Biology subjects

Gregor, T.

Publications and source records attributed to Gregor, T..

13 recordsLinked to original sources

bioRxiv2026

Structural survey of HIF-2α reveals regulation of its subcellular localization and protein interactome

Hypoxia-inducible factor 2 (HIF-2) is a central regulator of cellular homeostasis and a known oncogenic driver in multiple cancers. Although HIF-2 is canonically defined as a nuclear transcription factor, its cytoplasmic presence and non-canonical functions remain poorly understood. Here, we performed a structural survey of HIF-2 to determine the mechanisms underlying subcellular localization, protein abundance, and activity using a deletion-construct library, transcriptional assays, and in vivo xenograft models. We found that the oxygen-dependent degradation domain (ODD), the N-terminal intrinsically disordered region (n-IDR) and the N-terminal transactivation domain (NTAD) promote cytoplasmic localization, whereas the C-terminal IDR drives nuclear accumulation. Surprisingly, we found that HIF-2 nuclear localization occurs also in the absence of PAS A and B, the domains required for ARNT (HIF-1{beta}) dimerization, resolving the long-standing question in the field. These data suggest a dominant role for non-canonical cytoplasmic mechanisms in HIF-2-driven tumorigenesis. Strikingly, neither NTAD nor the C-terminal CTAD was required for tumor growth in vivo, in coherence with our transcriptional assays indicating that CTAD is dispensable for transactivation and NTAD functions as a suppressor rather than an activator of transcription. Proteomic analyses reveal HIF-2 interactions with regulators of mitochondrial function, translation initiation, RNA splicing, vesicular transport, and DNA replication. Together, these findings uncover previously unrecognized structural and functional complexity of HIF-2 compartmentalization and expand its role beyond canonical transcriptional regulation.

cell biology↗

bioRxiv2026

Spatially correlated fluctuations govern relative chromatin motion

Essential nuclear processes require pairs of chromosomal loci to find each other in three-dimensional space. Polymer models of chromosome dynamics typically assume that the stochastic forces driving such locus motion are spatially uncorrelated, implying that relative diffusion follows directly from single-locus dynamics. Here we show that this assumption fails in living cells. Using live-cell imaging in fly embryos and mouse embryonic stem cells, we find that pairwise locus distances diffuse markedly slower than predicted for independent fluctuations. Combining stochastic trajectory analysis with polymer simulations, we demonstrate that this slowdown arises from non-equilibrium spatially correlated fluctuations (SCFs) in the nucleoplasm, which cause nearby loci to move coherently. We establish three experimentally testable signatures of SCFs: fluctuation amplitudes plateau at large distances, are independent of genomic separation, and show an anomalous temporal scaling. All three predictions are confirmed experimentally, including for loci on separate chromosomes. ATP depletion and disruption of cohesin-mediated loop extrusion reveal that both active processes and crosslinking contribute to correlation magnitudes. Because SCFs slow relative motion preferentially at short distances, they reduce encounter frequencies while prolonging encounter durations, generating a trade-off with direct implications for gene regulation. Our results identify spatially correlated fluctuations as a fundamental determinant of relative motion in confined active polymers.

biophysics↗

bioRxiv2026

Loop Extrusion Accelerates Long-Range Enhancer-Promoter Searches in Living Embryos

Long-range gene regulation underlies a variety of human developmental disorders including Cornelia de Lange syndrome1 and polydactyly2. Cohesin-mediated loop extrusion and tether-like elements are two major mechanisms implicated in fostering long-range enhancer-promoter (E-P) contacts3. However, our understanding of the contributions of these mechanisms to the kinetics of E-P interactions is limited. Here we employ a combination of quantitative single-cell imaging, genetic manipulations and polymer simulations to examine the interplay of cohesin-mediated loop extrusion and tethering elements4,5 in the timely activation of gene expression in living Drosophila embryos. Depletion of NIPBL or deletion of an enhancer-proximal CTCF anchor element reduced expression without changing the duration of individual transcriptional bursts. Genetic epistasis experiments recapitulated polymer simulations predicting complementation of tether deletions by augmenting the stability of cohesin via reduced levels of WAPL6. We propose a "scan and snag" model whereby directional cohesin-driven enhancer scanning promotes diffusion-mediated tethering interactions to produce successful E-P contacts and transcriptional activation. We discuss how modulating cohesin stability and the "stickiness" of looping factors7-10 can fine-tune the levels and timing of gene expression in mammalian developmental and disease processes.

molecular biology↗

bioRxiv2025

Fine-tuning mechanical constraints uncouple patterning and gene expression in murine pseudo-embryos

The interplay between mechanical forces and genetic programs is fundamental to embryonic development, yet how these factors independently or jointly influence morphogenesis and cell fate decisions remains poorly understood. Here, we fine-tune the mechanical environment of murine gastruloids, three-dimensional in vitro models of early embryogenesis, by embedding them in bioinert hydrogels with precisely tunable stiffness and timing of application. This approach reveals that external constraints can selectively influence transcriptional profiles, patterning, or morphology, depending on the level and timing of mechanical modulation. Gastruloids embedded in ultra-soft hydrogels (< 30 Pa) elongate robustly, preserving both anteroposterior patterning and transcriptional profiles. In contrast, embedding at higher stiffness disrupts polarization while leaving gene expression largely unaffected. Conversely, earlier embedding significantly impacts transcriptional profiles independently of polarization defects, highlighting the uncoupling of patterning and transcription. These findings suggest that distinct cellular states respond differently to external constraints. Live imaging and cell tracking imply that impaired cell motility underlies polarization defects, underscoring the role of mechanical forces in shaping morphogenesis independently of transcriptional changes. By allowing precise control over external mechanical boundaries, our approach provides a powerful platform to dissect how physical and biochemical factors interact to orchestrate early embryonic development.

developmental biology↗

bioRxiv2024

Size-dependent temporal decoupling of morphogenesis and transcriptional programs in gastruloids

Understanding the interplay between cell fate specification and morphogenetic changes remains a central challenge in developmental biology. Gastruloids, self-organizing stem cell-based models of post-implantation mammalian development, provide a powerful platform to address this question. Here, we show that physical parameters, particularly system size, critically influence the timing and outcomes of morphogenetic processes. Larger gastruloids exhibit delayed symmetry breaking, increased multipolarity, and prolonged axial elongation, with morphogenesis driven by system size. Despite these variations, transcriptional programs and cell fate composition remain remarkably stable across a broad size range. Notably, extreme sizes show distinct transcriptional modules and clear shifts in gene expression patterns. Intriguingly, size perturbation experiments rescued the morphogenetic and pattern phenotypes observed in extreme sizes, demonstrating the remarkable adaptability of gastruloids to their effective system size. These findings establish gastruloids as versatile models for studying spatiotemporal dynamics in mammalian embryogenesis and reveal how physical constraints decouple transcriptional from morphogenetic programs.

developmental biology↗

bioRxiv2023

Stochastic motion and transcriptional dynamicsof distal enhancer-promoter pairs on a compacted chromosome

Chromosomes in the eukaryotic nucleus are highly compacted. However, for many functional processes, including transcription initiation, the 3D pair-wise motion of distal chromosomal elements, such as enhancers and promoters, is essential and necessitates dynamic fluidity. Therefore, the interplay of chromosome organization and dynamics is crucial for gene regulation. Here, we use a live imaging assay to simultaneously measure the positions of pairs of enhancers and promoters and their transcriptional output in the developing fly embryo while systematically varying the genomic separation between these two DNA loci. Our analysis reveals a combination of a compact globular organization and fast subdiffusive dynamics. These combined features cause an anomalous scaling of polymer relaxation times with genomic separation and lead to long-ranged correlations compared to existing polymer models. This scaling implies that encounter times of DNA loci are much less dependent on genomic separation than predicted by existing polymer models, with potentially significant consequences for eukaryotic gene expression.

biophysics↗

bioRxiv2023

MOXD1 is a gate-keeper of organ homeostasis and functions as a tumor-suppressor in neuroblastoma

Neuroblastoma is a childhood cancer believed to result from dysfunctional development. Its origin during embryogenesis remains poorly understood. The lack of appropriate models has hindered in-depth mapping of tumor-driving events. Here, we identify a novel tumor-suppressor gene that predicts poor survival in high-risk disease, by applying bulk and single cell RNA sequencing data of neuroblastoma and human fetal adrenal glands. Trunk neural crest-specific MOXD1 discriminates cell populations during normal and tumor development, with implications for deciphering neuroblastoma cell origin. We created an embryonic conditional knockout model and show that cell type-specific loss of MOXD1 leads to disrupted organ homeostasis and failed adrenal gland formation, home for neuroblastoma. We show that MOXD1 is a tumor suppressor gene in zebrafish, chick, and mice in vivo models. One-Sentence SummaryNeural crest-specific MOXD1 is a de novo tumor-suppressor gene in childhood cancers arising during embryogenesis.

cancer biology↗

bioRxiv2022

Plastid DNA sequences and oospore characters of some European species of Tolypella section Tolypella (Obtusifolia, Characeae) indicate a new cryptic Tolypella species from the Mediterranean island Sardinia

In Europe, the genus Tolypella (Characeae) comprises four to eight Tolypella species in sections Rothia and Tolypella that have been distinguished by vegetative morphology and gametangial characters such as antheridial size and oospore cell wall ornamentation. However, morphological species differentiation is difficult in some cases due to overlapping and variable vegetative features, which in many cases are difficult to observe clearly. To clarify the taxonomic status of the five European species of Tolypella in section Tolypella, sequence data of the plastid genes atpB, rbcL and psbC for Tolypella glomerata (Desv.) Leonh., Tolypella hispanica Allen, Tolypella nidifica (O.F. Mull.) A. Braun, Tolypella normaniana (Nordst.) Nordst. and Tolypella salina Cor. were combined with data on oospore morphology, including oospore wall ornamentation. Gene sequence data identified five distinct clusters, but they differed from the morphologically identified five species. T. glomerata consisted of some of the samples morphologically identified as T. glomerata and seven samples of T. normaniana, while the remaining T. glomerata samples clustered with specimens of unclear affiliation ("Tolypella. sp."). T. hispanica I consisted of samples from various locations, whereas "T. hispanica II" consisted of samples of T. hispanica from the Mediterranean island, Sardinia. The remaining cluster consisted of all the specimens that had been determined as T. salina or T. nidifica in addition to two specimens of T. normaniana. Oospore morphology was most clearly distinguishable for T. glomerata. Oospore characteristics for all other taxa were not as informative but showed some geographical and/or environmentally influenced differences, especially for T. nidifica and T. salina. Our results suggest a significantly different taxonomy of Tolypella sect. Tolypella in which specimens normally identified as T. glomerata might be two different species, T. glomerata and an unidentified species; T. nidifica and T. salina are not separate species; T. normaniana is a diminutive variant of T. nidifica or T. salina; and T. hispanica comprises two different species, one from the Mediterranean island Sardnia.

plant biology↗

bioRxiv2021

Optogenetic control of the Bicoid morphogen reveals fast and slow modes of gap gene regulation

Developmental patterning networks are regulated by multiple inputs and feedback connections that rapidly reshape gene expression, limiting the information that can be gained solely from slow genetic perturbations. Here we show that fast optogenetic stimuli, real-time transcriptional reporters, and a simplified genetic background can be combined to reveal quantitative regulatory dynamics from a complex genetic network in vivo. We engineer light-controlled variants of the Bicoid transcription factor and study their effects on downstream gap genes in embryos. Our results recapitulate known relationships, including rapid Bicoid-dependent expression of giant and hunchback and delayed repression of Kruppel. In contrast, we find that the posterior pattern of knirps exhibits a quick but inverted response to Bicoid perturbation, suggesting a previously unreported role for Bicoid in suppressing knirps expression. Acute modulation of transcription factor concentration while simultaneously recording output gene activity represents a powerful approach for studying how gene circuit elements are coupled to cell identification and complex body pattern formation in vivo.

developmental biology↗

bioRxiv2021

Temporally dynamic antagonism between transcription and chromatin compaction controls stochastic photoreceptor specification

Stochastic mechanisms diversify cell fates during development. How cells randomly choose between two or more fates remains poorly understood. In the Drosophila eye, the random mosaic of two R7 photoreceptor subtypes is determined by expression of the transcription factor Spineless (Ss). Here, we investigated how cis-regulatory elements and trans factors regulate nascent transcriptional activity and chromatin compaction at the ss gene locus during R7 development. We find that the ss locus is in a compact state in undifferentiated cells. An early enhancer drives ss transcription in all R7 precursors to open the ss locus. In differentiating cells, transcription ceases and the ss locus stochastically remains open or compacts. In SsON R7s, ss is open and competent for activation by a late enhancer, whereas in SsOFF R7s, ss is compact and repression prevents expression. Our results suggest that a temporally dynamic antagonism, in which transcription drives decompaction and then compaction represses transcription, controls stochastic cell fate specification.

developmental biology↗

bioRxiv2019

Elucidation of protein-protein interactions necessary for maintenance of the BCR-ABL signaling complex

Approximately 50% of chronic myeloid leukemia (CML) patients in deep remission experience a return of clinical CML after withdrawal of tyrosine kinase inhibitors (TKIs). This suggests signaling of inactive BCR-ABL, which allows for survival of cancer cells, leading to relapse. Understanding the dynamics of BCR-ABL signaling complex holds a key to the mechanism of BCR-ABL signaling. Here, we demonstrate that TKIs inhibit catalytic activity of BCR-ABL, but do not dissolve the BCR-ABL core signaling complex consisting of CrkL, SHC1, Grb2, SOS1, cCbl, and SHIP2. We show that CrkL binds to proline-rich regions located in C-terminal, intrinsically disordered region of BCR-ABL, that deletion of pleckstrin homology domain of BCR-ABL diminishes interaction with SHC1, and that BCR-ABL sequence motif located in disordered region around phosphorylated tyrosine 177 mediates binding of at least three core complex members, the Grb2, SOS1 and cCbl. Introduction of Y177F substitution blocks association with Grb2, SOS1 and cCbl. Further, we identified SHIP2 binding sites within the src-homology and tyrosine kinase domains of BCR-ABL. We found that BCR-ABL is unable to phosphorylate SHC1 in cells lacking SHIP2. Reintroducing SHIP2 into Ship2 knock-out cells restored SHC1 phosphorylation, which depended on inositol phosphatase activity of SHIP2. Our findings provide characterization of protein-protein interactions in the BCR-ABL signaling complex, and support the concept of targeting BCR-ABL signaling in CML by inhibition of its interactions with the members of the core complex.

molecular biology↗

bioRxiv2018

Eco-evolutionary significance of ‘loners’

Loners, individuals out-of-sync with a coordinated majority, occur frequently in nature. Are loners incidental byproducts of large-scale synchronization attempts or are they part of a mosaic of life-history strategies? Here, we provide the first empirical evidence of naturally occurring heritable variation in loner behavior, using the social amoeba Dictyostelium discoideum. Moreover, we show that Dictyostelium loners--cells that do not join the multicellular life-stage-- result from a dynamic population-partitioning process. Underlying this partitioning, we find evidence that each cell makes a stochastic, signal-based decision resulting in an imperfectly synchronized multicellular development affected by both abiotic (environmental porosity) and biotic (strain-specific signaling) factors. Finally, we predict that when strains differing in their partitioning behavior co-occur, cross-signaling impacts slime-mold diversity across spatio-temporal scales. Loners are therefore critical to understanding collective and social behaviors, multicellular development, and ecological dynamics in D. discoideum. More broadly, across taxa, imperfect synchronization might be adaptive by enabling diversification of life-history strategies.

ecology↗

bioRxiv2017

Direct visualization of transcriptional activation by physical enhancer-promoter proximity

A long-standing question in metazoan gene regulation is how remote enhancers communicate with their target promoters over long distances. Combining genome editing and quantitative live imaging we simultaneously visualize physical enhancer-promoter communication and transcription in Drosophila embryos. Enhancers regulating pair rule stripes of even-skipped expression activate transcription of a reporter gene over a distance of 150 kb. We show in individual cells that activation only occurs after the enhancer comes into close proximity with its regulatory target and that upon dissociation transcription ceases almost immediately. We further observe distinct topological conformations of the eve locus, depending on the spatial identity of the activating stripe enhancer. In addition, long-range activation results in transcriptional competition at the endogenous eve locus, causing corresponding developmental defects. Overall, we demonstrate that sustained physical proximity and enhancer-promoter engagement are required for enhancer action, and we provide a path to probe the implications of long-range regulation on cellular fates.

biophysics↗