Research Article
Abstract
Estrogen receptor–negative (ER-negative) breast cancer is thought to be more malignant and devastating than ER-positive breast cancer. ER-negative breast cancer exhibits elevated NF-κB activity, but how this abnormally high NF-κB activity is maintained is poorly understood. The importance of linear ubiquitination, which is generated by the linear ubiquitin chain assembly complex (LUBAC), is increasingly appreciated in NF-κB signaling, which regulates cell activation and death. Here, we showed that epsin proteins, a family of ubiquitin-binding endocytic adaptors, interacted with LUBAC via its ubiquitin-interacting motif and bound LUBAC’s bona fide substrate NEMO via its N-terminal homolog (ENTH) domain. Furthermore, epsins promoted NF-κB essential modulator (NEMO) linear ubiquitination and served as scaffolds for recruiting other components of the IκB kinase (IKK) complex, resulting in the heightened IKK activation and sustained NF-κB signaling essential for the development of ER-negative breast cancer. Heightened epsin levels in ER-negative human breast cancer are associated with poor relapse-free survival. We showed that transgenic and pharmacological approaches eliminating epsins potently impeded breast cancer development in both spontaneous and patient-derived xenograft breast cancer mouse models. Our findings established the pivotal role epsins played in promoting breast cancer. Thus, targeting epsins may represent a strategy to restrain NF-κB signaling and provide an important perspective into ER-negative breast cancer treatment.
Authors
Kai Song, Xiaofeng Cai, Yunzhou Dong, Hao Wu, Yong Wei, Uma T. Shankavaram, Kui Cui, Yang Lee, Bo Zhu, Sudarshan Bhattacharjee, Beibei Wang, Kun Zhang, Aiyun Wen, Scott Wong, Lili Yu, Lijun Xia, Alana L. Welm, Diane R. Bielenberg, Kevin A. Camphausen, Yibin Kang, Hong Chen
Abstract
Inborn errors of TLR3-dependent IFN-α/β– and IFN-λ–mediated immunity in the CNS can underlie herpes simplex virus 1 (HSV-1) encephalitis (HSE). The respective contributions of IFN-α/β and IFN-λ are unknown. We report a child homozygous for a genomic deletion of the entire coding sequence and part of the 3′-UTR of the last exon of IFNAR1, who died of HSE at the age of 2 years. An older cousin died following vaccination against measles, mumps, and rubella at 12 months of age, and another 17-year-old cousin homozygous for the same variant has had other, less severe, viral illnesses. The encoded IFNAR1 protein is expressed on the cell surface but is truncated and cannot interact with the tyrosine kinase TYK2. The patient’s fibroblasts and EBV-B cells did not respond to IFN-α2b or IFN-β, in terms of STAT1, STAT2, and STAT3 phosphorylation or the genome-wide induction of IFN-stimulated genes. The patient’s fibroblasts were susceptible to viruses, including HSV-1, even in the presence of exogenous IFN-α2b or IFN-β. HSE is therefore a consequence of inherited complete IFNAR1 deficiency. This viral disease occurred in natural conditions, unlike those previously reported in other patients with IFNAR1 or IFNAR2 deficiency. This experiment of nature indicates that IFN-α/β are essential for anti–HSV-1 immunity in the CNS.
Authors
Paul Bastard, Jeremy Manry, Jie Chen, Jérémie Rosain, Yoann Seeleuthner, Omar AbuZaitun, Lazaro Lorenzo, Taushif Khan, Mary Hasek, Nicholas Hernandez, Benedetta Bigio, Peng Zhang, Romain Lévy, Shai Shrot, Eduardo J. Garcia Reino, Yoon-Seung Lee, Soraya Boucherit, Mélodie Aubart, Rik Gijsbers, Vivien Béziat, Zhi Li, Sandra Pellegrini, Flore Rozenberg, Nico Marr, Isabelle Meyts, Bertrand Boisson, Aurélie Cobat, Jacinta Bustamante, Qian Zhang, Emmanuelle Jouangy, Laurent Abel, Raz Somech, Jean-Laurent Casanova, Shen-Ying Zhang
Abstract
A growing number of long noncoding RNAs (lncRNAs) have emerged as vital metabolic regulators. However, most human lncRNAs are nonconserved and highly tissue specific, vastly limiting our ability to identify human lncRNA metabolic regulators (hLMRs). In this study, we established a pipeline to identify putative hLMRs that are metabolically sensitive, disease relevant, and population applicable. We first progressively processed multilevel human transcriptome data to select liver lncRNAs that exhibit highly dynamic expression in the general population, show differential expression in a nonalcoholic fatty liver disease (NAFLD) population, and respond to dietary intervention in a small NAFLD cohort. We then experimentally demonstrated the responsiveness of selected hepatic lncRNAs to defined metabolic milieus in a liver-specific humanized mouse model. Furthermore, by extracting a concise list of protein-coding genes that are persistently correlated with lncRNAs in general and NAFLD populations, we predicted the specific function for each hLMR. Using gain- and loss-of-function approaches in humanized mice as well as ectopic expression in conventional mice, we validated the regulatory role of one nonconserved hLMR in cholesterol metabolism by coordinating with an RNA-binding protein, PTBP1, to modulate the transcription of cholesterol synthesis genes. Our work overcame the heterogeneity intrinsic to human data to enable the efficient identification and functional definition of disease-relevant human lncRNAs in metabolic homeostasis.
Authors
Xiangbo Ruan, Ping Li, Yonghe Ma, Cheng-fei Jiang, Yi Chen, Yu Shi, Nikhil Gupta, Fayaz Seifuddin, Mehdi Pirooznia, Yasuyuki Ohnishi, Nao Yoneda, Megumi Nishiwaki, Gabrijela Dumbovic, John L. Rinn, Yuichiro Higuchi, Kenji Kawai, Hiroshi Suemizu, Haiming Cao
Abstract
Protection of the brain from viral infections involves the type I IFN (IFN-I) system, defects in which render humans susceptible to herpes simplex encephalitis (HSE). However, excessive cerebral IFN-I levels lead to pathologies, suggesting the need for tight regulation of responses. Based on data from mouse models, human HSE cases, and primary cell culture systems, we showed that microglia and other immune cells undergo apoptosis in the HSV-1–infected brain through a mechanism dependent on the cyclic GMP-AMP synthase/stimulator of interferon genes (cGAS/STING) pathway, but independent of IFN-I. HSV-1 infection of microglia induced cGAS-dependent apoptosis at high viral doses, whereas lower viral doses led to IFN-I responses. Importantly, inhibition of caspase activity prevented microglial cell death and augmented IFN-I responses. Accordingly, HSV-1–infected organotypic brain slices or mice treated with a caspase inhibitor exhibited lower viral load and an improved infection outcome. Collectively, we identify an activation-induced apoptosis program in brain immune cells that downmodulates local immune responses.
Authors
Line S. Reinert, Ahmad S. Rashidi, Diana N. Tran, Georgios Katzilieris-Petras, Astrid K. Hvidt, Mette Gohr, Stefanie Fruhwürth, Chiranjeevi Bodda, Martin K. Thomsen, Mikkel H. Vendelbo, Ahmad R. Khan, Brian Hansen, Petra Bergström, Lotta Agholme, Trine H. Mogensen, Maria H. Christensen, Jens R. Nyengaard, Ganes C. Sen, Henrik Zetterberg, Georges MGM Verjans, Søren R. Paludan
Abstract
Microglia maintain homeostasis in the brain. However, with age, they become primed and respond more strongly to inflammatory stimuli. We show here that microglia from aged mice had upregulated mTOR complex 1 signaling controlling translation, as well as protein levels of inflammatory mediators. Genetic ablation of mTOR signaling showed a dual yet contrasting effect on microglia priming: it caused an NF-κB–dependent upregulation of priming genes at the mRNA level; however, mice displayed reduced cytokine protein levels, diminished microglia activation, and milder sickness behavior. The effect on translation was dependent on reduced phosphorylation of 4EBP1, resulting in decreased binding of eIF4E to eIF4G. Similar changes were present in aged human microglia and in damage-associated microglia, indicating that upregulation of mTOR-dependent translation is an essential aspect of microglia priming in aging and neurodegeneration.
Authors
Lily Keane, Ignazio Antignano, Sean-Patrick Riechers, Raphael Zollinger, Anaelle A. Dumas, Nina Offermann, Maria E. Bernis, Jenny Russ, Frederike Graelmann, Patrick Neil McCormick, Julia Esser, Dario Tejera, Ai Nagano, Jun Wang, Claude Chelala, Yvonne Biederbick, Annett Halle, Paolo Salomoni, Michael T. Heneka, Melania Capasso
Abstract
Although platelets are the cellular mediators of thrombosis, they are also immune cells. Platelets interact both directly and indirectly with immune cells, impacting their activation and differentiation, as well as all phases of the immune response. Megakaryocytes (Mks) are the cell source of circulating platelets, and until recently Mks were typically only considered bone marrow–resident (BM-resident) cells. However, platelet-producing Mks also reside in the lung, and lung Mks express greater levels of immune molecules compared with BM Mks. We therefore sought to define the immune functions of lung Mks. Using single-cell RNA sequencing of BM and lung myeloid-enriched cells, we found that lung Mks, which we term MkL, had gene expression patterns that are similar to antigen-presenting cells. This was confirmed using imaging and conventional flow cytometry. The immune phenotype of Mks was plastic and driven by the tissue immune environment, as evidenced by BM Mks having an MkL-like phenotype under the influence of pathogen receptor challenge and lung-associated immune molecules, such as IL-33. Our in vitro and in vivo assays demonstrated that MkL internalized and processed both antigenic proteins and bacterial pathogens. Furthermore, MkL induced CD4+ T cell activation in an MHC II–dependent manner both in vitro and in vivo. These data indicated that MkL had key immune regulatory roles dictated in part by the tissue environment.
Authors
Daphne N. Pariser, Zachary T. Hilt, Sara K. Ture, Sara K. Blick-Nitko, Mark R. Looney, Simon J. Cleary, Estheany Roman-Pagan, Jerry Saunders II, Steve N. Georas, Janelle Veazey, Ferralita Madere, Laura Tesoro Santos, Allison Arne, Nguyen P.T. Huynh, Alison C. Livada, Selena M. Guerrero-Martin, Claire Lyons, Kelly A. Metcalf-Pate, Kathleen E. McGrath, James Palis, Craig N. Morrell
Abstract
Mutations in the core RNA splicing factor SF3B1 are prevalent in leukemias and uveal melanoma, but hotspot SF3B1 mutations are also seen in epithelial malignancies such as breast cancer. Although hotspot mutations in SF3B1 alter hematopoietic differentiation, whether SF3B1 mutations contribute to epithelial cancer development and progression is unknown. Here, we identify that SF3B1 mutations in mammary epithelial and breast cancer cells induce a recurrent pattern of aberrant splicing leading to activation of AKT and NF-κB, enhanced cell migration, and accelerated tumorigenesis. Transcriptomic analysis of human cancer specimens, MMTV-cre Sf3b1K700E/WT mice, and isogenic mutant cell lines identified hundreds of aberrant 3′ splice sites (3′ss) induced by mutant SF3B1. Consistently between mouse and human tumors, mutant SF3B1 promoted aberrant splicing (dependent on aberrant branchpoints as well as pyrimidines downstream of the cryptic 3′ss) and consequent suppression of PPP2R5A and MAP3K7, critical negative regulators of AKT and NF-κB. Coordinate activation of NF-κB and AKT signaling was observed in the knockin models, leading to accelerated cell migration and tumor development in combination with mutant PIK3CA but also hypersensitizing cells to AKT kinase inhibitors. These data identify hotspot mutations in SF3B1 as an important contributor to breast tumorigenesis and reveal unique vulnerabilities in cancers harboring them.
Authors
Bo Liu, Zhaoqi Liu, Sisi Chen, Michelle Ki, Caroline Erickson, Jorge S. Reis-Filho, Benjamin H. Durham, Qing Chang, Elisa de Stanchina, Yiwei Sun, Raul Rabadan, Omar Abdel-Wahab, Sarat Chandarlapaty
Abstract
Dysfunction of immune and vascular systems has been implicated in aging and Alzheimer disease; however, their interrelatedness remains poorly understood. The complement pathway is a well-established regulator of innate immunity in the brain. Here, we report robust age-dependent increases in vascular inflammation, peripheral lymphocyte infiltration, and blood-brain barrier (BBB) permeability. These phenotypes were subdued by global inactivation and by endothelial cell–specific ablation of C3ar1. Using an in vitro model of the BBB, we identified intracellular Ca2+ as a downstream effector of C3a/C3aR signaling and a functional mediator of vascular endothelial cadherin junction and barrier integrity. Endothelial C3ar1 inactivation also dampened microglia reactivity and improved hippocampal and cortical volumes in the aging brain, demonstrating a crosstalk between brain vasculature dysfunction and immune cell activation and neurodegeneration. Further, prominent C3aR-dependent vascular inflammation was also observed in a tau-transgenic mouse model. Our studies suggest that heightened C3a/C3aR signaling through endothelial cells promotes vascular inflammation and BBB dysfunction and contributes to overall neuroinflammation in aging and neurodegenerative disease.
Authors
Nicholas E. Propson, Ethan R. Roy, Alexandra Litvinchuk, Jörg Köhl, Hui Zheng
Abstract
Therapeutic strategies designed to target TP53-deficient cancer cells remain elusive. Here, we showed that TP53 loss initiated a pharmacologically actionable secretory process that drove lung adenocarcinoma (LUAD) progression. Molecular, biochemical, and cell biological studies showed that TP53 loss increased the expression of Golgi reassembly and stacking protein 55 kDa (G55), a Golgi stacking protein that maintains Golgi organelle integrity and is part of a GOLGIN45 (G45)–myosin IIA–containing protein complex that activates secretory vesicle biogenesis in the Golgi. TP53 loss activated G55-dependent secretion by relieving G55 and myosin IIA from miR-34a–dependent silencing. G55-dependent secreted proteins enhanced the proliferative and invasive activities of TP53-deficient LUAD cells and promoted angiogenesis and CD8+ T cell exhaustion in the tumor microenvironment. A small molecule that blocks G55-G45 interactions impaired secretion and reduced TP53-deficient LUAD growth and metastasis. These results identified a targetable secretory vulnerability in TP53-deficient LUAD cells.
Authors
Xiaochao Tan, Lei Shi, Priyam Banerjee, Xin Liu, Hou-Fu Guo, Jiang Yu, Neus Bota-Rabassedas, B. Leticia Rodriguez, Don L. Gibbons, William K. Russell, Chad J. Creighton, Jonathan M. Kurie
Abstract
Oligodendrocytes express low-density lipoprotein receptor (LDLR) to endocytose cholesterol for the maintenance of adulthood myelination. However, the potential role of LDLR in chronic cerebral ischemia–related demyelination remains unclear. We used bilateral carotid artery stenosis (BCAS) to induce sustained cerebral ischemia in mice. This hypoxic-ischemic injury caused a remarkable decrease in oligodendroglial LDLR, with impaired oligodendroglial differentiation and survival. Oligodendroglial cholesterol levels, however, remained unchanged. Mouse miR-344e-3p and the human homolog miR-410-3p, 2 miRNAs directly targeting Ldlr, were identified in experimental and clinical leukoaraiosis and were thus implicated in the LDLR reduction. Lentiviral delivery of LDLR ameliorated demyelination following chronic cerebral ischemia. By contrast, Ldlr–/– mice displayed inadequate myelination in the corpus callosum. Ldlr–/– oligodendrocyte progenitor cells (OPCs) exhibited reduced ability to differentiate and myelinate axons in vitro. Transplantation with Ldlr–/– OPCs could not rescue the BCAS-induced demyelination. Such LDLR-dependent myelin restoration might involve a physical interaction of the Asn-Pro-Val-Tyr (NPVY) motif with the phosphotyrosine binding domain of Shc, which subsequently activated the MEK/ERK pathway. Together, our findings demonstrate that the aberrant oligodendroglial LDLR in chronic cerebral ischemia impairs myelination through intracellular signal transduction. Preservation of oligodendroglial LDLR may provide a promising approach to treat ischemic demyelination.
Authors
Yi Xie, Xiaohao Zhang, Pengfei Xu, Nana Zhao, Ying Zhao, Yunzi Li, Ye Hong, Mengna Peng, Kang Yuan, Ting Wan, Rui Sun, Deyan Chen, Lili Xu, Jingjing Chen, Hongquan Guo, Wanying Shan, Juanji Li, Rongrong Li, Yunyun Xiong, Dezhi Liu, Yuhui Wang, George Liu, Ruidong Ye, Xinfeng Liu
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Copyright © 2021 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)