cohesin, and the mitotic chromosome-associated condensins. While these complexes function at different stages of the cell cycle, they co-exist on chromatin during the G2/M-phase transition, when genome structure undergoes a dramatic reorganization. Yet, how distinct SMC complexes affect each other and how their mutual interplay orchestrates the dynamic folding of 3D genome remains elusive. Here, we engineered all possible cohesin/condensin configurations on mitotic chromosomes to delineate the concerted, mutual influential action of SMC complexes. We find that: (i) The mitotic SMC complex condensin disrupts the focal accumulation of extrusive-cohesin at CTCF binding sites, thereby promoting the disassembly of interphase TADs and chromatin loops during mitotic progression. Conversely, extrusive-cohesin can impair condensin activity and alter mitotic chromosome helicity. (ii) Condensin diminishes cohesive-cohesin focal enrichment and, conversely, cohesive-cohesin can counteract condensin function and impede mitotic chromosome longitudinal shortening. (iii) The co-presence of extrusive- and cohesive-cohesin synergistically antagonizes condensin function and dramatically delays mitotic chromosome condensation. (iv) Extrusive-cohesin positions cohesive-cohesin at CTCF binding sites. However, cohesive-cohesin by itself is insufficient to mediate the formation of TADs or chromatin loop, implying non-overlapping function with extrusive-cohesin. Instead, cohesive-cohesin restricts chromatin loop expansion, potentially by limiting extrusive-cohesin movement. Collectively, our data describe a comprehensive three-way interplay among major SMC complexes that dynamically sculpts chromatin architecture during cell cycle progression.

cell biology↗

bioRxiv2023

Genome folding principles revealed in condensin-depleted mitotic chromosomes

During mitosis, condensin activity interferes with interphase chromatin structures. Here, we generated condensin-free mitotic chromosomes to investigate genome folding principles. Co- depletion of condensin I and II, but neither alone, triggered mitotic chromosome compartmentalization in ways that differ from interphase. Two distinct euchromatic compartments, indistinguishable in interphase, rapidly emerged upon condensin loss with different interaction preferences and dependence on H3K27ac. Constitutive heterochromatin gradually self-aggregated and co-compartmentalized with the facultative heterochromatin, contrasting with their separation during interphase. While topologically associating domains (TADs) and CTCF/cohesin mediated structural loops remained undetectable, cis-regulatory element contacts became apparent, providing an explanation for their quick re-establishment during mitotic exit. HP1 proteins, which are thought to partition constitutive heterochromatin, were absent from mitotic chromosomes, suggesting, surprisingly, that constitutive heterochromatin can self-aggregate without HP1. Indeed, in cells traversing from M- to G1-phase in the combined absence of HP1, HP1{Pi} and HP1{gamma}, re-established constitutive heterochromatin compartments normally. In sum, "clean-slate" condensin-deficient mitotic chromosomes illuminate mechanisms of genome compartmentalization not revealed in interphase cells.

genomics↗

bioRxiv2022

Structure of the pre-mRNA leakage 39-kDa proteinreveals a single domain of integrated zf-C3HC and Rsm1 modules

In Saccharomyces cerevisiae, the pre-mRNA leakage 39-kDa protein (ScPml39) was reported to retain unspliced pre-mRNA prior to export through nuclear pore complexes (NPCs). Pml39 homologs outside the Saccharomycetaceae family are currently unknown, and mechanistic insight into Pml39 function is lacking. Here we determined the crystal structure of ScPml39 at 2.5 [A] resolution to facilitate the discovery of orthologs beyond Saccharomycetaceae, e.g. in Schizosaccharomyces pombe or human. The crystal structure revealed integrated zf-C3HC and Rsm1 modules, which are tightly associated through a hydrophobic interface to form a single domain. Both zf-C3HC and Rsm1 modules belong to the Zn-containing BIR (Baculovirus IAP repeat)-like super family, with key residues of the canonical BIR domain being conserved. Features unique to the Pml39 modules refer to the spacing between the Zn-coordinating residues, giving rise to a substantially tilted helix aC in the zf-C3HC and Rsm1 modules, and an extra helix AB in the Rsm1 module. Conservation of key residues responsible for its distinct features identifies S. pombe Rsm1 and Homo sapiens NIPA/ZC3HC1 as structural orthologs of ScPml39. Based on the recent functional characterization of NIPA/ZC3HC1 as a scaffold protein that stabilizes the nuclear basket of the NPC, our data suggest an analogous function of ScPml39 in S. cerevisiae.

biochemistry↗

bioRxiv2021

CTCF and transcription influence chromatin structure re-configuration after mitosis

During mitosis, transcription is globally attenuated and chromatin architecture is dramatically reconfigured. Here we exploited the M- to G1-phase progression to interrogate the contributions of the architectural factor CTCF and the process of transcription to re-sculpting the genome in newborn nuclei. Depletion of CTCF specifically during the M- to G1-phase transition altered the re-establishment of local short-range compartmentalization after mitosis. Chromatin domain boundary reformation was impaired upon CTCF loss, but a subset ([~]27%) of boundaries, characterized by transitions in chromatin states, was established normally. Without CTCF, structural loops failed to form, leading to illegitimate contacts between cis-regulatory elements (CREs). Transient CRE contacts that are normally resolved after telophase persisted deeply into G1-phase in CTCF depleted cells. CTCF loss-associated gains in transcription were often linked to increased, normally illegitimate enhancer-promoter contacts. In contrast, at genes whose expression declined upon CTCF loss, CTCF seems to function as a conventional transcription activator, independent of its architectural role. CTCF-anchored structural loops facilitated formation CRE loops nested within them, especially those involving weak CREs. Transcription inhibition did not elicit global architectural changes and left transcription start site-associated boundaries intact. However, ongoing transcription contributed considerably to the formation of gene domains, regions of enriched contacts spanning the length of gene bodies. Notably, gene domains formed rapidly in ana/telophase prior to the completion of the first round of transcription, suggesting that epigenetic features in gene bodies contribute to genome reconfiguration prior to transcription. The focus on the de novo formation of nuclear architecture during G1 entry yielded novel insights into how CTCF and transcription contribute to the dynamic re-configuration of chromatin architecture during the mitosis to G1 phase progression.

genomics↗

bioRxiv2019

Spatial genome re-organization between fetal and adult hematopoietic stem cells

Fetal hematopoietic stem cells (HSCs) undergo a developmental switch to become adult HSCs. The functional properties of the HSCs change dramatically during this switch, including their cycling behavior, hematopoietic lineage outputs and proliferation rate. The relationship between three-dimensional (3D) genome organization, epigenomic state, and transcriptome is poorly understood during this critical developmental transition. Here we conducted a comprehensive survey of the 3D genome, epigenome and transcriptome of fetal and adult HSCs in mouse. We found that chromosomal compartments and topologically associating domains (TAD) are largely conserved between fetal and adult HSCs. However, there is a global trend of increased compartmentalization and TAD boundary strength in adult HSCs. In contrast, dynamics of intra-TAD chromatin interactions is much higher and more widespread, involving over a thousand gene promoters and distal enhancers. Such dynamic interactions target genes involved in cell cycle, metabolism, and hematopoiesis. These developmental-stage-specific enhancer-promoter interactions appear to be mediated by different sets of transcription factors in fetal and adult HSCs, such as TCF3 and MAFB in fetal HSCs, versus NR4A1 and GATA3 in adult HSCs. Loss-of-function studies of TCF3 confirms the role of TCF3 in mediating condition-specific enhancer-promoter interactions and gene regulation in fetal HSCs. In summary, our data suggest that the fetal-to-adult transition is accompanied by extensive changes in intra-TAD chromatin interactions that target genes underlying the phenotypic differences between fetal and adult HSCs.

molecular biology↗

bioRxiv2019

Re-configuration of Chromatin Structure During the Mitosis-G1 Phase Transition

Higher-order chromatin organization such as A/B compartments, TADs and chromatin loops are temporarily disrupted during mitosis. These structures are thought to organize aspects of gene regulation, and thus it is important to understand how they are re-established after mitosis. We examined the dynamics of chromosome reorganization by Hi-C at defined time points following exit from mitosis in highly purified, synchronous cell populations. We observed that A/B compartments are rapidly established and progressively gain in strength following mitotic exit. Contact domain formation occurs from the \"bottom-up\" with smaller sub-TADs forming initially, followed by convergence into multi-domain TAD structures. CTCF is strongly retained at a significant fraction of sites on mitotic chromosomes and immediately resumes full binding at ana/telophase, the earliest tested time point. In contrast, cohesin is completely evicted from mitotic chromosomes and resumes focal binding with delayed kinetics. The formation of CTCF/cohesin co-anchored structural loops follows the kinetics of cohesin positioning. Stripe-shaped contacts anchored by CTCF grow in length, consistent with a loop extrusion process after mitosis. Interactions between cis-regulatory elements can form rapidly, preceding CTCF/cohesin anchored structural loops. Strikingly, we identified a group of rapidly emerging transient contacts between cis-regulatory elements in ana/telophase, that are dissolved upon G1 entry, co-incident with the establishment of inner boundaries or nearby interfering loops. Our findings indicate that distinct but mutually influential forces drive post-mitotic chromatin re-configuration to shape compartments, contact domains, cis-element contacts, and CTCF/cohesin dependent loops.

genomics↗

bioRxiv2017

A dementia-associated risk variant near TMEM106B alters chromatin architecture and gene expression

Neurodegenerative diseases pose an extraordinary threat to the worlds aging population, yet no disease-modifying therapies are available. While genome-wide association studies (GWAS) have identified hundreds of novel risk loci for neurodegeneration, the mechanisms by which these loci influence disease risk are largely unknown. Indeed, of the many thousands of SNP-trait associations identified by GWAS over the past ~10 years, very few are understood mechanistically. Here, we investigate the association of common genetic variants at the 7p21 locus with risk for the neurodegenerative disease frontotemporal lobar degeneration. We show that variants associated with disease risk correlate with increased brain expression of the 7p21 gene TMEM106B, and no other genes. Furthermore, incremental increases in TMEM106B levels result in incremental increases in lysosomal phenotypes and cell toxicity. We then combine fine-mapping, bioinformatics, and bench-based approaches to functionally characterize all candidate causal variants at this locus. This approach identified a noncoding variant, rs1990620, which differentially recruits CTCF, influencing CTCF-mediated long-range chromatin looping interactions between multiple cis-regulatory elements, including the TMEM106B promoter. Our findings thus provide an in-depth analysis of the 7p21 locus linked by GWAS to frontotemporal lobar degeneration, nominating a causal variant and a causal mechanism for allele-specific expression and disease association at this locus. Finally, we show that genetic variants associated with risk for neurodegenerative diseases beyond frontotemporal lobar degeneration are enriched in brain CTCF-binding sites genome-wide, implicating CTCF-mediated gene regulation in risk for neurodegeneration more generally.

genetics↗