Biology subjects

Weiss, T.

Publications and source records attributed to Weiss, T..

At least 37 records · Page 2Linked to original sources

bioRxiv2023

Structural basis of CMKLR1 signaling induced by chemerin9

Chemokine-like receptor 1 (CMKLR1), also known as chemerin receptor 23 (ChemR23) or chemerin receptor 1, is a chemoattractant G protein-coupled receptor (GPCR) that responds to the adipokine chemerin and is highly expressed in innate immune cells, including macrophages and neutrophils. The signaling pathways of CMKLR1 can lead to both pro- and anti-inflammatory effects depending on the ligands and physiological contexts. To understand the molecular mechanisms of CMKLR1 signaling, we determined a high-resolution cryo-electron microscopy (cryo-EM) structure of the CMKLR1-Gi signaling complex with chemerin9, a nanopeptide agonist derived from chemerin, which induced complex phenotypic changes of macrophages in our assays. The cryo-EM structure, together with molecular dynamics simulations and mutagenesis studies, revealed the molecular basis of CMKLR1 signaling by elucidating the interactions at the ligand-binding pocket and the agonist-induced conformational changes. Our results are expected to facilitate the development of small molecule CMKLR1 agonists that mimic the action of chemerin9 to promote the resolution of inflammation.

biophysics↗

bioRxiv2023

Efficient protein tagging and cis-regulatory element engineering via precise and directional oligonucleotide-based targeted insertion in plants

Efficient and precise targeted insertion holds great promise but remains challenging in plant genome editing. An efficient NHEJ-mediated targeted insertion method was recently developed by combining CRISPR-Cas9 with phosphorothioate modified double-stranded oligodeoxynucleotides (dsODNs). Yet this approach often led to imprecise insertions with no control over the insertion direction. In this study, we first quantitatively compared the impact of the chemical protection on efficiency of targeted insertion. With the observation that CRISPR-SpCas9 could frequently induce staggered cleavages with 5' 1-nucleotide overhangs, we then evaluated the impact of the donor end structures on the direction and preciseness of targeted insertions. Our study demonstrated that the chemically protected dsODNs with 5' 1-nt overhangs could significantly improve the precision and direction control of target insertions in all tested CRIPSR targeting sites. Lastly, we applied this method to endogenous gene tagging in Setaria viridis, and cis-regulatory element engineering for disease resistance in rice. Two distinct TAL effector binding elements were directionally inserted into the promoter region of a recessive rice bacterial blight resistance gene at up to 24.4% efficiency. The resulting rice lines with heritable insertions exhibited strong resistance to the infection of Xanthomonas oryzae pv. oryzae pathogen in an inducible and strain-specific manner.

plant biology↗

bioRxiv2023

BCAT1 inhibition affects CD8+ T cell activation, exhaustion, and tumoral immunity by altering iron homeostasis

The present study explores the role of the cytosolic branched chain amino acid aminotransferase (BCAT1) in CD8+ T cell activation, in general, and tumor immunity, in particular, and identifies a non-canonical function of the protein in iron homeostasis. Pharmacologic inhibition of BCAT1 using the novel drug ERG245 abrogates the effector functions of CD8+ T cells in vitro and metabolically reprograms the cells towards increased OXPHOS. In vivo, it suppresses activation of CD8+ T cells in DSS colitis leading to improved disease outcomes. Remarkably, withdrawal of BCAT1 inhibition further amplifies OXPHOS and gives rise to CD8+ T cells with increased cytotoxicity in vitro and in vivo. When combined with an anti-PD-1 treatment, temporal BCAT1 inhibition dramatically increases anti-PD-1 efficacy inducing complete and durable tumor regressions in the moderately immunogenic CT26 tumor model. Single cell RNA-seq data link expression of Bcat genes to exhausted T cells within the tumor microenvironment of human cancer patients, whereas in vitro assays indicate that BCAT1 inhibition partially prevents the adoption of a terminally exhausted phenotype by CD8+ T cells. We propose BCAT1 as a target for cancer combinatory therapies. SIGNIFICANCEThe study explores for the first time the role of BCAT1 in CD8+ T cell activation and proposes novel strategies for using BCAT1 inhibitors in cancer and beyond. It demonstrates that BCAT1 exerts its function without significantly altering branched chain amino acid (BCAA) levels through a mechanism that controls iron homeostasis, a novel non-canonical mechanism of action, and implicates BCAT1 in the adoption of an exhausted phenotype by T cells found in human cancers. While the majority of metabolic drugs temper OXPHOS, it demonstrates that an agent that increases OXPHOS in CD8+ T cells can be used successfully as an immune-oncology drug.

immunology↗

bioRxiv2022

The Siglec-sialic acid-axis is a target for innate immunotherapy of glioblastoma

Glioblastoma (GBM) is the most aggressive form of primary brain tumor, for which effective therapies are urgently needed. Cancer cells are capable of evading clearance by phagocytes such as microglia and monocyte-derived cells through engaging tolerogenic programs. Here, we found that high level of Siglec-9 expression correlates with reduced survival in GBM patients. Using conditional knockouts of Siglec-E, the murine functional homologue of Siglec-9, together with single-cell RNA sequencing, we demonstrated significant pro-phagocytosis effects in microglia and monocyte-derived cells in the absence of Siglec-E. Loss of Siglec-E on monocyte-derived cells enhances antigen cross-presentation and production of pro-inflammatory cytokines, resulting in more efficient T cell priming. This bridging of innate and adaptive responses delays tumor growth and results in prolonged survival. Further, we showed synergistic activity of Siglec-E blockade in combinatorial immunotherapies and demonstrate its translational potential against GBM.

cancer biology↗

bioRxiv2022

Targeting tumour-intrinsic neural vulnerabilities of glioblastoma

Glioblastoma is the most common yet deadliest primary brain cancer1. The neural behavior of glioblastoma, including the formation of synaptic circuitry and tumour microtubes, is increasingly understood to be pivotal for disease manifestation2-9. Nonetheless, the few approved treatments for glioblastoma target its oncological nature, while its neural vulnerabilities remain incompletely mapped and clinically unexploited. Here, we systematically survey the neural molecular dependencies and cellular heterogeneity across glioblastoma patients and diverse model systems. In 27 surgical patient samples, we identify cancer cell morphologies indicative of poor prognosis, and discover repurposable neuroactive drugs with anti-glioblastoma efficacy by image-based drug screening. Glioblastoma cells exhibit functional dependencies on highly expressed neuroactive drug targets, while interpretable molecular machine learning (COSTAR) reveals their downstream convergence on AP-1-driven tumour suppression. This drug-target connectivity signature is confirmed by accurate in silico drug screening on >1 million compounds, as well as by multi-omic profiling of glioblastoma drug responses. Thus, Ca2+-driven AP-1 pathway induction represents a tumour-intrinsic vulnerability at the intersection of oncogenesis and neural activity-dependent signaling. Opportunities for clinical translation of this neural vulnerability are epitomized by the antidepressant Vortioxetine synergizing with current standard of care treatments in vivo. Together, the results presented here provide a mechanistic foundation and conceptual framework for the treatment of glioblastoma based on its neural origins.

cancer biology↗

bioRxiv2022

Precise and versatile microplate reader-based analyses of biosensor signals from arrayed microbial colonies

Genetically encoded fluorescent biosensors have emerged as a powerful tool to support phenotypic screenings of microbes. Optical analyses of fluorescent sensor signals from colonies grown on solid media can be challenging as imaging devices need to be equipped with appropriate filters matching the properties of fluorescent biosensors. Towards versatile fluorescence analyses of different types of biosensor signals derived from arrayed colonies, we investigate here the use of monochromator equipped microplate readers as an alternative to imaging approaches. Indeed, for analyses of the LacI-controlled expression of the reporter mCherry in Corynebacterium glutamicum, or promoter activity using GFP as reporter in Saccharomyces cerevisiae, an improved sensitivity and dynamic range was observed for a microplate reader-based analyses compared to their analyses via imaging. The microplate reader allowed us to capture signals of ratiometric fluorescent reporter proteins (FRPs) with a high sensitivity and thereby to further improve the analysis of internal pH via the pH-sensitive FRP mCherryEA in Escherichia coli colonies. Applicability of this novel technique was further demonstrated by assessing redox states in C. glutamicum colonies using the FRP Mrx1-roGFP2. By the use of a microplate reader, oxidative redox shifts were measured in a mutant strain lacking the non-enzymatic antioxidant mycothiol (MSH), indicating its major role for maintaining a reduced redox state also in colonies on agar plates. Taken together, analyses of biosensor signals from microbial colonies using a microplate reader allows comprehensive phenotypic screenings and thus facilitates further development of new strains for metabolic engineering and systems biology.

synthetic biology↗

bioRxiv2022

Combined sensor-based monitoring of mycothiol redox potential and DNA-damage response in Corynebacterium glutamicum

Excessive amounts of reactive oxygen species (ROS) can cause irreversible damages to essential cellular components such as DNA. Genetically encoded biosensors targeting oxidative stress and DNA-stress have emerged to a powerful analytical tool to assess physiological states in a non-invasive manner. In this study, we aimed to combine the redox biosensor protein Mrx1-roGFP2 with a transcriptional biosensor for DNA-damage based on the PrecA promoter fused to a reporter gene (e2-crimson) in Corynebacterium glutamicum. Therefore, the redox biosensor strains C. glutamicum WT_Mrx1-roGFP2 and the mycothiol (MSH)-deficient mutant strain C. glutamicum {Delta}mshC_Mrx1-roGFP2 were equipped with the DNA-stress reporter plasmid pJC1_PrecA_e2-crimson. Exposure of the double-sensor equipped C. glutamicum WT strain to hypochlorite resulted in an oxidative redox shift, accompanied by an induction of the DNA-stress reporter system. In absence of the major non-enzymatic antioxidant MSH, the induction of the DNA-stress response was even more pronounced. This confirms the linkage of oxidative stress and DNA-damage response, and therefore making antioxidants a crucial player to protect DNA. Furthermore, exposure of the double biosensor strains to a DNA-damage inducing agent resulted in an oxidative redox shift. These results suggest a direct link of DNA-damage and oxidative stress response in C. glutamicum. Finally, we observed that inhibition of cell wall biosynthesis by penicillin caused both an oxidative redox shift and a DNA-damage response in C. glutamicum. The excellent compatibility of Mrx1-roGFP2 with E2-Crimson shown here provides a powerful combinatorial biosensor concept for in-depth studies of redox-related physiology in future studies.

microbiology↗

bioRxiv2022

Human repair-related Schwann cells adopt functions of antigen-presenting cells in vitro

The plastic potential of Schwann cells (SCs) is increasingly recognized to play a role after nerve injury and in diseases of the peripheral nervous system. In addition, reports on the interaction between SCs and immune cells indicate their involvement in inflammatory processes. However, data about the immunocompetence of human SCs are primarily derived from neuropathies and it is currently unknown whether SCs directly regulate an adaptive immune response after nerve injury. Here, we performed a comprehensive analysis of the immunomodulatory capacities of human repair-related SCs (hrSCs), which recapitulate SC response to nerve injury in vitro. We used our previously established protocol for the culture of primary hrSCs from human peripheral nerves and analyzed the transcriptome, secretome, and cell surface proteins for signatures and markers relevant in innate and adaptive immunity, performed phagocytosis assays, and monitored T-cell subset activation in co-cultures with autologous human T-cells. Our findings show that hrSCs are highly phagocytic, which is in line with high MHCII expression. In addition, hrSCs express co-regulatory molecules, such as CD40, CD80, B7H3, CD58, CD86, HVEM, release a plethora of chemoattractants, matrix remodelling proteins and pro- as well as anti-inflammatory cytokines, and upregulate the T-cell inhibiting PD-L1 molecule upon pro-inflammatory stimulation with IFN{gamma}. Furthermore, hrSC contact reduced the number and activation status of allogenic CD4+ and CD8+ T-cells. This study demonstrates that hrSCs possess features and functions typical for professional antigen presenting cells in vitro, and suggest a new role of these cells as negative regulators of T-cell immunity during nerve regeneration. Main pointsO_LIHuman repair-related Schwann cells (hrSC) function as professional antigen presenting cells. C_LIO_LIHrSCs up-regulate PD-L1 upon pro-inflammatory IFN{gamma} stimulation. C_LIO_LIHrSCs hamper CD4+ and CD8+ T-cell activation. C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/483322v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@1b5fbb7org.highwire.dtl.DTLVardef@a0de85org.highwire.dtl.DTLVardef@1710ee9org.highwire.dtl.DTLVardef@189e3bc_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2022

Drastic differential CRISPR-Cas9 induced mutagenesis influenced by DNA methylation and chromatin features

CRISPR-Cas9 mediated genome editing has been widely adopted for basic and applied biological research in eukaryotic systems. While many studies considered DNA sequences of CRISPR target sites as the primary determinant for CRISPR mutagenesis efficiency and mutation profiles, increasing evidence revealed the significant role of chromatin context. Nonetheless, most of the prior studies were limited by the lack of sufficient epigenetic resources and/or by only transiently expressing CRISPR-Cas9 in a short time window. In this study, we leveraged the wealth of high-resolution epigenomic resources in Arabidopsis thaliana to address the impact of chromatin features on CRISPR-Cas9 mutagenesis using stable transgenic plants. Our results indicated that DNA methylation and chromatin features could lead to significant variations in mutagenesis efficiency by up to 250 folds. Low mutagenesis efficiencies were mostly associated with repressive heterochromatic features. This repressive effect appeared to persist through cell divisions but could be alleviated through substantial reduction of DNA methylation at CRISPR target sites. Moreover, specific chromatin features, such as H3K4me1, H3.3, and H3.1, appear to be associated with significant variations in CRISPR-Cas9 mutation profiles reflected by the 1 bp insertion rates. Our findings provided strong evidence that specific chromatin features could have significant and lasting impacts on both CRISPR-Cas9 mutagenesis efficiency and DNA double strand break repair outcomes.

plant biology↗

bioRxiv2022

Limited consequences for loss of RNA-directed DNA methylation in Setaria viridis domains rearranged methyltransferase (DRM) mutants

The Domains Rearranged Methyltransferases (DRMs) are crucial for RNA-directed DNA methylation (RdDM) in plant species. Setaria viridis is a model monocot species with a relatively compact genome that has limited transposable element content. CRISPR-based genome editing approaches were used to create loss-of-function alleles for the two putative functional DRM genes in S. viridis to probe the role of RdDM. The analysis of drm1ab double mutant plants revealed limited morphological consequences for the loss of RdDM. Whole-genome methylation profiling provided evidence for wide-spread loss of methylation in CHH sequence contexts, particularly in regions with high CHH methylation in wild-type plants. Evidence was also found for locus-specific loss of CG and CHG methylation, even in some regions that lack CHH methylation. Transcriptome profiling identified a limited number of genes with altered expression in the drm1ab mutants. The majority of genes with elevated CHH methylation directly surrounding the transcription start site or in nearby promoter regions do not have altered expression in the drm1ab mutant even when this methylation is lost, suggesting limited regulation of gene expression by RdDM. Detailed analysis of the expression of transposable elements identified several transposons that are transcriptionally activated in drm1ab mutants. These transposons likely require active RdDM for maintenance of transcriptional repression. Significance statementMethylation profiling of Setaria viridis plants that lack functional Domains Rearranged Methyltransferase genes reveal widespread loss of DNA methylation in the CHH sequence context. Transcriptome analysis reveals a small set of genes and transposons that are silenced by RNA-directed DNA methylation.

plant biology↗

bioRxiv2021

Impedance flow cytometry for viability analysis of Corynebacterium glutamicum

Corynebacterium glutamicum efficiently produces glutamate when growth is inhibited. Analyses of viability in this non-growing state requires time consuming plating and determination of colony forming units. We here establish impedance flow cytometry measurements to assess the viability of non-growing, glutamate producing C. glutamicum cultures within minutes.

microbiology↗

bioRxiv2021

The epigenetic evolution of gliomas is determined by their IDH1 mutation status and treatment regimen

Tumor adaptation or selection is thought to underlie therapy resistance of gliomas. To investigate the longitudinal epigenetic evolution of gliomas in response to therapeutic pressure, we performed an epigenomic analysis of 143 matched initial and recurrent patients with IDH-wildtype (IDHwt) and IDH-mutant (IDHmut) gliomas. IDHwt gliomas showed a longitudinally stable epigenome with relatively low levels of global methylation, whereas the epigenome of IDHmut gliomas showed initial high levels genome-wide of DNA methylation that was progressively reduced to levels similar to those of IDHwt tumors. By integrating DNA methylation and gene expression data, adaptive changes of putative master regulators of the cell cycle and of differentiation were seen in IDHmut recurrent tumors. Furthermore, relapses of IDHmut tumors were accompanied by histological progression which in turn influenced survival, as validated in an independent cohort. Finally, the initial cell composition of the tumor microenvironment differed between IDHwt and IDHmut tumors and changed differentially following treatment, suggesting increased neo-angiogenesis and T-cell infiltration upon treatment for IDHmut gliomas. Our study provides one of the largest cohorts of paired glioma samples profiled with epigenomics, transcriptomics and genomics; and our results demonstrate that the treatment of IDHmut gliomas reshapes the epigenome towards an IDHwt-like phenotype. Accordingly, the prevalent practice of early genotoxic treatment in this patient population may need to be revisited.

genomics↗

bioRxiv2021

Visualizing the pH in Escherichia coli colonies via the sensor protein mCherryEA allows high-throughput screening of mutant libraries

Cytoplasmic pH is tightly regulated by diverse active mechanisms and interconnected regulatory processes in bacteria. Many processes and regulators underlying pH-homeostasis have been identified via phenotypic screening of strain libraries towards non-growth at low or high pH values. Direct screens with respect to changes of the internal pH in mutant strain collections are limited by laborious methods including fluorescent dyes or radioactive probes. Genetically encoded biosensors equip single organisms or strain libraries with an internal sensor molecule already during the generation of the strain. In this study, we used the pH-sensitive mCherry variant mCherryEA as ratiometric pH biosensor. We visualized the internal pH of E. coli colonies on agar plates by the use of a Gel-Doc imaging system. Combining this imaging technology with robot-assisted colony picking and spotting allowed us to screen and select mutants with altered internal pH values from a small transposon mutagenesis derived E. coli library. Identification of the TN- insertion sites in strains with altered internal pH levels revealed that the transposon was inserted into trkH (encoding a transmembrane protein of the potassium uptake system) or the rssB gene (encoding the anti-adaptor protein RssB which mediates the proteolytic degradation of the general stress response regulator RpoS), two genes known to be associated with pH-homeostasis and pH stress adaptation. This successful screening approach demonstrates that the pH- sensor based analysis of arrayed colonies on agar plates is a sensitive approach for the fast identification of genes involved in pH-homeostasis or pH stress adaptation in E. coli. ImportancePhenotypic screening of strain libraries on agar plates has become a versatile tool to understand gene functions and to optimize biotechnological platform organisms. Screening is supported by genetically encoded biosensors that allow to easily measure intracellular processes. For this purpose, transcription-factor-based biosensors have emerged as the sensor-type of choice. Here, the target stimulus initiates the activation of a response gene (e.g. a fluorescent protein) followed by transcription, translation and maturation. Due to this mechanistic principle, biosensor readouts are delayed and cannot report the actual intracellular state of the cell in real-time. To capture fast intracellular processes adequately, fluorescent reporter proteins are extensively applied. But these sensor-types are not utilized for phenotypic screenings so far. To take advantage of their properties, we here established an imaging method, which allows to apply a fast ratiometric sensor protein for assessing the internal pH of colonies in a high-thoughput manner.

microbiology↗

bioRxiv2021

Resolving the molecular architecture of the photoreceptor active zone by MINFLUX nanoscopy

Cells assemble macromolecular complexes into scaffoldings that serve as substrates for catalytic processes. Years of molecular neurobiology indicate that neurotransmission depends on such optimization strategies, yet the molecular topography of the presynaptic Active Zone (AZ) where transmitter is released upon synaptic vesicle (SV) fusion remains to be visualized. Therefore, we implemented MINFLUX optical nanoscopy to resolve the AZ of rod photoreceptors. To facilitate MINFLUX nanoscopy of the AZ, we developed and verified an immobilization technique, we name Heat Assisted Rapid Dehydration (HARD). Here fresh retinal slices are directly stamped onto glass coverslips yielding a single layer of rod AZs. These AZs exhibited excellent labeling efficiency and minimal signal redundancy in the Z-direction. Our data indicate that the SV release site is a molecular complex of bassoon-Rab3-binding molecule 2 (RIM2)-ubMunc13-2-CAST. The complexes are serially duplicated longitudinally, and reflected in register along the axis of symmetry of the synaptic ribbon. One sentence summaryStructural motifs formed by active zone proteins at the photoreceptor synapse.

neuroscience↗

bioRxiv2021

Longitudinal analysis of diffuse glioma reveals cell state dynamics at recurrence associated with changes in genetics and the microenvironment

To interrogate the factors driving therapy resistance in diffuse glioma, we collected and analyzed RNA and/or DNA sequencing data from temporally separated tumor pairs of 292 adult patients with IDH-wild-type or IDH-mutant glioma. Tumors recurred in distinct manners that were dependent on IDH mutation status and attributable to changes in histological feature composition, somatic alterations, and microenvironment interactions. Hypermutation and acquired CDKN2A deletions associated with an increase in proliferating stem-like malignant cells at recurrence in both glioma subtypes, reflecting active tumor growth. IDH-wild-type tumors were more invasive at recurrence, and their malignant cells exhibited increased expression of neuronal signaling programs that reflected a possible role for neuronal interactions in promoting glioma progression. Mesenchymal transition was associated with the presence of a specific myeloid cell state defined by unique ligand-receptor interactions with malignant cells. Collectively, our results uncover recurrence-associated changes that could be targetable to shape disease progression following initial diagnosis.

cancer biology↗

bioRxiv2020

A drug-screening platform based on organotypic cultures identifies vulnerabilities to prevent local relapse and treat established brain metastasis

Exclusion of brain metastases from clinical trials is a major cause of the limited therapeutic options for this growing population of cancer patients. Here, we report a medium-throughput drug-screening platform (METPlatform) based on organotypic cultures that allows to evaluate inhibitors against metastases growing in situ. By applying this approach to brain metastasis, we identified several hits from a library of FDA approved inhibitors and others being tested in clinical trials. A blood-brain barrier permeable HSP90 inhibitor showed high potency against mouse and human brain metastases at clinically relevant stages of the disease, including a novel model of local relapse after neurosurgery. Furthermore, in situ proteomic analysis applied to organotypic cultures with metastases treated with the chaperone inhibitor revealed novel biomarkers in human brain metastasis and actionable mechanisms of resistance. Our work validates METPlatform as a potent resource for metastasis research integrating drug-screening and unbiased omic approaches that is fully compatible with human samples. We envision that METPlatform could be established as a clinically relevant strategy to personalize the management of metastatic disease in the brain and elsewhere. SummarySystemic spread of cancer continues to be the key aspect associated with lethality. In this publication, Zhu et al. describes a drug-screening platform specifically designed to study vulnerabilities of metastasis when colonizing secondary organs and demonstrates its value in difficult-to-treat brain metastasis using new models and patient-derived samples.

cancer biology↗

bioRxiv2020

A Human Aggression Pheromone Elicits a Sexually Dimorphic Behavioral and Brain Response

Body-volatiles can effectively trigger or block conspecific aggression in terrestrial mammals. Here we tested whether hexadecanal (HEX), a human body-volatile implicated as a mammalian-wide social cue, impacts human aggression. Using validated behavioural paradigms, we observed a remarkable dissociation: sniffing HEX blocked aggression in men, but triggered aggression in women. Next, using functional brain imaging, we uncovered a pattern of brain activity mirroring behaviour: In both men and women, HEX increased activity in the left angular gyrus, an area implicated in perception of social cues. Hex then modulated functional connectivity between the angular gyrus and a brain network implicated in social appraisal (temporal pole) and aggressive execution (amygdala and orbitofrontal cortex) in a sex-dependent manner consistent with behaviour: increasing connectivity in men, but decreasing connectivity in women. These findings implicate sex-specific social chemosignaling at the mechanistic heart of human aggressive behaviour.

neuroscience↗

bioRxiv2020

Lineage and Spatial Mapping of Glioblastoma-associated Immunity

The diversity of molecular states and cellular plasticity of immune cells in the glioblastoma environment is still poorly understood. Here, we performed scRNA sequencing of the immune compartment and mapped potential cellular interactions leading to an immunosuppressive microenvironment and dysfunction of T cells. Through inferring the dynamic adaptation during T cell activation, we identified three different terminal states with unique transcriptional programs. Modeling of driver genes for terminal T cell fate identified IL-10 signaling alterations in a subpopulation of HAVCR2(+) T cells. To explore in depth cellular interactions, we established an in-silico model by the integration of spatial transcriptomic and scRNA-sequencing, and identified a subset of HMOX1+ myeloid cells defined by IL10 release leading to T cell exhaustion. We found a spatial overlap between HMOX(+) myeloid and HAVCR2(+) T cells, suggesting that myeloid-lymphoid interaction causes immunosuppression present in tumor regions with enriched mesenchymal gene expression. Using human neocortical GBM model, coupled with patient-derived T cells, we confirmed that the functional interaction between myeloid and lymphoid cells, leads to a dysfunctional state of T cells. This IL-10 driven T cell exhaustion was found to be rescued by JAK/STAT inhibition. A comprehensive understanding of the cellular states and plasticity of lymphoid cells in GBM will aid towards successful immunotherapeutic approaches.

cancer biology↗