Research Article
Abstract
Lymphatics maintain fluid homeostasis, immune surveillance, and tissue integrity. Here, we identified the E26 transformation-specific transcription factors Erg and Fli1 as essential cooperative regulators of lymphatic integrity and function. Using inducible lymphatic endothelial cell–specific deletion in mice, we demonstrated that combined loss of Erg and Fli1 in adults results in fatal lymphatic failure, including chylothorax, chylous ascites, and impaired lymphatic drainage. Single-cell transcriptomic analysis revealed that loss of Erg and Fli1 causes disrupted lymphatic heterogeneity and dysregulation of key lymphatic genes, including valve-specific gene profiles. Erg and Fli1 coordinated lymphatic-immune crosstalk by transcriptionally regulating C-C motif chemokine ligand 21, which mediates DC trafficking. Erg or Fli1 loss also induced proinflammatory and prothrombotic gene expression, further contributing to lymphatic dysfunction. During embryonic development, the codeletion led to lymphatic mispatterning and loss of valve-initiating lymphatic endothelial cell clusters. The impact of loss of Erg and Fli1 function on lymphatic development in mice is consistent with FOXC2 mutations in lymphedema-distichiasis syndrome or ERG gene variants underlying primary lymphedema in humans. Moreover, Erg and Fli1 were required for regenerative lymphangiogenesis and lymphatic repair following injury in adults. Our findings establish Erg and Fli1 as core transcriptional regulators of lymphatic identity, integrity, and function.
Authors
Myung Jin Yang, Seok Kang, Seon Pyo Hong, Hokyung Jin, Jin-Hui Yoon, Cheolhwa Jin, Chae Min Yuk, Lydia Getachew Gebeyehu, Junho Jung, Sung-Hwan Yoon, Hyuek Jong Lee, Gou Young Koh
Abstract
Atopic dermatitis (AD) is a chronic inflammatory skin condition characterized by a type 2 immune response that is not fully understood. Single-cell RNA-seq of human AD skin and murine models of type 2 inflammation identified transcriptionally distinct fibroblast clusters, revealing IL-4Rα–dependent populations of immune-acting fibroblasts (IAFs). These unbiased findings prompted further investigation into the role of dermal fibroblasts during allergic inflammation. These studies demonstrated that, in an inflammatory environment including TNF-α, IL-1β, and IL-17A, the cytokines IL-4 and IL-13 stimulated both mouse and human fibroblasts to produce multiple chemokines, including CCL8, which activated CCR3 to attract T cells. In the skin, fibroblasts were the primary source of many of these chemokines, and targeted deletion of IL-4Rα in mouse fibroblasts reduced T cell infiltration in a mouse model of AD. Additionally, pharmacologic inhibition of CCR3, the receptor shared by many chemokines produced by fibroblasts, decreased T cell infiltration and skin inflammation in mouse models of AD. These findings demonstrate that dermal fibroblasts are more than passive structural cells; they actively participate in the type 2 immune response and contribute to AD by producing chemokines that increase inflammation. Targeting the functions of IAFs could offer an alternative therapeutic approach for AD.
Authors
Tomofumi Numata, Michael Shia, Yoshiyuki Nakamura, Fengwu Li, Hung Chan, Teruaki Nakatsuji, Kellen J. Cavagnero, Jared Simmons, Henry Li, Aaroh Anand Joshi, Marta Palomo-Irigoyen, Richard L. Gallo
Abstract
Large-cohort GWAS for alcohol use disorder (AUD) drug treatment outcomes and AUD risk have repeatedly identified genetic loci that are splicing quantitative trait loci for the fibronectin III domain containing 4 (FNDC4) gene in the brain. However, FNDC4 function in the brain and how it might contribute to AUD pathophysiology remain unclear. In the present study, we characterized GWAS loci–associated FNDC4 splice isoforms and demonstrated that FNDC4 alternative splicing results in loss of function for FNDC4. We also investigated FNDC4 function using CRISPR/Cas9 editing and the creation of human induced pluripotent stem cell–derived (iPSC-derived) neural organoids joined with single-nucleus RNA sequencing, a series of studies that showed that FNDC4 KO resulted in a striking shift in the relative proportions of glutamatergic and GABAergic neurons in iPSC-derived forebrain organoids as well as changes in their electrical activity. We further explored a potential mechanism(s) of FNDC4-dependent neurogenesis, and the results suggested a role for FNDC4 in mediating neural cell surface interactions. In summary, this series of experiments indicates that FNDC4 plays a role in regulating cerebral cortical neurogenesis in the brain. This regulation may contribute to the response to AUD pharmacotherapy as well as the effects of alcohol on the brain.
Authors
Xiujuan Zhu, August J. John, Sooan Kim, Li Wang, Enci Ding, Jing Zheng, Ateka Saleh, Irene Marín-Goñi, Abedalrahman Jomaa, Huanyao Gao, Meijie Wang, Ching Man Wai, Irene Moon, Cindy Chen, Alireza Agahi, Brandon J. Coombes, Tony M. Kerr, Nobuyoshi Suto, Liewei Wang, Mark A. Frye, Joanna M. Biernacka, Victor M. Karpyak, Hu Li, Richard M. Weinshilboum, Duan Liu
Abstract
Glioblastomas (GBMs) are highly lethal brain tumors with limited treatment options and resistance to immune checkpoint inhibitors due to their immunosuppressive tumor microenvironment. Here, we identify OLIG2 as a key regulator of immune evasion in GBM stem-like cells, which inhibits CD8+ T cell–dependent antitumor immunity while promoting protumor macrophage polarization. Mechanistically, OLIG2 recruited HDAC7 to repress CXCL10 transcription, inducing STAT3 activation in tumor-associated macrophages (TAMs) and decreasing CD8+ T cell infiltration and activation. Genetic deletion of OLIG2 significantly increased CXCL10 secretion, shifting TAMs toward an antitumor phenotype and enhancing CD8+ T cell activities. Furthermore, upregulated OLIG2 expression was correlated with resistance to immune checkpoint inhibitors in patients with GBMs. OLIG2 inhibition by either genetic deficiency or pharmacological targeting with CT-179 sensitized GBM tumors to anti–PD-L1 therapy, enhancing antitumor immune responses and prolonging survival. Our findings reveal OLIG2+ glioma stem-like cells as critical mediators of immune evasion and identify the OLIG2/HDAC7/CXCL10 axis as a potential therapeutic target to enhance immune checkpoint inhibitor efficacy and improve immunotherapy outcomes in aggressive GBMs.
Authors
Xinchun Zhang, Jinjiang Xue, Cunyan Zhao, Chenqiuyue Zeng, Jiacheng Zhong, Gangfeng Yu, Xi Yang, Yao Ling, Dazhen Li, Jiaxiao Yang, Yun Xiu, Hongda Li, Shiyuan Hong, Liangjun Qiao, Song Chen, Q. Richard Lu, Yaqi Deng, Zhaohua Tang, Fanghui Lu
Abstract
Epidermal growth factor receptor–activating (EGFR-activating) mutations are established biomarkers of resistance to immune checkpoint blockade (ICB) in lung cancer, yet the precise molecular mechanism and effective therapeutic strategies remain elusive. In this study, we show that EGFR overexpression and amplification recapitulated the negative effect of EGFR driver mutations on the ICB response, indicating a proactive involvement of EGFR signaling in antagonizing the antitumor immune response. Functional studies unveiled that EGFR activation suppressed the cellular response to IFN-γ following ICB treatment across multiple cancer models. This impairment in IFN-γ responsiveness further limited the upregulation of T cell–recruiting chemokines and antigen presentation, resulting in reduced T cell infiltration and activation, ultimately undermining antitumor immunity. Mechanistically, EGFR promotes Src homology 2-containing protein tyrosine phosphatase 2 (SHP2) activation to accelerate STAT1 dephosphorylation, leading to premature termination of the IFN-γ response. SHP2 inhibition restored ICB sensitivity in EGFR-activated tumors, significantly reducing tumor burden while maintaining a favorable safety profile. Our findings suggest that the EGFR/SHP2 axis functions as a molecular brake to disrupt the initiation and amplification of the IFN-γ–mediated antitumor response during immunotherapy. This discovery unveils a potential avenue to overcome immunotherapy resistance in EGFR-driven tumors, particularly lung cancer, through SHP2-targeted combination strategies.
Authors
Wei-Tao Zhuang, Lan-Lan Pang, Li-Yang Hu, Jun Liao, Jian-Hua Zhan, Ting Li, Ri-Xin Chen, Jia-Ni Zheng, An-Lin Li, Wen-Yan Yu, Tian-Qin Mao, Liang Chen, Yu-Jian Huang, Shao-Dong Hong, Jing Li, Jun-Han Wu, Yi-Ming Zeng, Meng-Juan Yang, Hai-Qing Zeng, Ya-Xiong Zhang, Li Zhang, Wen-Feng Fang
Abstract
Drug-associated environmental cues can trigger drug-seeking behavior and precipitate relapse. In this study, we determined that the claustrum (CL) connects the ventral tegmental area (VTA) with the medial prefrontal cortex (mPFC), forming the VTA–CL–mPFC circuit. Using a methamphetamine (METH) conditioned place preference (CPP) model in male mice, we found that manipulating the VTA–CL–mPFC circuit or CL neuronal ensemble receiving projections from VTA and projecting to mPFC (VTA–CL–mPFC) could disrupt the retrieval of METH-paired context memory, resulting in the blockage of the acquisition of METH CPP in male mice. During the process, dopamine release and dopamine 1-like receptor–mediated activation of CL neurons were required for the retrieval of METH-induced reward memory in male mice. These findings reveal a midbrain–cortical circuit orchestrated by CL neurons that plays an essential role in the retrieval of drug-paired environmental cue memory.
Authors
Ziheng Zhao, Yuhong He, Yang Liu, Quying Feng, Hee Young Kim, Yu Fan, Xiaowei Guan
Abstract
Malignant tumors with TP53 mutations exhibit poor therapeutic outcomes and high recurrence rates. T cell receptor–based (TCR-based) T cell therapy shows great promise for targeting intracellular cancer neoantigens. However, the immunogenic potential of TP53 hotspot mutations remains poorly characterized. Here, we identified an immunogenic neoantigen derived from the recurrent TP53R248Q mutation, presented by the prevalent HLA-A*11:01 allele. Additionally, we isolated a TP53R248Q-reactive TCR that specifically recognized the TP53R248Q mutation without any discernible cross-activity with cognate WT TP53 or other TP53 mutants at the same codon position. Functional characterization revealed that TP53R248Q TCR-T cells exhibited selective cytotoxicity against tumor cells expressing both the TP53R248Q mutation and HLA-A*11:01 in vitro. Importantly, the adoptive transfer of TP53R248Q TCR-T cells exhibited significant antitumor activity in a clinically relevant patient-derived xenograft model engrafted with TP53R248Q/HLA-A*11:01–positive human tumor tissues. Collectively, our study validates the immunogenicity of the TP53R248Q hotspot mutation and provides a TCR with high therapeutic potential for the development of T cell therapies targeting TP53R248Q/HLA-A*11:01–positive cancers.
Authors
Lianghua Shen, Ziyu Chen, Jian Xu, Qiaomei He, Changmeng Zhang, Xiao Zhou, Xiaodan Ding, Jinan Fang, Fanlin Li, Ming Jiao, Yuqin Yang, Baoxia Dong, Liping Wan, Xueying Ding, Yan Zheng, Jingyi Zhou, Chijian Zuo, Tian Min, Ming Zhu, Bin Ma, Yuhua Wan, Qiufang Guo, Hua Zhang, Jian Hua, Pengran Wang, Qi Li, Jiang Long, Xianmin Song, Yan Zhang
Abstract
MRE11, a breast tumor suppressor and component of the MRE11-RAD50-NBS1 complex, plays a critical role in DNA end resection and initiation of ataxia-telangiectasia mutation–dependent (ATM-dependent) DNA damage signaling. However, the precise mechanisms governing MRE11 function in the DNA damage response (DDR) remain incompletely understood. Here, we found that MRE11 is deacetylated by the SIRT2 sirtuin deacetylase and breast tumor suppressor, which promotes DNA binding to facilitate DNA end resection and ATM-dependent signaling. SIRT2 deacetylase activity promoted DNA end resection. SIRT2 further complexed with and deacetylated MRE11 at conserved lysine 393 (K393) in response to DNA double-stranded breaks (DSBs), which promoted MRE11 localization and DNA binding at DSBs but not interaction with RAD50, NBS1, or CtIP. Moreover, MRE11 K393 deacetylation by SIRT2 promoted ATM-dependent signaling. Our findings define a mechanism regulating MRE11 binding to DNA through SIRT2 deacetylation, elucidating a critical upstream signaling event directing MRE11 function in the DDR and providing insight into how SIRT2 dysregulation leads to genomic instability and tumorigenesis.
Authors
Fatmata Sesay, Hui Zhang, Priya Kapoor-Vazirani, Andrew T. Jung, Mark E. Essien, Amanda J. Bastien, Nho C. Luong, Xu Liu, PamelaSara E. Head, Duc M. Duong, Xiaofeng Yang, Zachary S. Buchwald, Xingming Deng, Nicholas T. Seyfried, David S. Yu
Abstract
While current antivirals primarily target viral proteins, host-directed strategies remain underexplored. Here, we performed a genome-wide CRISPR inhibition (CRISPRi) screening to identify the host protein, hepatocyte growth factor-regulated tyrosine kinase substrate (HGS), facilitating the pan-coronavirus infection both in vitro and in vivo. Mechanistically, HGS interacts with the viral membrane (M) protein, facilitating its trafficking to the ER-Golgi intermediate compartment for virion assembly. Conversely, HGS deficiency caused M retention in the ER, blocking assembly. Leveraging this interaction, we designed M-derived peptides and screened over 5,000 FDA-approved or commonly used drugs, identifying riboflavin tetrabutyrate (RTB). Both the peptides and RTB bind HGS and disrupt its interaction with the M protein, leading to M retention in the ER and subsequent blockade of virion assembly. These agents demonstrated broad anti-pan-coronavirus activity in vitro and in vivo. Collectively, our findings establish HGS as a druggable host target and identify RTB as a promising broad-spectrum antiviral candidate.
Authors
Xubing Long, Rongrong Chen, Rong Bai, Buyun Tian, Yu Cao, Kangying Chen, Fuyu Li, Yiliang Wang, Yongjie Tang, Qi Yang, Liping Ma, Fan Wang, Maoge Zhou, Xianjie Qiu, Yongzhi Lu, Jie Zheng, Peng Zhou, Xinwen Chen, Qian Liu, Xuepeng Wei, Yongxia Shi, Yanhong Xue, Jincun Zhao, Wei Ji, Liqiao Hu, Jinsai Shang, Tao Xu, Zonghong Li
Abstract
Warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome is an immunodeficiency caused by autosomal dominant hyperfunctional mutations in chemokine receptor CXCR4 that promote panleukopenia due to BM retention. We previously reported a preclinical gene therapy protocol involving allele-nonspecific Cxcr4 CRISPR/Cas9 inactivation, leveraging the known in vivo dominance of Cxcr4+/o (+, WT; o, inactivated) hematopoietic stem cells (HSCs) for autologous BM engraftment and leukocyte reconstitution over HSCs with other Cxcr4 genotypes. Here, we show that without BM conditioning, this approach is not able to correct leukopenia in WHIM mice. We therefore modified the protocol by adding conditioning with a nongenotoxic CD117-targeted immunotoxin, CD117-antibody-saporin-conjugate. With this change, donor-derived blood cells rapidly reached ~95% chimerism after transplantation, which was stable without adverse events. Mice receiving edited HSCs showed rapid normalization of absolute myeloid cell counts, the key blood subset responsible for WHIM syndrome. In competitive transplants using equal numbers of edited and unedited donor HSCs, over 80% of blood cells originated from the edited population, predominantly with the Cxcr4+/o genotype. These results provide proof of principle that CRISPR/Cas9-mediated inactivation of the Cxcr4 disease allele, combined with nongenotoxic HSC-targeted conditioning, may offer a safe and effective gene therapy strategy generalizable to all WHIM-causing mutations.
Authors
Ji-Liang Gao, Zhanzhuo Li, Rafael Calderon-Perez, Antonia Pavek, Lina Kim, David H. McDermott, Philip M. Murphy
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Copyright © 2026 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)