Biology subjects

Vos, H. R.

Publications and source records attributed to Vos, H. R..

8 recordsLinked to original sources

bioRxiv2026

Degenerated intervertebral disc environment impairs notochordal cell-derived extracellular vesicles release and their matrix anabolic effect

At the onset of intervertebral disc degeneration, within the disc core, the pH and osmolarity decrease, and the residing notochordal cells (NCs) gradually transition towards nucleus pulposus cells (NPCs). How these microenvironmental cues shape the NCs extracellular vesicles (EV)-enriched secretome, and thus EV-mediated communication with NPCs during this transition, remains poorly understood. To study this, we collected the secretome from pig NC-rich tissue cultured for 4 days in either healthy or degenerate disc media to mimic these changes. In both conditions, NC-rich tissues were largely comparable at the histological and biochemical levels. Despite, tissues released glycosaminoglycans (GAGs), depleting the extracellular matrix. Surprisingly, degenerative media did not differentially release inflammatory regulators, though it reduced PGE2 release. We asked whether this extended to EV-enriched secretome media (SM_EV+), and found that the degenerative media reduced the number of EVs without altering their morphology or size. We then determined NC-EV association of inflammatory and matrix regulators. NC-EV isolation enriched MMP1, IL6 and IL10 and depleted soluble GAGs. Conversely, EV-depletion (SM_EV-) removed most GAGs without affecting MMP1, IL6, and IL10, suggesting that they contribute to the NC-EV soft corona. Functionally, healthy SM_EV+ improved GAG production by NPCs, but attenuated TBXT expression. Degenerate SM_EV+ did not elicit detectable EV-specific effects. These findings suggest that, in health, secretome-mediated communication from NCs to NPCs is only partially EV-mediated. At the onset of IVD degeneration, low pH and osmolarity impair the release of NC-EVs and negate the EV-specific beneficial matrix-anabolic effects on NPCs, contributing to the NC-to-NPC transition.

cell biology↗

bioRxiv2026

Hydrogen Peroxide induces resistance to DNA damage in a localization and p53 dependent manner.

Organisms need to be able to adapt to a changing environment in order to survive. The adaptive response invoked by a low dose of a stressor resulting in resistance to high levels of that stressor is known as hormesis and can even lead to lifespan extension of organisms. The exact mechanisms underlying stress-induced hormesis are unknown, although multiple studies pose mitochondria-derived Reactive Oxygen Species (ROS, e.g. H2O2) as an important contributor. Here we used chemo-genetic H2O2 production as a model to study ROS-dependent adaptive responses in a localization-dependent manner. We found that brief, sublethal H2O2 production at the nucleosomes provides p53-dependent resistance to a subsequent high dose of H2O2, whereas mitochondrial H2O2 production, surprisingly, does not. A multi-omics approach revealed that p53-induced hormesis is accompanied by metabolic rewiring that boosts reductive capacity, and that the increased stress resistance can mostly be attributed to its downstream target p21. Importantly, brief p53 stabilization also mounted protection against chemotherapy-induced DNA damage, suggesting that p53-dependent hormesis could be exploited to selectively protect healthy, p53-wildtype tissue from chemotherapy in the treatment of patients with p53 mutant tumors.

cancer biology↗

bioRxiv2025

Multi-omics analysis of TNBC organoids identifies phosphorylation of the membrane trafficking machinery as a key event associated with FER-mediated invasion

Triple Negative Breast Cancer (TNBC) is characterised by unfavourable outcome due to the combination of its metastatic propensity, chemo-refractory behaviour and the lack of effective targeted interventions. Expression of the feline sarcoma-related (FER) kinase constitutes an independent prognostic factor that correlates with poor patient survival. FER promotes invasive behaviour in TNBC cells by regulating endosomal sorting and recycling (ESR) of adhesion proteins. Yet, the ESR machinery supporting invasion in TNBC, particularly within 3D environments, remains poorly understood. Here, we used FER-expressing TNBC patient-derived xenograft organoids (PDXOs) and MDA-MB-231 cells to identify the membrane trafficking machinery promoting invasion. Using a combination of proteomics, phospho-proteomics, and single cell RNA-sequencing, we show that the invasion of FER-expressing PDXO cells in collagen-I is mainly associated with the differential phosphorylation of membrane trafficking regulators, including SEC16A and a marked increase in Rab4-positive tubules. SEC16A depletion impairs cell invasion and reduces the number of focal adhesions and Rab4-positive tubules. Importantly, FER regulates SEC16A levels and localization, specifically in TNBC. Altogether, we identified SEC16A as a key player in FER-driven TNBC invasion, highlighting the membrane trafficking machinery as a promising target for the development of new therapeutic strategies.

cancer biology↗

bioRxiv2025

Unconventional components complement the cryptic kinetochore of the ciliate Tetrahymena thermophila

The accurate segregation of chromosomes is mediated by kinetochores, multi-protein structures that connect centromeric chromatin to the dynamic microtubules of the spindle apparatus. Comparative genomics surveys predict a complex kinetochore in the last eukaryotic common ancestor (LECA) and recurrent loss or replacement of its subcomplexes across the eukaryotic tree of life. Understanding kinetochore composition and organization in diverse lineages can reveal the trajectories of kinetochore evolution in eukaryotes and aid in dissecting the function of each subcomplex. Tetrahymena thermophila is a unicellular eukaryote of the phylum Ciliophora with a largely elusive kinetochore composition. Here, we leverage proximity proteomics coupled to deep homology detection approaches to identify 16 kinetochore proteins in T. thermophila, dubbed KiTTs (Kinetochore of Tetrahymena thermophila 1-16). We find that nine KiTTs (3-9 + 15-16) are cryptic orthologs of conventional kinetochore proteins that previously remained undetected due to extensive sequence divergence. Four KiTTs (10-13) are not orthologous to known subunits and therefore represent unconventional kinetochore proteins. Super-resolution imaging places three of these novel proteins (KiTT10/11/13) at the inner kinetochore, whereas the fourth (KiTT12) localizes near the MIS12 complex at the outer kinetochore. RNAi-mediated depletion of KiTT12 reduces levels of the outer kinetochore protein KiTT1NDC80 and causes chromosome segregation errors, showcasing a bona fide role at the kinetochore. Our work reveals a unique kinetochore composition in a ciliate, providing new insights into the evolution of an essential cellular protein machine.

cell biology↗

bioRxiv2025

Bioengineering Developmentally Inspired Matrix Vesicles as Designer Nanotherapeutics for Bone Regeneration

Extracellular vesicles (EVs) are emerging as promising acellular nanotherapeutics for musculoskeletal repair. Matrix vesicles, a matrix-bound subset of EVs, are essential mediators of endochondral ossification in bone development and fracture repair. This study aims to design bioengineered matrix vesicles from hypertrophic cartilage microtissues to drive endochondral ossification for bone repair. Human mesenchymal stromal cell (hBMSC) microtissues were differentiated with/without BMP2 in chondrogenic or hypertrophic medium. Isolated matrix vesicles were characterized for physiochemical properties and biological functionality. BMP2 and hypertrophic conditioning significantly increased vesicle yield (1.5-fold), alkaline phosphatase activity (3.24-fold), calcium binding capacity (8.82-fold), and growth factor content (BMP2, VEGF). These vesicles promoted proliferation, migration, and mineralization of hBMSCs and enhanced angiogenesis in human endothelial colony forming cells (hECFCs), with BMP2 and hypertrophically conditioned vesicles showing the most pronounced effects. Proteomics analysis confirmed the enrichment of proteins involved in extracellular matrix remodelling, mineral deposition and vascularization within these hypertrophically engineered vesicles. These findings demonstrate that hypertrophic induction of cartilaginous microtissues substantially improves the yield and therapeutic potential of matrix vesicles. Taken together, this research unveils a powerful strategy to bioengineer developmentally inspired vesicles that not only recapitulate key cues of endochondral ossification but offers a tailorable, multifunctional nanotherapeutic platform for improved bone regeneration strategies. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=112 SRC="FIGDIR/small/684111v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@17327a9org.highwire.dtl.DTLVardef@1310583org.highwire.dtl.DTLVardef@160f095org.highwire.dtl.DTLVardef@152eff0_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2025

Phosphoproteomics Maps Calcineurin-NFAT-DSCR1.4 Signaling as Druggable Axis in Gαq-R183Q-Driven Capillary Malformations

Vascular malformations are congenital lesions caused by somatic and germline mutations that disrupt developmental signaling pathways. Capillary malformations (CMs) typically present as port-wine stains in the skin and can also affect ocular and cerebral tissues in Sturge Weber Syndrome (SWS), leading to aesthetic, ophthalmic, and neurological complications. CMs are caused by a somatic mutation in the GNAQ gene in endothelial cells, leading to a p.R183Q substitution in the Gq protein. The underlying mechanisms of Gq-R183Q-driven CMs formation remain unclear. To address this, we generated CRISPR/Cas9-engineered human dermal microvascular endothelial cells lacking endogenous Gq, whilst expressing the Gq-R183Q mutant instead. The Gq-R183Q mutation strongly impaired endothelial cell migration and angiogenic sprouting capacity compared to wild-type controls. Next, using SILAC-based quantitative proteomics, we investigated the Gq-R183Q-induced changes in the endothelial phosphoproteome. These analyses revealed prominent activation of the calcineurin-NFAT signaling pathway in Gq-R183Q-expressing endothelial cells, leading to dephosphorylation of NFAT1 and NFAT2 and the selective expression of their transcriptional target DSCR1.4. Immunofluorescence of patient-derived skin biopsies confirmed deregulation of NFAT1/2 and the expression of DSRC1 in endothelial cells, validating their potential importance in CMs. We further demonstrate that pharmacological inhibition of calcineurin with tacrolimus (FK506) could partially restore NFAT signaling in Gq-R183Q endothelial cells. Intriguingly, the genetic depletion of the NFAT target DSCR1 in Gq-R183Q cells fully restored calcineurin/NFAT signaling to normal levels, enabling proper endothelial migration and sprouting. In summary, we uncovered a calcineurin-NFAT-DSCR1.4 signal transduction axis that is driven by Gq-R183Q and established its importance for endothelial angiogenic properties. These findings highlight the calcineurin/NFAT signaling axis as a promising therapeutic target to restore endothelial function in CMs.

cell biology↗

bioRxiv2025

E-cadherin mechanotransduction activates EGFR-ERK signaling in epithelial cells by inducing ADAM-mediated ligand shedding

The behavior of cells is governed by signals originating from their local environment, including mechanical forces that cells experience. Forces are transduced by mechanosensitive proteins, which can impinge on signaling cascades that are also activated by growth factor receptors upon ligand binding. We investigated the crosstalk between these mechanical and biochemical signals in the regulation of intracellular signaling networks in epithelial monolayers. Phosphoproteomic and transcriptomic analyses on epithelial monolayers subjected to mechanical strain revealed ERK signaling as a predominant strain-activated hub, initiated at the level of the upstream epidermal growth factor receptor (EGFR). Strain-induced EGFR-ERK signaling depends on mechanosensitive E-cadherin adhesions. Proximity labeling identified the metalloproteinase ADAM17, an enzyme that mediates shedding of soluble EGFR ligands, to be closely associated with E-cadherin. We developed a novel probe for monitoring ADAM-mediated shedding, which demonstrated that mechanical strain induced ADAM activation. Mechanically-induced ADAM activation was essential for mechanosensitive signaling from E-cadherin adhesions towards EGFR-ERK. Collectively, our data demonstrate that mechanical strain transduced by E-cadherin adhesion triggers the shedding of EGFR ligands that stimulate downstream downstream ERK activity. Our findings illustrate how mechanical signals and biochemical ligands can operate within a single, linear signaling cascade. Significance statementCells integrate different types of information that they receive from their local environment to regulate their behavior. This includes biochemical signals, such as growth factors binding to their dedicated receptor. Similarly, cells respond to mechanical forces that they are subjected to. Although biochemical and mechanical signals can elicit similar signaling responses in cells, the interplay between these types of signals is not well understood. Here we unveil that mechanical strain of epithelia modulates the activity of the EGFR-ERK signaling pathway by controlling the availability of growth factors that bind and activate EGFR. This finding demonstrates that biochemical and mechanical signals do not act in a segregated fashion, but rather can function in a linear cascade, shedding light on fundamental principles governing cellular regulation.

cell biology↗

bioRxiv2023

A farnesyl-dependent structural role for CENP-E in expansion of the fibrous corona

Correct chromosome segregation during cell division depends on proper connections between spindle microtubules and kinetochores. During prometaphase, kinetochores are temporarily covered with a dense protein meshwork known as the fibrous corona. Formed by oligomerization of ROD/ZW10/ZWILCH-SPINDLY (RZZ-S) complexes, the fibrous corona promotes spindle assembly, chromosome orientation and spindle checkpoint signaling. The molecular requirements for formation of the fibrous corona are not fully understood. Here we show that the fibrous corona depends on the mitotic kinesin CENP-E, and that poorly expanded fibrous coronas after CENP-E depletion are functionally compromised. This previously unrecognized role for CENP-E does not require its motor activity but instead is driven by farnesyl modification of its C-terminal kinetochore-and microtubule-binding domain. We show that in cells CENP-E interacts with RZZ-S complexes in a farnesyl-dependent manner. CENP-E is recruited to kinetochores following RZZ-S, and - while not required for RZZ-S oligomerization per se - promotes subsequent fibrous corona expansion. Our comparative genomics analyses suggest that the farnesylation motif in CENP-E orthologs emerged alongside the full RZZ-S module in an ancestral lineage close to the fungi-animal split (Obazoa), revealing potential conservation of the mechanisms for fibrous corona formation. Our results show that proper spindle assembly has a potentially conserved non-motor contribution from the kinesin CENP-E through stabilization of the fibrous corona meshwork during its formation.

cell biology↗