Biology subjects

Tol, S.

Publications and source records attributed to Tol, S..

5 recordsLinked to original sources

bioRxiv2025

Transcriptome-Proteome analysis of human naive and memory B cell subsets reveal isotype and subclass-specific phenotypes

Antibodies produced by B cells aid in recognition and clearance of pathogens and is the cornerstone of vaccination strategies. Humans produce nine different antibody isotypes and their effector functions differ according to the type of antigen and route of exposure. Phenotypic variation between isotype-switched B cell subsets is expected but not studied in detail. To obtain a molecular definition of isotype-defined cell identity, we performed proteomics and transcriptomics on isotype-defined populations of human naive and memory B cells (MBCs): CD27-IgM+IgD+, CD27+CD38lo/-IgM+IgD+, CD27+CD38lo/-IgM+IgD-, and IgA1, IgA2, IgG1, IgG2, IgG3, and IgG4 MBCs (CD27+CD38lo/-Ig+). Combined proteome and transcriptome analysis revealed that mRNA and protein expression profiles separate isotype-defined B cell subsets according to their differentiation status. mRNA and protein expression levels correlated reasonably well for many genes. IgG4-switched B cells were most distinct from naive B cells in terms of mRNA as well as protein expression profiles. Besides a distinct expression profile of cytokine and Fc receptors, we identified a high expression of IgE-coding mRNA in IgG4-switched B cells. SDR16C5 was identified as uniquely upregulated in IgG4-switched B cells. Taken together, this study highlights the distinct phenotypic profile of IgG4-switched B cells.

immunology↗

bioRxiv2024

Metastatic tumor cells in bone marrow differ from paired neuroblastoma tumor and contain subsets with therapy-resistant characteristics

Bone marrow (BM) is a common site for solid tumor metastasis, often causing poor outcome. Here, we define the characteristics of BM-disseminated tumor cells (DTCs) using neuroblastoma as a model. We combined single-cell RNA-sequencing (scRNA-seq) and cell-surface protein analysis using 7 paired BM and primary tumor (PT) samples and found that DTCs contain a higher percentage of cycling cells and higher expression of neurodevelopmental genes compared to corresponding PT cells. In 6 patients, the copy number variation profile differed between PT cells and DTCs, indicating spatial heterogeneity. Within the BM, we detected dormant DTCs with potentially reduced chemosensitivity; this population contained cells expressing low levels of the immunotherapeutic antigen GD2 and increased NGFR expression. In conclusion, we characterized DTCs that are particularly challenging to target, offering new avenues for developing therapeutic strategies designed to target all subpopulations within the highly complex metastatic site, thereby preventing the development of drug-resistant clones.

cancer biology↗

bioRxiv2022

Opto-RhoGEFs: an optimized optogenetic toolbox to reversibly control Rho GTPase activity on a global to subcellular scale, enabling precise control over vascular endothelial barrier strength

The inner layer of blood vessels consists of endothelial cells, which form the physical barrier between blood and tissue. This vascular barrier is tightly regulated to allow the passage of essential molecules like oxygen, carbon-dioxide, water, ions, and nutrients. The vascular endothelial barrier is defined by cell-cell contacts through adherens and tight junctions. To further investigate the signaling in the endothelium that regulates vascular barrier strength, we focused on Rho GTPases, regulators of the actin cytoskeleton and known to control junction integrity. Rho GTPase signaling is confined in space and time. To manipulate the signaling in a temporal and spatial manner we applied optogenetics. Guanine exchange factor (GEF) domains from ITSN1, TIAM1 and p63RhoGEF, activating Cdc42, Rac and Rho respectively, were integrated into the optogenetic recruitment tool iLID. This tool allows for activation at the subcellular level in a reversible and non-invasive manner and thereby to recruit a GEF to local areas at the plasma membrane, enabling the local activation of specific Rho GTPases. The membrane tag of iLID was optimized and a HaloTag was applied to gain more flexibility for multiplex imaging. The resulting Opto-RhoGEFs were tested in an endothelial cell monolayer and demonstrated precise temporal control of vascular barrier strength by a cell-cell overlap-dependent, VE-cadherin-independent, mechanism. Furthermore, Opto-RhoGEFs enabled precise optogenetic control in endothelial cells over morphological features such as cell-size, -roundness, local extension, and cell contraction. In conclusion, we have optimized and applied the optogenetic iLID GEF recruitment tool i.e. Opto-RhoGEFs, to study the role of Rho GTPases in the vascular barrier of the endothelium and found that membrane protrusions at the junction region can rapidly increase barrier integrity independent of VE-cadherin.

cell biology↗

bioRxiv2021

Visualizing endogenous RhoA activity with an improved localization-based, genetically encoded biosensor

Rho GTPases are regulatory proteins, which orchestrate cell features such as morphology, polarity and movement. Therefore, probing Rho GTPase activity is key to understanding processes such as development, cell migration and wound healing. Localization-based reporters for active Rho GTPases are attractive probes to study Rho GTPase-mediated processes, in real time with subcellular resolution in living cells and tissue. Until now, relocation RhoA biosensors seem to only be useful in certain organisms and have not been characterized well. In this paper, we systematically examined the contribution of the fluorescent protein and RhoA binding peptides, on the performance of localization-based sensors. To test the performance, we compared relocation efficiency and specificity in cell-based assays. We identified several improved localization-based, genetically encoded, fluorescent biosensors for detecting endogenous RhoA activity. This enables a broader application of RhoA relocation biosensors, which was demonstrated by using the improved biosensor to visualize RhoA activity, during cell division, during random migration, at the Golgi membrane and induced by G protein-coupled receptor signaling. Due to the improved avidity of the new biosensors for RhoA activity, cellular processes regulated by RhoA can be better understood. O_FIG O_LINKSMALLFIG WIDTH=134 HEIGHT=200 SRC="FIGDIR/small/430250v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@1847c24org.highwire.dtl.DTLVardef@f22c92org.highwire.dtl.DTLVardef@14c4aa8org.highwire.dtl.DTLVardef@1b9657d_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2021

Endothelial junctional membrane protrusions serve as hotspots for neutrophil transmigration

Upon inflammation, leukocytes rapidly transmigrate across the endothelium to enter the inflamed tissue. Evidence accumulates that leukocytes use preferred exit sites, though it is not yet clear how these hotspots in the endothelium are defined and how they are recognized by the leukocyte. Using lattice light sheet microscopy, we discovered that leukocytes prefer endothelial membrane protrusions at cell junctions for transmigration. Phenotypically, these junctional membrane protrusions are present in an asymmetric manner, meaning that one endothelial cell shows the protrusion and the adjacent one does not. Consequently, leukocytes cross the junction by migrating underneath the protruding endothelial cell. These protrusions depend on Rac1 activity and by using a photo-activatable Rac1 probe, we could artificially generate local exit-sites for leukocytes. Overall, we have discovered a new mechanism that uses local induced junctional membrane protrusions to facilitate/steer the leukocyte escape/exit from inflamed vessel walls.

cell biology↗