Biology subjects

Grosshans, J.

Publications and source records attributed to Grosshans, J..

12 recordsLinked to original sources

bioRxiv2026

HSPB8 regulates CTP synthase filaments to couple nucleotide metabolism and autophagy in tumors

Metabolic adaptation and proteostasis are essential for tumor survival under nutrient stress, yet how these processes are mechanistically integrated remains unclear. Here, we identify CTP synthase (CTPS) filament dynamics as a regulatory nexus linking nucleotide metabolism to autophagic flux. Under glutamine deprivation--a hallmark of poorly vascularized solid tumors--CTPS undergoes polymerization into filamentous assemblies that exhibit reduced enzymatic activity. We demonstrate that filament formation is driven by intracellular asparagine availability and stabilized by the accumulation of misfolded proteins. Using APEX2-based proximity labeling and proteomics, we identify the small heat shock protein HSPB8 as a filament-associated regulator. HSPB8, acting within the chaperone-assisted selective autophagy (CASA) pathway, antagonizes CTPS polymerization by promoting clearance of misfolded proteins, thereby restoring soluble, catalytically active CTPS. Filament disassembly increases CTP production and enhances synthesis of autophagy-related phospholipids, including phosphatidylinositol and phosphatidylethanolamine, resulting in accelerated autophagic flux. Cells expressing filament-deficient CTPS mutants display elevated autophagosome formation and increased LC3-II accumulation upon lysosomal blockade, confirming enhanced flux. In vivo, disruption of CTPS filament assembly impairs xenograft tumor growth and is associated with excessive autophagy. Analysis of TCGA datasets further reveals that high CTPS and low HSPB8 expression correlate with poor patient survival across multiple cancers. Collectively, our findings establish CTPS filament dynamics as a proteostasis-sensitive metabolic switch that coordinates nucleotide biosynthesis with autophagy, revealing a previously unrecognized vulnerability in tumors adapting to nutrient limitation.

cell biology↗

bioRxiv2024

Developmental control of E-cadherin junctions by mechanical contractility in Drosophila embryos

Adherens junctions are very plastic structures that change their composition and structure depending on the state of the epithelial tissue, involving mechanical stress and tissue dynamics. In Drosophila embryos, adherens junctions mature only during gastrulation following their formation a few minutes before during cellularization. Although the developmental maturation is obvious during gastrulation, the factors and conditions controlling it have remained unclear. Here, we assay the levels, distribution pattern, and mobility of E-cadherin during Drosophila gastrulation. Indicating a maturation, we find an increase in total levels at junctions and a drop in the mobile fraction of E-cadherin molecules. Both developmental changes depend on Myo-II contractility. Consistent with stereotypic Myo-II regulation, we find that interference with Rho signaling leads to corresponding changes in E-cadherin levels and mobility, at least in part. Besides Rho signaling, Src42A contributes to junction maturation, as tested by both loss-function and gain-of-function situations. Together, we demonstrate developmental maturation of adherens junctions and identify Rho signaling and Src42A as upstream regulatory pathways.

developmental biology↗

bioRxiv2023

Polychaetoid/ZO-1 strengthens cell junctions under tension while localizing differently than core adherens junction proteins

During embryonic development dramatic cell shape changes and movements re-shape the embryonic body plan. These require robust but dynamic linkage between the cell-cell adherens junctions and the force-generating actomyosin cytoskeleton. Our view of this linkage has evolved, and we now realize linkage is mediated by a mechanosensitive multiprotein complex assembled via multivalent connections. Here we combine genetic, cell biological and modeling approaches to define the mechanism of action and functions of an important player, Drosophila Polychaetoid, homolog of mammalian ZO-1. Our data reveal that Pyd reinforces cell junctions under elevated tension, and facilitates cell rearrangements. Pyd is important to maintain junctional contractility and in its absence cell rearrangements stall. We next use structured illumination microscopy to define the molecular architecture of cell-cell junctions during these events. The cadherin-catenin complex and Cno both localize to puncta along the junctional membrane, but are differentially enriched in different puncta. Pyd, in contrast, exhibits a distinct localization to strands that extend out from the region occupied by core junction proteins. We then discuss the implications for the protein network at the junction-cytoskeletal interface, suggesting different proteins localize and function in distinct ways but combine to produce robust connections.

cell biology↗

bioRxiv2022

PSC niche develops into immune-responsive blood cells capable of transdifferentiating into lamellocytes in Drosophila

Drosophila blood cells called hemocytes form an efficient barrier against infections and tissue damage. During metamorphosis, hemocytes undergo tremendous changes in their shape and behavior preparing them for tissue clearance. Yet, the diversity and functional plasticity of pupal blood cells have not been explored. Here, we combine single-cell transcriptomics and high-resolution microscopy to dissect the heterogeneity and plasticity of pupal hemocytes. We identified precursor and effector hemocytes with distinct molecular signatures and cellular functions clearly distinct from other stages of hematopoiesis. Strikingly, we identified that PSC cells, which function as lymph gland niche, are highly migratory and immune responsive cells in the pupa. PSC cells can transdifferentiate to lamellocytes triggered by wasp infection. Altogether, our data highlight a remarkable cell heterogeneity, and identifies a cell population that acts not only as a stem cell niche in larval hematopoiesis, but functions as cell reservoir to pupal and adult blood cells.

cell biology↗

bioRxiv2019

Exfoliated near infrared fluorescent CaCuSi4O10 nanosheets with ultra-high photostability and brightness for biological imaging

Imaging of complex (biological) samples in the near infrared (nIR) range of the spectrum is beneficial due to reduced light scattering, absorption, phototoxicity and autofluorescence. However, there are only few near infrared fluorescent materials known and suitable for biomedical applications. Here, we exfoliate the layered pigment CaCuSi4O10 (known as Egyptian Blue, EB) via facile tip sonication into nanosheets (EB-NS) with ultra-high nIR fluorescence stability and brightness. The size of EB-NS can be tailored by tip sonication to diameters < 20 nm and heights down to 1 nm. EB-NS fluoresce at 910 nm and the total fluorescence intensity scales with the number of Cu2+ ions that serve as luminescent centers. Furthermore, EB-NS display no bleaching and ultra-high brightness compared to other nIR fluorophores. The versatility of EB-NS is demonstrated by in vivo single-particle tracking and microrheology measurements in developing Drosophila embryos. Additionally, we show that EB-NS can be uptaken by plants and remotely detected in a low cost stand-off detection setup despite strong plant background fluorescence. In summary, EB-NS are a highly versatile, bright, photostable and biocompatible nIR fluorescent material that has the potential for a wide range of bioimaging applications both in animal and plant systems.

biophysics↗

bioRxiv2019

Emergence of the subapical domain is associated with the midblastula transition

Epithelial domains and cell polarity are determined by polarity proteins which are associated with the cell cortex in a spatially restricted pattern. Early Drosophila embryos are characterized by a stereotypic dynamic and de novo formation of cortical domains. For example, the subapical domain emerges at the transition from syncytial to cellular development during the first few minutes of interphase 14. The dynamics in cortical patterning is revealed by the subapical markers Canoe/Afadin and ELMO-Sponge, which widely distributed in interphase 13 but subapically restricted in interphase 14. The factors and mechanism determining the timing for the emergence of the subapical domain have been unknown. In this study, we show, that the restricted localization of subapical markers depends on the onset of zygotic gene expression. In contrast to cell cycle remodeling, the emergence of the subapical domain does not depend on the nucleo-cytoplasmic ratio. Thus, we define cortical dynamics and specifically the emergence of the subapical domain as a feature of the midblastula transition.\n\nAuthor summaryMidblastula transition is a paradigm of a developmental transition. Multiple processes such as cell cycle, cell mobility, onset of zygotic gene expression, degradation of maternal RNA and chromatin structure are coordinated to lead to defined changes in visible morphology. The midblastula transition in Drosophila embryos is associated with a change from fast nuclear cycles to a cell cycle mode with gap phase and slow replication, a strong increase in zygotic transcription and cellularization. The timing of the processes associated with the midblastula transition are controlled by the onset of zygotic gene expression or the nucleocytoplasmic ratio. Here we define the patterning of cortical domains, i. e. the emergence of a subapical domain as a novel feature of the midblastula transition whose appearance is controlled by the onset of zygotic transcription but not the nucleocytoplasmic ratio. Our findings will help to gain further understanding of the coordination of complex developmental processes during the midblastula transition.

developmental biology↗

bioRxiv2019

An emergent flow of the nuclear array in syncytial embryos

Many aspects in tissue morphogenesis are attributed to the collective behavior of the participating cells. Yet, the mechanism for emergence of dynamic tissue behavior is not understood completely. Here we report the "yo-yo"-like nuclear drift movement in Drosophila syncytial embryo displays typical emergent feature of collective behavior, which is associated with pseudo-synchronous nuclear division cycle. We uncover the direct correlation between the degree of asynchrony of mitosis and the nuclear collective movement. Based on experimental manipulations and numerical simulations, we find the ensemble of spindle elongation, rather than a nucleus own spindle, is the main driving force for its drift movement. The cortical F-actin acts as viscoelastic medium to dampen the movements and plays a critical role in restoring the nuclear positions after a mitosis cycle. Our study provides insights into how the interactions between cytoskeleton as individual elements leads to collective movement of the nuclear array on a macroscopic scale.

developmental biology↗

bioRxiv2018

Automated Segmentation of Epithelial Tissue Using Cycle-Consistent Generative Adversarial Networks

A central problem in biomedical imaging is the automated segmentation of images for further quantitative analysis. Recently, fully convolutional neural networks, such as the U-Net, were applied successfully in a variety of segmentation tasks. A downside of this approach is the requirement for a large amount of well-prepared training samples, consisting of image - ground truth mask pairs. Since training data must be created by hand for each experiment, this task can be very costly and time-consuming. Here, we present a segmentation method based on cycle consistent generative adversarial networks, which can be trained even in absence of prepared image - mask pairs. We show that it successfully performs image segmentation tasks on samples with substantial defects and even generalizes well to different tissue types.

bioinformatics↗

bioRxiv2018

Optochemical control of cell contractility at single cell resolution during tissue morphogenesis

AbstractsThe spatial and temporal dynamics of cell contractility plays a key role in tissue morphogenesis, wound healing and cancer invasion. Here we report a simple, single cell resolution, optochemical method to induce reversible minute-scale cell contractions in vivo during morphogenesis. We employed the photolabile Ca2+ chelator o-nitrophenyl EGTA to induce bursts of intracellular free Ca2+ by laser photolysis. Ca2+ bursts appear within seconds and are restricted to individual target cells. Cell contraction reliably followed within a minute, to about half of the cross-sectional area. Increased Ca2+ levels and contraction were reversible and the target cells further participated in tissue morphogenesis. Cell contractions are paralleled with non-muscle myosin-II accumulation in the apico-medial cortex, indicating that Ca2+ bursts trigger non-muscle myosin II activation. Our approach can be easily adapted to many experimental systems and species, as no specific genetic elements are required and a widely used reagent is employed.

developmental biology↗

bioRxiv2017

A "molecular guillotine" reveals an interphase function of Kinesin-5

Motor proteins are important for transport and force generation in a variety of cellular processes and morphogenesis. Here we design a general strategy for conditional motor mutants by inserting a protease cleavage site at the \"neck\" between the head domain and the stalk of the motor protein, making the protein susceptible to proteolytic cleavage at the neck by the corresponding protease. To demonstrate the feasibility of this approach, we inserted the cleavage site of TEV protease into the neck of the tetrameric motor Kinesin-5. Application of TEV protease led to a specific depletion and functional loss of Kinesin-5 in Drosophila embryos. By this, we revealed that Kinesin-5 stabilized the microtubule network during interphase in syncytial embryos. The \"molecular guillotine\" can potentially be applied to many motor proteins due to the conserved structures of kinesin, dynein and myosin with accessible necks.\n\nAuthor summaryWe design a general strategy for conditional motor mutants by inserting a protease cleavage site between head and stalk domain of the motor protein, making it susceptible to specific proteolytic cleavage. We demonstrate the feasibility of the approach with the motor Kinesin-5 and the protease TEV in Drosophila embryos. This approach can potentially be applied to motor proteins kinesin, dynein and myosin due to the conserved structures.

developmental biology↗

bioRxiv2017

Expression of lamina proteins Lamin Dm0 and Kugelkern suppresses stem cell proliferation

The nuclear lamina is involved in numerous cellular functions, such as gene expression, nuclear organization, nuclear stability, and cell proliferation. The mechanism underlying the involvement of lamina is often not clear, especially in physiological contexts. Here we investigate how the farnesylated lamina proteins Lamin Dm0 and Kugelkern are linked to proliferation control of intestinal stem cells (ISCs) in adult Drosophila flies by loss-of-function and gain-of-function experiments. We found that ISCs mutant for Lamin Dm0 or Kugelkern proliferate, whereas overexpression of Lamin Dm0 or Kugelkern strongly suppressed proliferation. The anti-proliferative activity is, at least in part, due to suppression of Jak/Stat but not Delta/Notch signalling. Lamin Dm0 expression suppresses Jak/Stat signalling by normalization of about 50% of the Stat target genes in ISCs.\n\nAuthor summaryThe nuclear lamina is a protein meshwork that lies beneath the inner side of the nuclear membrane and interacts with nuclear pores, chromatin and the cytoskeleton. Changes in proteins of the nuclear lamina cause a wide range of diseases which are often not well understood. It is hypothesized that impairment of stem cell function, as a result of lamina changes, might play a key role in some of those diseases. Here we use the well characterized Drosophila midgut as a system to investigate the role of lamina proteins Lamin Dm0 and Kugelkern on stem cell proliferation.

developmental biology↗

bioRxiv2017

Protein phosphatase V ensures timely cell cycle remodeling during the mid-blastula transition in Drosophila

Cell cycle remodeling from fast nuclear cycles to a generic cell cycle mode is a major feature of the mid-blastula transition (MBT) in Drosophila. Remodeling occurs when Twine/Cdc25 falls below a critical threshold. Timing is based on Twine destabilization induced by zygotic transcription. It is conceivable that appropriate starting levels are also important for timely reaching the threshold. Mechanisms for controlling Twine levels at the onset of MBT are unknown. Here we identify a function of the protein phosphatase V in this mechanism. Twine was increased in PpV mutants, whereas the decay rate was comparable to wildtype. PpV mutants frequently underwent an extra nuclear division. We detected PpV-dependent phosphosites in Twine. Phosphosite mutants contain higher Twine levels and frequently underwent an extra nuclear division, comparable to PpV mutants. Our data support a model that the cell cycle remodeling is controlled by induced destabilization and PpV-d ependent control of Twine levels.

developmental biology↗