Biology subjects

Sahlgren, C.

Publications and source records attributed to Sahlgren, C..

6 recordsLinked to original sources

bioRxiv2025

Receptor stoichiometry predicts artery-typical vulnerability to altered Notch signaling during smooth muscle differentiation.

The development and maintenance of arterial smooth muscle cells (SMCs) rely on Jagged1-Notch2/Notch3 signaling. While Notch2 and Notch3 are thought to function redundantly during SMC development, clinical and experimental evidence suggests artery-specific importance for the two receptors. Combining in vitro, in vivo, and in silico models, we report that the canonical Notch signaling during SMC differentiation is largely driven by Notch2. While Notch2 and Notch3 co-regulate a large group of genes in SMCs upon Jagged1 interaction, Notch3 is a less potent inducer of Notch signaling than Notch2 and requires higher doses to potentiate a meaningful transcriptional response due to its weak interaction with RBPJ{kappa}. Consequently, Notch2 depletion abolishes Notch signaling in the SMCs of the large elastic arteries. However, high Jagged1 and Notch3 expression in smaller arteries like those in the brain can compensate for Notch2 loss. This work refines our mechanistic understanding of Notch signaling in the SMCs and offers region-specific insights into the Notch-related arterial diseases.

developmental biology↗

bioRxiv2024

Phosphoproteomic analysis reveals the diversity of signaling behind ErbB inhibitor-induced phenotypes

The impact of kinase inhibitors on the phosphoproteome has been rarely investigated at a whole organism level. Here we performed a phosphoproteomic analysis in embryonic zebrafish to identify the signaling pathways perturbed by ErbB receptor tyrosine kinase inhibitors at the organism level. The phosphorylation of proteins associated with the PI3K/Akt, p38 MAPK, Notch, Hippo/Yap and {beta}-catenin signaling pathways were differentially regulated by the ErbB inhibitors. Gene set enrichment analyses indicated differential neurological and myocardial phenotypes of different ErbB inhibitors. To assess the neurological and myocardial effects, motility and ventricle growth assays were performed on zebrafish embryos treated with the ErbB and downstream signaling pathway inhibitors. The treatment with the inhibitors targeting the PI3K/Akt, p38 MAPK, and Notch signaling pathways along with the ErbB inhibitors AG1478 and Lapatinib perturbed the overall movement and ventricle wall growth of zebrafish embryos. Taken together, these results indicate that inhibitors with the same primary targets can affect different signaling pathways while eliciting similar physiological phenotypes.

cell biology↗

bioRxiv2023

Environmental stiffness restores mechanical homeostasis in vimentin-depleted cells

Recent experimental evidence indicates a role for the intermediate filament vimentin in regulating cellular mechanical homeostasis, but its precise contribution remains to be discovered. Mechanical homeostasis requires a balanced bi-directional interplay between the cells microenvironment and the cellular morphological and mechanical state - this balance being regulated via processes of mechanotransduction and mechanoresponse, commonly referred to as mechanoreciprocity. Here, we systematically analyze vimentin-expressing and vimentin-depleted cells in a swatch of in vitro cellular microenvironments varying in stiffness and/or ECM density. We find that vimentin-expressing cells maintain mechanical homeostasis by adapting cellular morphology and mechanics to micromechanical changes in the microenvironment. However, vimentin-depleted cells lose this mechanoresponse ability on short timescales, only to reacquire it on longer time scales. Indeed, we find that the morphology and mechanics of vimentin-depleted cell in stiffened microenvironmental conditions can get restored to the homeostatic levels of vimentin-expressing cells. Additionally, we observed vimentin-depleted cells increasing collagen matrix synthesis and its crosslinking, a phenomenon which is known to increase matrix stiffness, and which we now hypothesize to be a cellular compensation mechanism for the loss of vimentin. Taken together, our findings provide further insight in the regulating role of intermediate filament vimentin in mediating mechanoreciprocity and mechanical homeostasis.

biophysics↗

bioRxiv2021

Optogenetic control of NOTCH1 signalling

The Notch signalling pathway is a crucial regulator of cell differentiation as well as tissue organisation. Dysregulation of Notch signalling has been linked to the pathogenesis of different diseases. Notch plays a key role in breast cancer progression by controlling the interaction between the tumour cells and the microenvironment as well as by increasing cell motility and invasion. NOTCH1 is a mechanosensitive receptor, where mechanical force is required to activate the proteolytic cleavage and release of the Notch intracellular domain (NICD). Here, we circumvent this step by regulating Notch activity by light. To achieve this, we have engineered a membrane-bound optogenetic NOTCH1 receptor (optoNotch) to control the activation of NOTCH1 intracellular domain (N1ICD) and its downstream transcriptional activities. Using optoNotch we confirm that NOTCH1 activation increases cell proliferation in MCF7 and MDA-MB-468 breast cancer cells in 2D and spheroid 3D cultures. OptoNotch allows fine-tuning ligand-independent regulation of N1ICD to understand the spatiotemporal complexity of Notch signalling.

cell biology↗

bioRxiv2020

iGIST - a kinetic bioassay for pertussis toxin based on its effect on inhibitory GPCR signaling

Detection of pertussis toxin (PTX) activity is instrumental for the development and manufacturing of pertussis vaccines. These quality and safety measures require annually thousands of mice. Here, we describe iGIST (Interference in Gi-mediated Signal Transduction) - an animal-free kinetic bioassay for detection of PTX by measuring its effect on inhibitory G protein-coupled receptor (GPCR) signaling. PTX ADP-ribosylates inhibitory -subunits of the heterotrimeric G proteins, thereby perturbing the inhibitory GPCR signaling. iGIST is based on HEK293 cells co-expressing a somatostatin receptor 2 (SSTR2), which is an inhibitory GPCR controllable by a high affinity agonist octreotide, and a luminescent 35-cyclic adenosine monophosphate (cAMP) probe. iGIST has a low sensitivity threshold in picogram/ml range of PTX, surpassing by 100-fold in a parallel analysis the currently used in vitro end-point technique to detect PTX, the cluster formation assay (CFA) in Chinese hamster ovary cells. iGIST also detects PTX in complex samples, i.e. a commercial PTX- toxoid containing pertussis vaccine that was spiked with an active PTX. iGIST has an objective digital readout and is observer-independent, offering prospects for automation. iGIST emerges as a promising animal-free alternative to detect PTX activity in the development and manufacturing of pertussis vaccines. iGIST is also expected to facilitate basic PTX research, including identification and characterization of novel compounds interfering with PTX.

microbiology↗

bioRxiv2019

Dynamic-ultrastructural cell volume (3D) correlative microscopy facilitated by intracellular fluorescent nanodiamonds as multi-modal probes

Three-dimensional correlative light and electron microscopy (3D CLEM) are attaining popularity as a potential technique to explore the functional aspects of a cell together with high-resolution ultrastructural details across the cell volume. In order to perform such a 3D CLEM experiment, there is an imperative requirement for multi-modal probes that are both fluorescent and electron-dense. These multi-modal probes will serve as landmarks in matching up the large full cell volume datasets acquired by different imaging modalities. Fluorescent nanodiamonds (FNDs) are a unique nanosized, fluorescent, and electron-dense material from the nanocarbon family. We hereby propose a novel and straightforward method for executing 3D CLEM using FNDs as multi-modal landmarks. We demonstrate that FNDs is biocompatible and easily identified both in living cell fluorescence imaging and in serial block-face scanning electron microscopy (SB-EM). We illustrate the 3D CLEM method by registering multi-modal datasets.

cell biology↗