Biology subjects

Dean, K. M.

Publications and source records attributed to Dean, K. M..

At least 19 recordsLinked to original sources

bioRxiv2026

Niche-dependent epistasis within the cancer genome shapes metastatic fitness

Epistasis, the genetic interaction between loci, directs non-additive genotype-to-phenotype outcomes. Within a multicellular organism, how the tissue niche (E) shapes gene-by-gene epistasis (GxG) and its phenotypic consequences remains poorly studied. Using tumor lung metastasis as an experimental model, we mapped GxGxE through combinatorial CRISPR perturbation coupled with depletion of defined host myeloid populations in mice. The metastatic niche first selects disseminated tumor cells on GxG epistatic fitness, then drives transcriptome convergence regardless of genotype. Removing anatomically distinct myeloid populations reshaped the metastatic epistasis landscape despite identical tumor genotypes. Gene-by-gene-by-environment epistasis therefore operates at the organismal level, with interaction magnitude and sign set by niche cells. Beyond metastasis, our data suggest that the tissue niche conditions how genetic interactions translate into divergent phenotypes.

genetics↗

bioRxiv2026

Deciphering the Multiscale Morphology of Somatic Oncogenic Alterations in Hepatocellular Carcinoma

Somatic oncogenic mutations are typically defined by their molecular alterations, yet how they reorganize cellular architecture within intact tissues remain largely unknown. Here, we demonstrate that distinct oncogenic drivers produce unique multiscale architectural phenotypes that can be quantitatively resolved in intact liver tissue. Using iterative expansion microscopy, multiscale light-sheet imaging, and three-dimensional morphometric analysis, we systematically mapped structural remodeling from single-cell morphology to mitochondrial architecture in mosaic mouse models of hepatocellular oncogene activation. NRAS and CTNNB1 induced fundamentally different morphological programs. NRAS activation drove extensive remodeling of cell morphology, membrane curvature, and surface irregularity, whereas CTNNB1 activation largely preserved global cell morphology while selectively altering mitochondrial organization and shape. We first established a zonation-aware reference state for interpreting oncogene-associated organelle remodeling by resolving mitochondrial differences between periportal and pericentral hepatocytes. Within this framework, CTNNB1 activation shifted mitochondrial features toward a pericentral-like state, consistent with the role of Wnt/{beta}-catenin signaling in hepatic zonation and metabolic identity. Furthermore, integrating cellular morphology, membrane geometry, and mitochondrial architecture improved discrimination of oncogenic states beyond any individual structural feature, demonstrating that mutation-specific phenotypes arise through coordinated remodeling across multiple biological scales. Together, these findings establish multiscale structural phenotyping as a framework for linking oncogenic genotype to three-dimensional cellular organization and reveal that distinct oncogenic drivers remodel different architectural compartments during liver oncogene activation.

cancer biology↗

bioRxiv2026

Transient architecture of the embryonic pancreas determines endocrine mass

During organogenesis, epithelial tissues undergo extensive three-dimensional (3D) remodeling while simultaneously generating specialized cell types. Whether these transient architectural states actively instruct lineage allocation remains unclear. Here we identify a morphogenetic stage in which resolution of epithelial stratification is required for lineage allocation and establishment of endocrine cell mass. We show that loss of the Hippo pathway regulator Merlin disrupts lumen morphogenesis and prevents formation of the transient 3D epithelial architecture that characterizes normal pancreas development. Failure to establish this architectural state alters lineage allocation, impairing acinar differentiation, markedly reducing adult endocrine cell mass, and disrupting glucose homeostasis. Mosaic analyses reveal that these lineage defects arise non-cell autonomously, demonstrating that epithelial architecture itself instructs cell fate decisions. Mechanistically, Merlin coordinates PI3K-regulated polarized membrane trafficking required for apical membrane biogenesis and lumen formation. Together, these findings identify Merlin-dependent membrane trafficking as a mechanism coupling epithelial morphogenesis to lineage allocation and demonstrate that transient developmental architectures can determine the cellular composition of mature organs.

developmental biology↗

bioRxiv2026

Altair-dvOPM: an open-access platform for large-field three-dimensional tissue imaging

Altair-dvOPM is an open-access direct-view oblique plane microscope designed for large-field, three-dimensional imaging of cleared and expanded tissue sections. By combining photographic-lens-based detection, externally launched oblique illumination and precision-registered modular baseplates, the system achieves micrometer-scale lateral resolution over a ~5.4 mm field of view without custom objectives or highly specialized alignment procedures. We demonstrate imaging across scales, from subcellular structures in expanded cells to centimeter-scale expanded tissue sections, and provide documentation, CAD files, Zemax models and open-source control software to support replication and extension.

biophysics↗

bioRxiv2026

Modulation of Oncogenic KRAS Signaling by Branched Actin-driven Cell Membrane Protrusions

For over three decades, we have known that oncogenic RAS alters the actin cytoskeleton organization and cell surface morphology1,2. RAS activates the GTPase RAC1, which triggers the growth of branched actin networks to promote cell membrane protrusions3,4. In melanoma, the hyperactive RAC1 mutant, Rac1P29S, was recently shown to drive extended lamellipodia, which then empower cell proliferation through sequestration and localized inhibition of the merlin tumor suppressor5. This discovery illustrates cell morphological programs not only as outputs but also as regulators of human oncogenic signals. Hence, we wondered whether the pronounced branched actin-driven membrane protrusions (BAMPs) downstream of oncogenic RAS are not mere outputs of RAS signaling but rather an active component in mediating the oncogenic penetrance of RAS mutants. We used volumetric light sheet microscopy and biochemical approaches to investigate the role of BAMPs in regulating the molecular signaling of oncogenic KRAS in pancreatic and lung cancer models. We found that elevated BAMP formation regulated the interaction of oncogenic KRAS with downstream effectors, specifically with the RAC1 GEF TIAM1. This implies that BAMPs amplify their own upstream regulators in a positive feedback. This meritorious cycle upregulates cyclin D1 expression by inactivating the merlin tumor suppressor, independently of the mitogen activated protein kinase pathway (MAPK). In the absence of BAMPs, cells carrying oncogenic KRAS mutations are unable to attain their full penetrance in proliferation. Overall, this work unveils the long-overlooked role of branched actin-driven cell morphology in the functionalization of KRAS mutants as potent oncogenes.

cancer biology↗

bioRxiv2026

Leptin Receptor+ cells create a perisinusoidal niche for thrombopoiesis in the bone marrow by synthesizing CXCL14

Leptin Receptor-expressing (LepR+) stromal cells in the bone marrow are a critical source of growth factors for the maintenance of hematopoietic stem cells (HSCs) and most restricted hematopoietic progenitors. An important unresolved question is whether they also regulate terminal differentiation in some hematopoietic cells. We found that LepR+ cells promote thrombopoiesis by synthesizing the chemokine CXCL14, which is expressed in the bone marrow by a subset of LepR+ cells. Cxcl14-expressing LepR+ cells extend fine processes that wrap around perisinusoidal megakaryocytes. Deletion of Cxcl14 from LepR+ cells did not significantly alter HSC function or most aspects of bone marrow hematopoiesis, including megakaryocyte generation; however, it significantly reduced the numbers of proplatelet-forming megakaryocytes in the bone marrow and platelets in the blood. CXCL14 promoted platelet formation by remodeling lipid metabolism in megakaryocytes, increasing fatty acid transporter expression and enabling megakaryocytes to take up more polyunsaturated fatty acids from circulation. A high fat diet rescued the formation of proplatelet-forming megakaryocytes and platelets in Lepr-cre; Cxcl14 fl/fl mice. CXCL14 protein was sufficient to promote platelet formation by megakaryocytes in vitro and in vivo. LepR+ cells thus create a perisinusoidal niche for thrombopoiesis by producing CXCL14, which regulates lipid metabolism and terminal differentiation in megakaryocytes. Key pointsO_LILeptin Receptor+ stromal cells regulate terminal differentiation in megakaryocytes in addition to maintaining stem and progenitor cells C_LIO_LICXCL14 from Leptin Receptor+ cells promotes the formation of platelets by remodeling lipid metabolism in megakaryocytes in the bone marrow C_LI

cell biology↗

bioRxiv2026

In vivo evidence for bleb-induced survival signaling in metastatic melanoma

Bleb signaling is a cellular process in which pressure-driven plasma membrane protrusions generate localized micron-scale membrane curvature that recruits cytosolic septin complexes, assembling signaling hubs that promote cell survival1. Our prior work showed that this morphology-encoded signaling pathway is necessary to sustain anchorage-independent survival of BRAF and NRAS mutant melanoma cells in vitro. However, whether bleb-induced signaling occurs in vivo and contributes to cancer progression is unclear. Here, we develop complementary in vivo and ex vivo assays spanning human patient samples and mouse xenograft models to mechanistically interrogate bleb signaling directly in physiologic contexts. We observe that bleb-associated septin hubs are exclusively formed in poorly adherent amoeboid tumor cells at the invasive margin, within malignant effusions, and at distant metastatic sites, while well-adhered cells in the tumor interior show no signs of septin hub formation. Accordingly, pharmacological pathway disruption specifically kills disseminated tumor cells within these low-adhesion microenvironments while having no appreciable effect on adhered cells in the tumor interior, resulting in reduced metastatic burden and delayed disease recurrence in vivo. This work confirms septin-mediated bleb signaling as a previously unrecognized vulnerability of disseminated cancer cells and demonstrates that this morphology-encoded pathway operates in vivo to support disease progression, preserving cancer cell viability under conditions in which cell survival signals from the environment are muted. These findings suggest novel opportunities to target survival signaling in micrometastatic disseminated cancer cells. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/697601v1_ufig1.gif" ALT="Figure 1"> View larger version (61K): org.highwire.dtl.DTLVardef@1770384org.highwire.dtl.DTLVardef@87e567org.highwire.dtl.DTLVardef@1e5d6b2org.highwire.dtl.DTLVardef@724c5f_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2026

Unraveling Subcellular Ultrastructure with Cyclically Multiplexed Expansion Microscopy

Despite advances in fluorescence microscopy, spectral overlap and limited resolution hinder the dense mapping of the cellular ultrastructure. To overcome these challenges, we developed Cy-ExM, a high-plex imaging strategy that integrates optimized cryo-fixation for antigen preservation, expansion microscopy, and iterative immunofluorescence labeling. Using oblique plane microscopy, we perform three-dimensional super-resolution imaging of 20 biological targets encompassing the full cellular volume of individual mammalian cells.

cell biology↗

bioRxiv2025

Multi-immersion Oblique Plane Microscope (miOPM): A reconfigurable platform for high-resolution Light-Sheet Fluorescence Microscopy.

Light-Sheet Fluorescence Microscopy (LSFM) enables gentle, rapid, and efficient volumetric imaging of biological specimens. Despite its potential, LSFM has fragmented into multiple variants, each narrowly optimized for specific samples or imaging regimes, limiting its broad applicability. To overcome this barrier, we introduce the multi-immersion oblique plane microscope (miOPM), an adaptable LSFM platform for high-resolution imaging ranging from subcellular dynamics to whole organisms and cleared tissues. miOPM uniquely supports seamless interchangeability of oil, water, and air objectives, and maintains diffraction limited performance across a refractive index range of 1.33 - 1.51. It is compatible with standard sample mounting, current clearing protocols and high-throughput 3D imaging. We leverage miOPM to image diverse biological specimens at multiple spatial scales. By greatly expanding the adaptability and ease of use of LSFM, miOPM is poised to democratize advanced three-dimensional imaging.

bioengineering↗

bioRxiv2025

Altair-LSFM: A High-Resolution, Easy-to-Build Light-Sheet Microscope for Sub-Cellular Imaging

Although several open-source, easy-to-assemble light-sheet microscope platforms already exist--such as mesoSPIM, OpenSPIM, and OpenSpin--they are optimized for imaging large specimens and lack the resolution required to visualize sub-cellular features, such as organelles or cytoskeletal architectures. In contrast, Latice Light-Sheet Microscopy (LLSM) achieves the resolution necessary to resolve such fine structures but, in its open-source implementation, can be alignment- and maintenance-intensive, often requiring specialist expertise. To address this gap, we developed Altair-LSFM, a high-resolution, open-source, sample-scanning light-sheet microscope specifically designed for sub-cellular imaging. By optimizing the optical pathway in silico, we created a custom baseplate that greatly simplifies alignment and assembly. The system integrates streamlined optoelectronics and optomechanics with seamless operation through our open-source software, navigate. Altair-LSFM achieves lateral and axial resolutions of approximately 235 nm and 350 nm, respectively, across a 266-micron field of view after deconvolution. We validate the systems capabilities by imaging sub-diffraction fluorescent nanospheres and visualizing fine structural details in mammalian cells, including microtubules, actin filaments, nuclei, and Golgi apparatus. We further demonstrate its live-cell imaging capabilities by visualizing microtubules and vimentin intermediate filaments in actively migrating cells.

biophysics↗

bioRxiv2025

Feature-Driven Whole-Tissue Imaging with Subcellular Resolution

Understanding how subcellular structures and events shape tissue-wide phenomena remains a central challenge in biology. We present Multiscale Cleared Tissue Axially Swept Light-Sheet Microscopy (MCT-ASLM), a platform combining cm-scale imaging with targeted high-resolution interrogation of intact tissues in human-guided or autonomous modes. Capable of capturing fields of view up to 21 mm at micron-scale resolution, MCT-ASLM can seamlessly transition to submicron (~300 nm) isotropic resolution for targeted imaging. This versatility enables comprehensive studies of hierarchical organization and spatially complex processes, including mapping neuronal circuits in rat brains, visualizing glomerular innervation in mouse kidneys, and examining metastatic tumor microenvironments. By bridging subcellular to tissue-level scales, MCT-ASLM offers a powerful method for unraveling how local events contribute to global biological phenomena.

biophysics↗

bioRxiv2024

A general algorithm for consensus 3D cell segmentation from 2D segmented stacks

Cell segmentation is the foundation of a wide range of microscopy-based biological studies. Deep learning has revolutionized 2D cell segmentation, enabling generalized solutions across cell types and imaging modalities. This has been driven by the ease of scaling up image acquisition, annotation, and computation. However, 3D cell segmentation, requiring dense annotation of 2D slices still poses significant challenges. Manual labeling of 3D cells to train broadly applicable segmentation models is prohibitive. Even in high- contrast images annotation is ambiguous and time-consuming. Here we develop a theory and toolbox, u- Segment3D, for 2D-to-3D segmentation, compatible with any 2D method generating pixel-based instance cell masks. u-Segment3D translates and enhances 2D instance segmentations to a 3D consensus instance segmentation without training data, as demonstrated on 11 real-life datasets, >70,000 cells, spanning single cells, cell aggregates, and tissue. Moreover, u-Segment3D is competitive with native 3D segmentation, even exceeding when cells are crowded and have complex morphologies.

bioinformatics↗

bioRxiv2024

Mechanically Sheared Axially Swept Light-Sheet Microscopy

We present a mechanically sheared image acquisition format for upright and open-top light-sheet microscopes that automatically places data in its proper spatial context. This approach, which reduces computational post-processing and eliminates unnecessary interpolation or duplication of the data, is demonstrated on an upright variant of Axially Swept Light-Sheet Microscopy (ASLM) that achieves a field of view, measuring 774 x 435 microns, that is 3.2-fold larger than previous models and a raw and isotropic resolution of [~]420 nm. Combined, we demonstrate the power of this approach by imaging sub-diffraction beads, cleared biological tissues, and expanded specimens.

biophysics↗

bioRxiv2024

navigate: an open-source platform for smart light-sheet microscopy.

navigate is a turnkey, open-source software solution designed to enhance light-sheet fluorescence microscopy (LSFM) by integrating smart microscopy techniques into a user-friendly framework. It enables automated, intelligent imaging with a Python-based control system that supports GUI-reconfigurable acquisition routines and the integration of diverse hardware sets. As a comprehensive package, navigate democratizes access to advanced LSFM capabilities, facilitating the development and implementation of smart microscopy workflows without requiring deep programming knowledge or specialized expertise in light-sheet microscopy.

biophysics↗

bioRxiv2023

Keratin isoform shifts modulate motility signals during wound healing

Keratin intermediate filaments form strong mechanical scaffolds that confer structural stability to epithelial tissues, but the reason this function requires a protein family with 54 isoforms is not understood. During skin wound healing, a shift in keratin isoform expression alters the composition of keratin filaments. How this change modulates cellular function to support epidermal remodeling remains unclear. We report an unexpected effect of keratin isoform variation on kinase signal transduction. Increased expression of wound-associated keratin 6A, but not of steady-state keratin 5, potentiated keratinocyte migration and wound closure without compromising epidermal stability by activating myosin motors. This pathway depended on isoform-specific interaction between intrinsically disordered keratin head domains and non-filamentous vimentin shuttling myosin-activating kinases. These results substantially expand the functional repertoire of intermediate filaments from their canonical role as mechanical scaffolds to include roles as isoform-tuned signaling scaffolds that organize signal transduction cascades in space and time to influence epithelial cell state.

cell biology↗

bioRxiv2022

Increasing the Field-of-View in Oblique Plane Microscopy via optical tiling

Fast volumetric imaging of large fluorescent samples with high-resolution is required for many biological applications. Oblique plane microscopy (OPM) provides high spatiotemporal resolution, but the field of view is typically limited by its optical train and the pixel number of the camera. Mechanically scanning the sample or decreasing the overall magnification of the imaging system can partially address this challenge, albeit by reducing the volumetric imaging speed or spatial sampling, respectively. In this Letter, we introduce a novel dual-axis scan unit for OPM that enables rapid and high-resolution volumetric imaging throughout a volume of 800 x 500 x 200 microns. This enables imaging of model organisms, such as zebrafish embryos, with subcellular resolution. Furthermore, we combined this microscope with a real-time and multi-perspective projection imaging technique to increase the volumetric interrogation rate to more than 10 Hz.

bioengineering↗

bioRxiv2022

Resolution doubling in light-sheet microscopy via oblique plane structured illumination

Structured illumination microscopy (SIM) doubles the spatial resolution of a fluorescence microscope without requiring high laser powers or specialized fluorophores. However, the excitation of out-of-focus fluorescence can accelerate photobleaching and phototoxicity. In contrast, light-sheet fluorescence microscopy (LSFM) largely avoids exciting out-of-focus fluorescence, thereby enabling volumetric imaging with low photo-bleaching and intrinsic optical sectioning. Combining SIM with LSFM would enable gentle 3D imaging at doubled resolution. However, multiple orientations of the illumination pattern, which are needed for isotropic resolution doubling in SIM, are challenging to implement in a light-sheet format. Here we show that multidirectional structured illumination can be implemented in oblique plane microscopy, a LSFM technique that uses a single objective for excitation and detection, in a straightforward manner. We demonstrate isotropic lateral resolution below 150nm, combined with lower photo-toxicity compared to traditional SIM systems and volumetric acquisition speed exceeding 1Hz.

biophysics↗

bioRxiv2022

A Microtubule Mechanostat Enables Cells to Navigate Confined Environments

Cells migrating through complex 3D environments experience considerable physical challenges including tensile stress and compression. To move, cells need to resist these forces whilst also squeezing the large nucleus through confined spaces. This requires highly coordinated cortical contractility. Microtubules can both resist compressive forces and sequester key actomyosin regulators to ensure appropriate activation of contractile forces. Yet, how these two roles are integrated to achieve nuclear transmigration in 3D is largely unknown. Here, we demonstrate that compression triggers reinforcement of a dedicated microtubule structure at the rear of the nucleus by the mechanoresponsive recruitment of CLASPs (cytoplasmic linker-associated proteins) which dynamically strengthens and repairs the lattice. These reinforced microtubules form the mechanostat: an adaptive feedback mechanism that allows the cell to both withstand compressive force and spatiotemporally organise contractility signalling pathways. The microtubule mechanostat facilitates nuclear positioning and coordinates force production to enable the cell to pass through constrictions. Disruption of the mechanostat imbalances cortical contractility, stalling migration and ultimately resulting in catastrophic cell rupture. Our findings reveal a new role for microtubules as cellular sensors which detect and respond to compressive forces, enabling movement and ensuring survival in mechanically demanding environments. One Sentence SummaryMechanically tuned microtubules form a mechanostat to coordinate contractility and nuclear positioning in confined migration.

cell biology↗