Research
Abstract
GATA6 is a master regulator of differentiation in the pancreas and its expression levels determine the two main molecular subtypes of pancreatic cancer. High GATA6 contributes to the “classical” pancreatic cancer subtype, which is associated with a higher degree of tumor differentiation and better disease prognosis. However, why GATA6 expression varies across pancreatic cancers and what regulate GATA6 expression remain elusive. Here we report that the oncogenic KRAS-activated ERK signaling suppresses GATA6 transcription in pancreatic cancers. GATA6 mRNA levels inversely correlated with KRAS/ERK activity in pancreatic tumors. A genome-wide CRISPR screen in a GATA6-EGFP reporter knockin cell line identified JUNB as the ERK-regulated transcriptional repressor for GATA6. Active ERK stabilizes JUNB protein while KRAS/ERK inhibition led to ubiquitin-independent proteasomal degradation of JUNB and increased transcription of GATA6. Up-regulation of GATA6 enhanced chemosensitivity of pancreatic cancer cells and KRAS/ERK inhibitors synergized with chemotherapy in a GATA6-dependent manner. Our study identifies how oncogenic KRAS/ERK signaling suppresses GATA6 to cause dedifferentiation in pancreatic cancer. Combining KRAS/ERK inhibitors with standard-of-care chemotherapies could be a promising therapeutic strategy for treating pancreatic cancers.
Authors
Zheng Zhong, Xinang Cao, Pei-Ju Liao, Raman Sethi, Jeffrey A. Klomp, Clint A. Stalnecker, Jinmiao Chen, Yue Wan, Channing J. Der, David M. Virshup
Abstract
During vascular injury, platelets are essential for halting bleeding and recruiting neutrophils to prevent microbial invasion. However, in antibody-mediated autoimmune diseases occurring without vascular damage, neutrophils infiltrate tissues and contribute to pathology. Here, we investigated whether the dependence of neutrophils on platelets is conserved in the context of antibody-driven inflammation. Using human cells from individuals with rheumatoid arthritis and a microfluidic system mimicking physiological shear over IgG-containing immune complexes, we demonstrate that despite expressing Fc receptors, neutrophils require platelets to stably adhere to immune complexes under flow. Platelet FcγRIIA binding was critical for resisting shear stress, while neutrophils used FcγRIIA and FcγRIIIB for immune complex recognition. Platelet P-selectin binding to neutrophil PSGL-1 was essential for recruitment, whereas Mac-1 was dispensable. In a mouse model of autoantibody-mediated arthritis, intravital imaging confirmed that neutrophil recruitment critically relies on PSGL-1. Importantly, expression of FcyRIIA aggravated arthritis, and blockade of PSGL-1 in these mice, but not of Mac-1, abrogated both the platelet and neutrophil interactions and disease. These findings identify key molecular interactions in platelet–neutrophil cooperation and reveal that platelets are essential enablers of FcR-mediated neutrophil adhesion in antibody-driven inflammation.
Authors
Marie Bellio, Isabelle Allaeys, Etienne Doré, Myriam Vaillancourt, Tania Lévesque, Mélina Monteil, Nicolas Vallières, Philippe Desaulniers, Nicolas Bertrand, Valance A. Washington, Yotis Senis, Steve Lacroix, Paul Fortin, Clémence Belleannée, Eric Boilard
Abstract
The role of CARD9 in the pathogenesis of various chronic fungal infections has been established; however, the precise mechanisms underlying the pathobiology of these infections remain unclear. We aimed to investigate the specific cellular mechanisms by which CARD9 deficiency contributes to the pathogenesis of chronic fungal infections. Using single-cell RNA sequencing (scRNA-seq), we analyzed the immune cell profiles in skin lesions from both murine and human samples. We focused on macrophage differentiation and signaling pathways influenced by CARD9 deficiency. We found that CARD9 deficiency promotes the differentiation of TREM2high macrophages following fungal stimulation, impairing their antifungal functions and inducing exhaustion-like T helper 1 (Th1) cells. Mechanistically, the NF-κB pathway activation was restricted in CARD9-deficient macrophages, leading to enhanced CREB activation, which in turn exerted a positive regulatory effect on Trem2 expression by activating C/EBPβ. Notably, targeting TREM2 enhanced the antifungal immune response in vivo and in vitro, thereby alleviating the severity of CARD9-deficient subcutaneous dematiaceous fungal infection. Our findings highlight the important role of CARD9 in regulating cutaneous antifungal immunity and identify potential targets for immunotherapy in chronic dematiaceous fungal infections.
Authors
Lu Zhang, Zhichun Tang, Yi Zhang, Wenjie Liu, Haitao Jiang, Li Yu, Kexin Lei, Yubo Ma, Yang-xin Fu, Ruoyu Li, Wenyan Wang, Fan Bai, Xiaowen Wang
Abstract
Emerging evidence demonstrates that chronic stress alters immunological, neurochemical and endocrinological functions, thereby promoting tumor progression. However, the underlying metabolic mechanism of chronic stress in tumor progression is still elusive. Using multi-omics analysis, we found that aminopeptidase N (ANPEP) was upregulated in tumors with chronic restraint, associating with the reprogramming of amino acid metabolism. Functional assays revealed that ANPEP promoted liver cancer growth and metastasis. Knockdown of ANPEP blocked chronic stress-induced liver cancer progression. Chronic stress-induced glucocorticoids promoted nuclear receptor subfamily 3 group C member 1 (NR3C1) nuclear translocation to activate ANPEP transcription by directly binding to its promoter. Furthermore, ANPEP promotes glutathione synthesis, subsequently inhibiting reactive oxygen species (ROS)-induced ferroptosis. Mechanistically, ANPEP interacted with solute carrier family 3 member 2 (SLC3A2) to block membrane associated ring-CH-type finger 8-mediated (MARCH8-mediated) lysosome-dependent degradation of SLC3A2, promoting intracellular L-cystine transport, thereby increasing glutathione synthesis. The combination of ANPEP silencing and sorafenib treatment showed a synergistic effect in inhibiting liver cancer progression. Finally, clinical data and mouse models demonstrated that chronic stress drove liver tumor progression via ANPEP-regulated SLC3A2. These findings reveal unanticipated communication between chronic stress and metabolic reprogramming during liver cancer progression, providing potential therapeutic implications for liver cancer.
Authors
Yongkang Wu, Yankun Zhang, Xiaojia Shi, Mengting Wu, Min Sun, Ying Feng, Wenmeng Ma, Xiule Jiang, Dingqi Fei, Mingjian Zhao, Zhuanchang Wu, Chunyang Li, Xiaohong Liang, Lifen Gao, Chunhong Ma, Xuetian Yue
Abstract
Epstein-Barr virus (EBV) is of growing interest for its potential role in neurodegenerative diseases such as multiple sclerosis (MS) and its possible utility as a therapeutic target in herpesvirus-associated chronic diseases. The effects of brincidofovir (BCV) on EBV reactivation were evaluated in vitro using EBV-infected spontaneous lymphoblastoid cell lines (SLCLs) and peripheral blood mononuclear cells (PBMCs) derived from MS patients and healthy controls. In addition, a B lymphoblastoid cell line and PBMCs from common marmosets (Callithrix jacchus) naturally infected with an EBV-related gammaherpesvirus (Callitrichine herpesvirus 3, CalHV-3) were used to measure BCV efficacy in a nonhuman primate model. BCV significantly inhibited gammaherpesvirus reactivation, with decreased lytic and latent viral transcript expression. These results suggest that BCV may be a useful antiviral for inhibiting EBV activity in MS patients. Additionally, this work further validates the utility of CalHV-3 in marmosets as a translational model for the investigation of successful EBV-targeting therapeutics.
Authors
Abaigeal Donaldson, Madeleine R. Druker, Maria Chiara Monaco, Emily H. Stack, Paige Zimmerman, Amanda Lee, Izabela Bialuk, William Frazier, Irene Cortese, Heather Narver, Masatoshi Hazama, Fuminori Yoshida, Xiaofan Li, Laurie T. Krug, Stacey L. Piotrowski, Steven Jacobson
Abstract
Acute myeloid leukemia (AML) is an aggressive cancer with very poor outcomes. To identify additional drivers of leukemogenesis, we analyzed sequencing data from 1,727 unique individual AML patients, which revealed mutations in ubiquitin ligase family genes in 11.2% of adult AML samples with mutual exclusivity. The SKP1/CUL1/F-box (SCF) E3 ubiquitin ligase complex gene, FBXO11, was the most significantly downregulated gene of the SCF complex in AML. We found that FBXO11 interacts with and catalyzes K63-linked ubiquitination of LONP1 in the cytosol, to promote LONP1 entry into mitochondria. We show that depletion of FBXO11 or LONP1 reduces mitochondrial respiration through impaired LONP1 chaperone activity to assemble electron transport chain Complex IV. Reduced mitochondrial respiration secondary to FBXO11 or LONP1 depletion imparted myeloid-biased stem cell properties in primary CD34+ hematopoietic stem and progenitor cells (HSPC) in vitro. In a human xenograft model, depletion of FBXO11 cooperated with AML1-ETO and mutant KRASG12D to generate serially transplantable AML. Our findings suggest that reduced FBXO11 cooperates to initiate AML by priming HSPC for myeloid-biased self-renewal through attenuation of LONP1-mediated regulation of mitochondrial respiration.
Authors
Hayle Kincross, Ya-Chi Angela Mo, Xuan Wang, Linda Chang, Gerben Duns, Franziska Mey, Jihong Jiang, Zurui Zhu, Naomi Isak, Harwood Kwan, Tammy T.Y. Lau, T. Roderick Docking, Pranav Garg, Jessica Tran, Shane Colborne, Se-Wing Grace Cheng, Shujun Huang, Nadia Gharaee, Elijah Willie, Jeremy D.K. Parker, Joshua Bridgers, Davis Wood, Ramon I. Klein Geltink, Gregg B. Morin, Aly Karsan
Abstract
Disorders of GABRA3, the only epilepsy-associated GABA-A receptor subunit gene on the X chromosome, have eluded clinical clarity due to ambiguous inheritance patterns and variable phenotypes. The long-standing assumption that all pathogenic variants cause loss-of-function further obscured genotype-phenotype relationships and hindered progress. Here, we curated a cohort of individuals with a GABRA3 variant, integrating deep phenotyping, genotyping, family history, electrophysiology, with a targeted mouse model. Among 43 individuals with 19 GABRA3 variants, functional analyses revealed both gain- and loss-of-function effects, each linked to distinct clinical profiles. Gain-of-function variants were associated with severe, treatment-resistant epilepsy and severe-profound intellectual disability, disproportionately affecting males, who were often non-ambulant and had cortical visual impairment. Loss-of-function variants produced milder phenotypes, with epilepsy rarely observed; affected males showed behavioural issues and language delay, while females were unaffected carriers. Our gain-of-function (Gabra3Q242L/+) mouse model mirrored these sex-specific differences, showing increased seizure susceptibility, early death, and marked cortical hyperexcitability. These insights not only resolve longstanding uncertainties surrounding GABRA3 but also redefine how X-linked disorders are interpreted. They demonstrate that it is the functional impact of a variant, not its mere presence, that determines whether a condition manifests dominantly or recessively. This distinction carries important implications for genetic counselling, precision medicine, and the broader interpretation of X-linked neurodevelopmental disorders.
Authors
Katrine M. Johannesen, Khaing Phyu Aung, Vivian W.Y. Liao, Nathan L. Absalom, Han C. Chua, Xue N. Gan, Miaomiao Mao, Chaseley E. McKenzie, Hian M. Lee, Sebastian Ortiz, Rebecca C. Spillmann, Vandana Shashi, Rodney A. Radtke, Ghayda M. Mirzaa, P. Anne Weisner, Josue Flores Daboub, Caroline Hagedorn, Pinar Bayrak-Toydemir, Desiree DeMille, Jian Zhao, Nandita Bajaj, Yline Capri, Boris Keren, Miriam Schmidts, Ingrid M.B.H. van de Laar, Marjon A. van Slegtenhorst, Rafal Ploski, Marta Bogotko, Danielle K. Bourque, Ebba Alkhunaizi, Lauren Chad, Nada Quercia, Houda Elloumi, Ingrid M. Wentzensen, Michael C. Kruer, Pritha Bisarad, Carolina I. Galaz-Montoya, Violeta Rusu, Dominique Braun, Katie Angione, Jessica C. Win, Camilo Espinosa-Jovel, Pia Zacher, Konrad Platzer, Samuel F. Berkovic, Ingrid E. Scheffer, Mary Chebib, Guido Rubboli, Rikke S. Møller, Christopher A. Reid, Philip K. Ahring
Abstract
Hormone Receptor positive (HR+) breast cancers (BC) are typically “immune-cold” poorly immune infiltrated tumors that do not respond to immune-checkpoint blockade (ICB) therapies. Using clinical data, we report that estrogen receptor (ERα) signaling associates with immunosuppressive pathways and lack of response to ICB in HR+ patients. In this study, we validate ER-mediated immunosuppression by engineering and modulating ER in preclinical models in vitro, in vivo and ex vivo. Mechanistically, we found that ERα hijacks LCOR, a nuclear receptor corepressor, thereby preventing LCOR’s function in the induction of tumor immunogenicity and immune infiltration, which is normally observed in the absence of ERα, such as in ER-negative BC. In HR+BC, we demonstrate that the molecular disruption of LCOR and ERα interaction using anti-ER therapies or using a mutant of the LCOR nuclear-receptor binding domain (LSKLL into LSKAA) that does not interact with ERα, restores LCOR’s immunogenic functions. Remarkably, the LCOR-ERα disruption converts HR+BC immune-cold tumors into immune-hot tumors responsive to ICB by increased antigen presentation machinery (APM) expression, immune infiltration, T cell recognition and mediated killing. In conclusion, ERα inhibition and the disruption of LCOR to ERα represent a novel therapeutic strategy and an opportunity to elicit immunotherapeutic benefit in HR+BC patients.
Authors
José Ángel Palomeque, Gabriel Serra-Mir, Sandra Blasco-Benito, Helena Brunel, Pau Torren-Duran, Iván Pérez-Núñez, Chiara Cannatá, Laura Comerma, Silvia Menendez, Sonia Servitja, Tamara Martos, Maria Castro, Rodrigo L. Borges, Joanna I. Lopez-Velazco, Sara Manzano, Santiago Duro-Sánchez, Joaquin Arribas, Maria M. Caffarel, Ander Urruticoechea, Jose A. Seoane, Lluis Morey, Joan Albanell, Toni Celià-Terrassa
Abstract
In pancreatic β-cells, misfolded proinsulin is a substrate for Endoplasmic Reticulum-Associated protein Degradation (ERAD) via HRD1/SEL1L. β-cell HRD1 activity is alternately reported to improve, or impair, insulin biogenesis. Further, while β-cell SEL1L deficiency causes HRD1 hypofunction and diminishes islet insulin content; reports conflict as to whether β-cell ERAD deficiency increases or decreases proinsulin levels. Here we’ve examined β-cell-specific Hrd1-KO mice (chronic deficiency), plus rodent (and human islet) β-cells treated acutely with HRD1 inhibitor. β-Hrd1-KO mice developed diabetes with decreased islet proinsulin yet a relative increase of misfolded proinsulin re-distributed to the ER; upregulated biochemical markers of β-cell ER stress and autophagy; electron microscopic evidence of ER enlargement and decreased insulin granule content; and increased glucagon-positive islet cells. Misfolded proinsulin was also increased in islets treated with inhibitors of lysosomal degradation. Preceding any loss of total proinsulin, acute HRD1 inhibition triggered increased nonnative proinsulin, increased phospho-eIF2ɑ with inhibited proinsulin synthesis, and increased LC3b-II (the abundance of which requires expression of SigmaR1). We posit a subset of proinsulin molecules undergoes HRD1-mediated disposal. When HRD1 is unavailable, misfolded proinsulin accumulates, accompanied by increased phospho-eIF2ɑ that limits further proinsulin synthesis, plus SigmaR1-dependent autophagy activation, ultimately lowering steady-state β-cell proinsulin (and insulin) levels — triggering diabetes.
Authors
Anoop Arunagiri, Leena Haataja, Maroof Alam, Noah F. Gleason, Emma Mastroianni, Chao-Yin Cheng, Sami Bazzi Onton, Jeffrey Knupp, Ibrahim Metawea, Anis Hassan, Dennis Larkin, Deyu Fang, Billy Tsai, Ling Qi, Peter Arvan
Abstract
While immune checkpoint blockade (ICB) therapy has revolutionized the antitumor therapeutic landscape, it remains successful in only a small subset of cancer patients. Poor or loss of MHC-I expression has been implicated as a common mechanism of ICB resistance. Yet the molecular mechanisms underlying impaired MHC-I remain to be fully elucidated. Herein, we identified USP22 as a critical factor responsible for ICB resistance through suppressing MHC-I-mediated neoantigen presentation to CD8 T cells. Both genetic and pharmacologic USP22 inhibition increased immunogenicity and overcome anti-PD-1 immunotherapeutic resistance. At the molecular level, USP22 functions as a deubiquitinase for the methyltransferase EZH2, leading to transcriptional silencing of MHC-I gene expression. Targeted Usp22 inhibition resulted in increased tumoral MHC-I expression and consequently enhanced CD8 T cell killing, which was largely abrogated by Ezh2 reconstitution. Multiplexed immunofluorescence staining detected a strong reverse correlation between USP22 expression and both 2M expression and CD8+ T lymphocyte infiltration in solid tumors. Importantly, USP22 upregulation was associated with ICB immunotherapeutic resistance in patients with lung cancer. Collectively, this study highlights the role of USP22 as a diagnostic biomarker for ICB resistance and provides a potential therapeutic avenue to overcome the current ICB resistance through inhibition of USP22.
Authors
Kun Liu, Radhika Iyer, Yi Li, Jun Zhu, Zhaomeng Cai, Juncheng Wei, Yang Cheng, Amy Tang, Hai Wang, Qiong Gao, Nikita Lavanya Mani, Noah Marx, Beixue Gao, D. Martin Watterson, Seema A. Khan, William J. Gradishar, Huiping Liu, Deyu Fang
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ISSN: 0021-9738 (print), 1558-8238 (online)