Biology subjects

Schulze, N.

Publications and source records attributed to Schulze, N..

5 recordsLinked to original sources

bioRxiv2026

Heparan sulfate-function is essential for Integrin-dependent cell-matrix interactions and regulates glycosaminoglycan synthesis synthesis through YAP

Extracellular matrix (ECM) is the main component of cartilage, making it an ideal environment to study cell-matrix interactions. Among ECM constituents, heparan sulfate (HS)-carrying proteoglycans (PGs) are of particular interest since they are not only structural components but are also involved in cell matrix adhesion and signalling processes. We previously demonstrated that transgenic mice with a clonal loss of HS synthesis in chondrocytes (Col2-rtTA-Cre;Ext1e2fl/e2fl) develop clusters of enlarged cells in the articular cartilage (AC), which are surrounded by a glycosaminoglycan (GAG)-rich ECM. This led to the questions how HS regulate the molecular composition and mechanical properties of the ECM, how they sense alterations in the HS structure and how they respond to it. We stained tissue sections of Col2-rtTA-Cre;Ext1e2fl/e2f animals and detected increased levels of chondroitin sulfate (CS), Aggrecan (Acan), Perlecan (Pcan), Matrilin (Matn)-3 and-4, Collagen type II (Col2) and Col9, while Col12 was abolished in the HS-deficient clusters. We assessed the stiffness of the mutant matrix by Atomic Force Microscopy (AFM) and found that it was markedly softer than the surrounding, HS-containing tissue. Likely in response to this altered texture, HS-deficient clones showed increased protein levels of Integrin pathway components. To model a loss of HS-function in vitro, we treated murine embryonic fibroblasts (MEFs) with the HS-antagonist Surfen. Treatment during cell adhesion resulted in impaired cell-substrate adhesion, increased formation of filopodia-like membrane protrusions, decreased cell polarisation and migration, reduced formation of FA and SF, and a translocation of YAP into the cytoplasm. Similarly, we observed reduced cell polarisation in HS-deficient CHO pgsD-667 cells, which could not be rescued by external presentation of HS. When MEFs were treated with Surfen after the completion of the initial cell adhesion process, inhibition of HS-function led to an increased formation of FA and SF, in line with the increased levels of Integrin pathway components observed in HS-deficient chondrocytes in vivo. We detected high levels of Yes1-associated protein (YAP) in the HS-deficient clusters, and we investigated the effect of YAP modulation on high density micromass cultures from primary murine chondroprogenitors. YAP activation induced an increased GAG synthesis similar to Surfen, while YAP inactivation partially abolished the effect of Surfen, showing that YAP acts downstream of HS function and controls GAG synthesis. Taken together, we demonstrated that HS-function is essential for Integrin-dependent cell-matrix interactions. Information on the impaired cell matrix adhesion upon loss of HS is conveyed into the nucleus via YAP, which at least partially controls the synthesis of GAGs in chondrocytes.

cell biology↗

bioRxiv2025

Targeted degradation of USP7 in solid cancer cells reveals disparate effects of deubiquitinase inhibition vs. acute protein depletion

Proteolysis-targeting chimeras (PROTACs) co-op the ubiquitin system for targeted protein degradation, creating opportunities to interrogate cellular functions of proteins through "chemical knockdown". However, matched pairs of protein degraders and inhibitors, that possess high specificity and chemical complementarity, for individual components of the ubiquitin system have remained scarce. This includes reagents to modulate activity and abundance of deubiquitinases (DUBs), which critically regulate ubiquitin-mediated signaling. Here, using an integrated chemical biology approach, we explored the cellular function of the DUB USP7 as a case study comparing inhibition and degradation of this DUB in melanoma and pancreatic cancer cells. Through the synthesis of a degrader library, we identified potent USP7 PROTACs for each cancer type, established BRET-based ternary complex formation and quantified degradation efficiency. USP7 degraders and their cognate inhibitor were subsequently employed to characterize treatment-induced phenotypic alterations. Proteomic and cellular analyses revealed that highly specific degradation of USP7 modulated both shared and distinct protein sets across cancer cell types, without impacting cell growth. Notably, cellular responses to USP7 degradation differed markedly from those to USP7 inhibition. Moreover, our data uncovered broad proteomic and metabolic changes induced by prolonged USP7 inhibitor treatment. Collectively, our work provides a chemical toolbox of comprehensively characterized reagents to distinguish on-target phenotypes which will aid the understanding of the role of USP7 in malignant diseases. More broadly, our data emphasize the importance of increased specificity via PROTAC-mediated degradation and the potential of this modality to distinguish catalytic from non-catalytic as well as cell-line specific functions of DUBs.

biochemistry↗

bioRxiv2025

ATXN3 regulates lysosome regeneration after damage by targeting K48-K63-branched ubiquitin chains

The cellular response to lysosomal damage involves fine-tuned mechanisms of membrane repair, lysosome regeneration and lysophagy, but how these different processes are coordinated is not fully understood. Here we show in human cells that the deubiquitinating enzyme ATXN3 helps restore integrity of the lysosomal system after damage by targeting K48-K63-linked branched ubiquitin chains on regenerating lysosomes. We find that ATXN3 translocates to lysosomes after different types of damage and there colocalizes with its partner, VCP/p97. ATXN3 recruitment occurs late after the initial repair of a subset of lysosomes and after phagophore formation on terminally damaged lysosomes. Of note, inactivation of ATXN3 by induced degradation, depletion or knock-out impairs clearance of damaged lysosomes and full restoration of lysosomal capacity. Mechanistically, ATXN3, along with VCP/p97, turns over K48-K63-branched ubiquitin conjugates on LAMP1-positive, phosphatidylinositol-(4,5)-bisphosphate-decorated compartments that are not yet fully re-acidified indicating involvement in lysosome regeneration. Our findings identify a key role of ATXN3 in restoring lysosomal function after lysosomal membrane damage and uncover K48-K63-branched ubiquitin chain-regulated regeneration as a critical element of the lysosomal damage stress response.

cell biology↗

bioRxiv2025

Optogenetic stimulation of Lbc GEF-mediated Rho activity dynamics promotes cell invasion

Cancer cell invasion relies on dynamic cell shape changes, which originate from protrusive and contractile intracellular forces. Previous studies revealed that contractile forces are controlled by positive-feedback amplification of the contraction regulator Rho by Lbc GEFs. These GEFs were previously linked to tumor progression, however, the underlying mechanisms are poorly understood. Here, we generated a mouse melanoma model, in which cytosolic levels of the Lbc GEF GEF-H1 are controlled by light. Using this model, we found that increased GEF-H1 levels strongly stimulate cell contraction dynamics. Interestingly, increased contraction dynamics rapidly induced expansion of tumor spheroids via a focal adhesion kinase-dependent mechanism. Furthermore, long-term stimulation led to the escape of individual cells from spheroids. These findings reveal new insights into the oncogenic roles of Lbc GEFs, and how they might promote tumor cell invasion. We propose a mechanism, in which increased cell contraction dynamics results in asymmetric pulling forces at the tumor border, promoting the detachment and escape of individual cells.

cell biology↗

bioRxiv2024

A non-autonomous protein quality control mechanism targeting tau aggregate propagation

Tauopathies such as Alzheimers disease, frontotemporal dementia with Parkinsonism, and other neurodegenerative disorders are characterized by the spread of tau pathology from an initial brain region to neuroanatomically connected areas. At the molecular level, spreading involves aggregation of tau in a donor cell, externalization of transmissible fragments of amyloid fibrils, internalization by an acceptor cell, followed by seeded aggregation of endogenous tau. However, the protein quality control mechanisms that counteract tau aggregation, and in particular its spreading process, are not well understood. In this context, a co-migrating factor performing location-independent interference of fibril formation and transmission would be an appropriate conceptual solution. Here, we show that the cell-to-cell transfer of the widely conserved serine protease HTRA1 impedes tau pathology by targeting multiple steps within the spreading process. Our results suggest a defense mechanism against the intercellular spread of pathogenic protein conformations.

biochemistry↗