Biology subjects

Rutkowska, A.

Publications and source records attributed to Rutkowska, A..

5 recordsLinked to original sources

bioRxiv2023

Integration of Thermal Proteome Profiling with phosphoproteomic and transcriptomic data via mechanistic network models decodes the molecular response to PARP inhibition

The deregulation of complex diseases often spans multiple molecular processes. A multimodal functional characterization of these processes can shed light on the disease mechanisms and the effect of drugs. Thermal Proteome Profiling (TPP) is a mass-spectrometry based technique assessing changes in thermal protein stability that can serve as proxies of functional changes of the proteome. These unique insights of TPP can complement those obtained by other omics technologies. Here, we show how TPP can be integrated with phosphoproteomics and transcriptomics in a network-based approach using COSMOS, a framework for causal integration of multi-omics, to provide an integrated view of transcription factors, kinases and proteins with altered thermal stability. This allowed us to recover known mechanistic consequences of PARP inhibition in ovarian cancer cells on cell cycle and DNA damage response in detail and to uncover new insights into drug response mechanisms related to interferon and hippo signaling. We found that TPP complements the other omics data and allowed us to obtain a network model with higher coverage of the main underlying mechanisms. These results illustrate the added value of TPP, and more generally the power of network models to integrate the information provided by different omics technologies. We anticipate that this strategy can be used to broadly integrate functional proteomics with other omics to study complex molecular processes. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=63 SRC="FIGDIR/small/553354v1_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@af952eorg.highwire.dtl.DTLVardef@16b1d1borg.highwire.dtl.DTLVardef@1440787org.highwire.dtl.DTLVardef@14a4227_HPS_FORMAT_FIGEXP M_FIG C_FIG

systems biology↗

bioRxiv2023

Systemic inflammation differentially modulates the levels of EBI2 and CH25H/CYP7B1 enzymes in the brain microvascular cells.

The endogenous ligand for the EBI2 receptor, oxysterol 7,25OHC, crucial for immune responses, is finely regulated by CH25H, CYP7B1 and HSD3B7 enzymes. Lymphoid stromal cells and follicular dendritic cells within T cell follicles maintain a gradient of 7,25OHC, with stromal cells increasing and dendritic cells decreasing its concentration. This gradient is pivotal for proper B cell positioning in lymphoid tissue. In the animal model of multiple sclerosis, the experimental autoimmune encephalomyelitis, the levels of 7,25OHC rapidly increase in the central nervous system driving the migration of EBI2 expressing immune cells through the blood-brain barrier (BBB). To explore if blood vessel cells in the brain express these enzymes, we examined normal mouse brain microvessels and studied their expression changes during inflammation. EBI2 was abundantly expressed in endothelial cells, pericytes/smooth muscle cells, and astrocytic endfeet. CH25H, CYP7B1, and HSD3B7 were variably detected in each cell type, suggesting their active involvement in oxysterol 7,25OHC synthesis and gradient maintenance under normal conditions. Under acute inflammatory conditions, EBI2 and synthesizing enzyme modulation occurred in brain vasculature, with variations based on the enzyme and cell type. Significant species-specific differences emerged in EBI2 and enzyme levels between mouse and human BBB-forming cells. Overall, our investigation suggest a direct role of the brain microvascular cells in regulating 7,25OHC levels, shedding light on potential therapeutic targets for neuroinflammatory disorders.

neuroscience↗

bioRxiv2023

The proton-activated receptor TDAG8 is upregulated in oligodendrocytes during maturation and under acidic conditions.

Acidosis is one of the hallmarks of demyelinating central nervous system (CNS) lesions in multiple sclerosis (MS). Response to acidic pH is primarily mediated by a family of G protein-coupled proton-sensing receptors: OGR1, GPR4, and TDAG8. These receptors are inactive at alkaline pH, while at acidic pH they are maximally activated. Genome-wide association studies identified a locus within the TDAG8 gene to be associated with several autoimmune diseases including MS. Notably, we here found that TDAG8 expression is upregulated in MS plaques which prompted us to explore the expression and function of TDAG8 in the CNS in human MO3.13 oligodendrocytes in vitro and in vivo in the lipopolysaccharide-induced neuroinflammation model. We found that TDAG8 is upregulated in maturing oligodendrocytes and temporarily under acidic conditions. Acidic pH also induces oligodendrocyte branching, inhibits chemotaxis and affects the expression of oligodendrocyte maturation markers, PDGFR and MBP in vitro. Even though myelination was not affected in the adult TDAG8-deficient mice, the expression of human and murine TDAG8 was strongly regulated upon inflammation in vivo in the brain and in vitro in lipopolysaccharide and pro-inflammatory cytokine-treated oligodendrocytes. Together these findings point toward a potential role of TDAG8 in oligodendrocyte biology, neuroinflammation and pathophysiology of MS and provide new directions for further scientific enquiry.

neuroscience↗

bioRxiv2018

Proteome thermal stability reflects organ physiology and identifies drug-target engagement in vivo

Studying biological processes at a molecular level and monitoring drug-target interactions is established for simple cell systems but challenging in vivo. We introduce and apply a methodology for proteome-wide thermal stability measurements to characterize organ physiology and activity of many fundamental biological processes across tissues, such as energy metabolism and protein homeostasis. This method, termed tissue thermal proteome profiling (tissue-TPP), also enabled target and off-target identification and occupancy measurements in tissues derived from animals dosed with the non-covalent histone deacetylase inhibitor, panobinostat. Finally, we devised blood-CETSA, a thermal stability-based method to monitor target engagement in whole blood. Our study generates the first proteome-wide map of protein thermal stability in tissue and provides tools that will be of great impact for translational research.

cell biology↗

bioRxiv2018

Characterization of novel inhibition of indoleamine 2,3-dioxygenase by targeting its apo form.

Indoleamine-2,3-dioxygenase 1 (IDO1) is a heme-containing enzyme that catalyzes the rate-limiting step in the kynurenine pathway of tryptophan (TRP) metabolism. As an inflammation-induced immunoregulatory enzyme, pharmacological inhibition of IDO1 activity is currently being pursued as a potential therapeutic tool for the treatment of cancer and other disease states. As such, a detailed understanding of the mechanism of action of established and novel IDO1 inhibitors remains of great interest. Comparison of a newly-developed IDO1 inhibitor (GSK5628) to the existing best-in-class compound, epacadostat (Incyte), allows us to report on a unique inhibition mechanism for IDO1. Here, we demonstrate that GSK5628 inhibits IDO1 by competing with heme for binding to a heme-free conformation of the enzyme (apo-IDO1) while epacadostat coordinates its binding with the iron atom of the IDO1 heme cofactor. Comparison of these two compounds in cellular systems reveals a long-lasting inhibitory effect of GSK5628, undescribed for other known IDO1 inhibitors. Detailed characterization of this apo-binding mechanism for IDO1 inhibition may help design superior inhibitors or may confer a unique competitive advantage over other IDO1 inhibitors vis-a-vis specificity and pharmacokinetic parameters.

biochemistry↗