Research
Abstract
Metabolic dysfunction-associated steatotic liver disease (MASLD) and steatohepatitis (MASH) are leading causes of cirrhosis and hepatocellular carcinoma. Defects in autophagy contribute to the development of MASLD, however, the role of the Unc-51-like autophagy-activating kinase 1 (ULK1) in the pathophysiology of MASLD remains unclear. Herein, we show that ULK1, a serine/threonine kinase and core autophagy protein, is significantly repressed in human MASH livers, and that hepatocyte-specific loss of ULK1, unexpectedly, promotes hepatic steatosis and progression to liver fibrosis, without affecting basal autophagy flux. Phospho-proteomics identified the transcriptional coactivator NCOA3 as a downstream phospho-target of ULK1. Mechanistically, ULK1 phosphorylates NCOA3 to repress its transcriptional activity and restrain the CREB/CBP-mediated de novo lipogenic program. Accordingly, a phosphorylation-deficient NCOA3 mutant drives CREB/CBP-mediated lipogenesis, whereas genetic or pharmacological NCOA3 inhibition prevents steatosis, hepatic inflammation, and profibrotic signaling. Hence, ULK1-mediated NCOA3 phosphorylation is a fundamental and druggable checkpoint against the entire MASLD spectrum.
Authors
Young Do Koo, Romilia Tatiana Castillo, Asha Sukumaran Nair, Michael Garneau, Chad Gochee, Zachary V. Campbell, Tashya Shreyas Vakil, Jua Ha, Alex Marti, Jamie Soto, Debajyoti Das, Nuria Martinez-Lopez, Shipra Sharma, Yennifer Delgado, Callie Phung, Immy A. Ashley, Edmund D. Kapelczak, Rashel Jacobo, Eric T. Weatherford, Dao-Fu Dai, Jihane N. Benhammou, Andrea G. Marshall, Antentor Hinton Jr, Ling Yang, Renata O. Pereira, Tara TeSlaa, Mehdi Bouhaddou, Rajat Singh, E. Dale Abel
Abstract
Early initiation of antiretroviral therapy (ART) in perinatally HIV-infected children significantly limits the establishment of the viral reservoir. However, the long-term impact of this intervention remains unclear. We measured the frequency of inducible, translation-competent, and replication-competent proviruses in samples from 62 children who initiated ART early (median 9.9 weeks) and remained virally suppressed for up to 9.9 years. Only a small fraction of HIV genomes produced HIV transcripts (1.8%), viral proteins (<0.9%) or infectious virions (<0.05%). Accordingly, replication-competent virus was detected in only 15% of the participants. Despite the predominance of naïve cells in pediatric blood, most proviruses were detected in memory CD4+ T cells, especially central memory cells (contribution 41%). Longitudinal analysis revealed a biphasic decay in HIV DNA: an initial decline followed by long-term stability, which was associated with extensive expansions of infected T-cell clones. In contrast, inducible proviruses declined continuously and became undetectable in most children after five years. Near full-length sequencing of 1,305 HIV genomes revealed a dramatic reduction in genetically intact proviruses, from 40% pre-ART to 0.3% after 7 years of ART. Together, these findings suggest that the intact viral reservoir rapidly decays in early-treated children, offering critical insights for pediatric HIV cure strategies.
Authors
Marta Massanella, Caroline Dufour, Amélie Pagliuzza, Audrée Lemieux, Corentin Richard, Jintanat Ananworanich, Louise Leyre, Thidarat Jupimai, Supranee Buranapraditkun, Rapisa Nantanee, Julie L. Mitchell, Panadda Sawangsinth, Mark de Souza, Piyarat Suntarattiwong, Suparat Kanjanavanit, Pope Kosalaraksa, Thitiporn Borkird, Witaya Petdachai, Kulkanya Chokephaibulkit, Lydie Trautmann, Rémi Fromentin, Thanyawee Puthanakit, Nicolas Chomont
Abstract
Crimean-Congo hemorrhagic fever virus (CCHFV) is an emerging arboviral and zoonotic bunyavirus. CCHFV can infect livestock, wild animals, and humans. Here we report the isolation of a panel of monoclonal antibodies (mAbs) from the B cells of an immune individual following a natural nosocomial infection. We determined that the panel comprised antibodies that bound to two glycoproteins: 1) the carboxy-terminal glycoprotein (Gc) that serves as the fusion protein and 2) the glycoprotein 38 (GP38). By antibody variable gene analysis, we identified genetic diversity in the B cell response to CCHFV within a single donor for both Gc- and GP38-specific responses. Protection against most bunyavirus-associated diseases is mediated principally by neutralizing antibodies, but here, we found that neutralization activity was not associated with protection. Gc-specific antibodies to diverse antigenic sites neutralized only weakly and did not protect against heterologous virus challenge. GP38-specific antibodies bound to two dominant antigenic sites on the glycoprotein. Although GP38-specific antibodies did not neutralize the virus, one mediated protection against heterologous virus challenge in an experimental model of infection in mice primarily by complement-mediated activity. These studies support the model of development of CCHFV countermeasures that induce protection against GP38 in vivo.
Authors
Nathaniel S. Chapman, Viktoriya Borisevich, Nurgun Kose, Luke Myers, Stephen Priest, Éric Bergeron, Elena Trigo Esteban, María Paz Sanchez-Seco, Jose Melero, Thomas Geisbert, Robert W. Cross, James E. Crowe Jr.
Abstract
Cilia are cellular organelles extruding from the surface of various cell types, serving either sensory or motile functions. Retinitis pigmentosa GTPase regulator (RPGR) variants affect both photoreceptor sensory cilia and airway motile cilia, leading to retinitis pigmentosa (RP) and primary ciliary dyskinesia (PCD), respectively. Not all patients develop PCD, and it remains unclear which RPGR variants predispose patients to PCD. Here, we leverage 2D organoids, super-resolution microscopy, and live-cell imaging to characterize the multiciliated cells (MCCs) from patients with different RPGR variants and CRISPR-modified RPGR KO MCCs. We demonstrate that MCCs with RPGR variants have reduced ciliation, shorter cilia, impaired cilia beat, or cilia beat incoordination, potentially resulting in compromised mucociliary clearance and lung diseases. Moreover, we show that RPGR regulates motile cilia through interfering with F-actin dynamics, evidenced by the undissolved F-actin meshwork in RPGR-deficient MCCs, and the defects can be ameliorated with either Latrunculin A or Y27632 treatment. Though PCD was observed only in patients with variants that affect both isoforms, patients with RPGRORF15 variants also showed cilia and airway anomalies. All RPGR variants affect motile cilia one way or another, and the mechanisms involve the accumulation of apical F-actin.
Authors
Yang Wu, Erika Tavares, Binrun Liang, Wallace B Wee, Vito Mennella, Han-Chao Feng, Jiaying Cao, Pui Yee Wong, Jiayi Zheng, Mu He, Kirk AJ Stephenson, Liran Hanan Hochma, Janice Min Li, Nan-Peng Chen, Sharon D Dell, Elise Heon, ZHEN LIU
Abstract
Epigenetic dysregulation is associated with immune evasion and immune checkpoint blockade (ICB) resistance. Here, using in vivo CRISPR-Cas9 screens targeting epigenetic-related factors in mouse tumor models treated with ICB, we identified Chromobox 4 (CBX4) as a key negative regulator of immune tumor microenvironment. Single-cell RNA sequencing and spatial transcriptomics analyses of patients receiving neoadjuvant anti-PD-1 therapy revealed high CBX4 expression in both tumor cells and immunosuppressive tumor-associated macrophage subpopulations, with preferential accumulation in non-responders. Deficiency of CBX4 in macrophages or tumor cells, induced robust anti-tumor immunity, increased infiltration and cytotoxic activity of CD8+ T cells and NK cells, thereby heightening the sensitivity of ICB treatment. Mechanistically, CBX4 targeted H3K9me3 and H3K27me3-marked endogenous retroelements such as RLTR4-Mm-int. Loss of CBX4 derepressed retrotransposons, activating cytosolic RNA-sensing pathways and triggering type I interferon response, ultimately leading to robust inflamed TME. Moreover, we uncovered a negative correlation between CBX4 expression and immune responses, retrotransposon levels as well as the prognosis of patients with hepatocellular carcinoma (HCC) undergoing ICB therapy. Our study establishes CBX4 as an epigenetic immune checkpoint through the epigenetic silencing of retrotransposons, remodeling immune TME and thus providing a promising therapeutic target to enhance tumor immunogenicity and overcoming immunotherapy resistance.
Authors
Zhibo Ma, Wenlong Jia, Xi Zhou, Jing Liu, Qingwen Li, Ruizhi Chang, Shiqi Gu, Naonao Yuan, Zhishui Chen, Peixiang Lan
Abstract
VPS13A is an intracellular lipid transfer protein comprising over 3,000 amino acids. Mutations in human VPS13A cause VPS13A disease, a neurodegenerative disorder that affects movement and cognition. VPS13A forms a complex with the membrane protein XK to mediate ATP-induced phospholipid scrambling in the plasma membrane. Here, we established a mouse cell system expressing full-length mouse VPS13A and examined its interaction with XK. Mutational analysis revealed that VPS13A binds to XK through a C-terminal β-strand that interacts with a β-hairpin in the central region of XK, an interaction essential for scramblase activity. The XK paralog XKR2, which contains a similar β-hairpin structure, also associates with VPS13A and supports phospholipid scrambling. We analyzed ten mouse VPS13A variants corresponding to patient mutations and classified them into four groups: (1) L67P, I90K, and W2453R, which showed reduced expression; (2) A1091P and M3080R, which were normally expressed but lacked scramblase activity; (3) S1446P, Q2689H, Y2713C, and R3084H, which modestly impaired expression or activity; and (4) I2763R, which altered cell size, and disrupted ER independently of XK. These findings define the VPS13A–XK interaction interface, clarify the functional impact of disease-causing mutations, and reveal an unexpected gain-of-function mutation of a VPS13A variant.
Authors
Xing Lin, Yuta Ryoden, Chigure Suzuki, Hiroyuki Ishikawa, Takaharu Sakuragi, Yasuo Uchiyama, Shigekazu Nagata
Abstract
Resistance to HIF-2α inhibitors such as Belzutifan underscores the need to better understand how HIF-2α is transcriptionally regulated in clear cell renal cell carcinoma (ccRCC). Here, we uncover a cytokine-driven enhancer mechanism that sustains HIF-2α expression through the JAK1–STAT3 signaling pathway. Using a genome-wide CRISPR screen in VHL-deficient ccRCC cells, we identified SOCS3 as a key negative regulator of HIF-2α. Mechanistically, loss of SOCS3 activates JAK1–STAT3 signaling, leading to the recruitment of STAT3 to distal enhancers upstream of EPAS1 that physically loop to its promoter to drive HIF-2α transcription. This cytokine–enhancer circuit was recapitulated in ccRCC patient samples and functionally validated using CRISPR interference, which disrupted enhancer–promoter looping and reduced tumor growth in HIF-2α–dependent models. SOCS3 overexpression or pharmacologic inhibition of JAK1/STAT3 markedly suppressed HIF-2α expression and tumor progression both in vitro and in vivo. Unlike prior studies focusing on VHL/HIF occupancy–driven enhancer activation, this work defines a trans-acting cytokine–JAK1–STAT3 pathway that transcriptionally controls EPAS1. Together, these findings reveal a targetable enhancer mechanism that sustains HIF-2α expression and suggest that combined inhibition of JAK1/STAT3 and HIF-2α may overcome therapeutic resistance in kidney cancer.
Authors
Jun Fang, Jeremy M Simon, Tao Wang, Yunpeng Gao, Xianju Bi, Lianxin Hu, Chengheng Liao, Cheng Zhang, Yayoi Adachi, Jin Zhou, Hongyi Liu, Qian Liang, James A. Nathan, Ram Mani, James Brugarolas, Qing Zhang
Abstract
The lymphatic system plays a central role in lipid absorption by transporting triglyceride-rich particles called chylomicrons (CMs) from the small intestine to the systemic circulation. However, the molecular mechanism by which CMs get into the intestinal lymphatics is unknown. Here we demonstrated that GPR182, an atypical chemokine receptor in lymphatic endothelial cells, mediates dietary fat absorption. GPR182 knockout mice exhibit a selective increase in circulating high-density lipoproteins and are resistant to dietary-induced obesity. GPR182 ablation in mice leads to poor lipid absorption and thereby a delay in growth during development. GPR182 broadly interacts with and transports lipoproteins. Transmission electron microscopy analysis reveals that mechanistically, loss of GPR182 prevents CMs from entering the lacteal lumen of the small intestine. Consistent with this, GPR182 blockade with monoclonal antibodies protects mice from diet- induced obesity and treats existing obesity. Together, our study identifies GPR182 as a lipoprotein receptor that mediates dietary fat absorption and supports GPR182 blockade as a feasible approach to treat obesity and related disorders.
Authors
Zhiwei Sun, Robert J. Torphy, Emily N. Miller, Anza Darehshouri, Isaac Vigil, Taichi Terai, Eck Eleanor, Yi Sun, Yujie Guo, Dustin P. Fykstra, Elliott J. Yee, Junyi Hu, Ross M. Kedl, Erika L. Lasda, Jay R. Hesselberth, Julie A. Siegenthaler, Paul S. MacLean, Kimberley D. Bruce, Gwendalyn J. Randolph, Richard D. Schulick, Yuwen Zhu
Abstract
The peritoneal cavity contains a large population of GATA6-expressing large peritoneal macrophages (LPMs), known to support healing of intra-abdominal organs. In this study, we aimed to explore their full sphere of influence by examining their ability to perform wound healing at distant sites outside the cavity. In a mouse model combining a remote skin injury with peritoneal stimulation we observed a significant acceleration of skin wound healing in response to LPM activation. Tracking GATA6-expressing LPMs, we demonstrated that LPMs do not migrate to distant wound sites following peritoneal activation. Using parabiosis experiments and administration of activated peritoneal contents indicated an important role of molecules secreted by LPMs in remote skin wound healing. More specifically, proteomic and transcriptomic analyses identified fibronectin as a key factor produced by activated LPMs. In fact, depletion of LPMs or genetic knockout of fibronectin in myeloid cells eliminated the enhanced healing effect. These findings highlight the endocrine function of LPMs in systemic tissue repair, challenging the traditional perspective of plasma fibronectin being exclusively liver-derived. Our results suggest that LPMs, strategically positioned in the peritoneal cavity, serve as a source of circulating fibronectin, promoting matrix formation and accelerating wound healing at distant sites.
Authors
Lilian Salm, Simone N. Zwicky, Daniel Spari, Tural Yarahmadov, Marie Siwicki, Fernanda VS Castanheira, Jonas Zbinden, Deborah Stroka, Joel Zindel, Antoine Dufour, Paul Kubes, Guido Beldi
Abstract
Single-agent anti-PD-1 antibody is ineffective for pancreatic ductal adenocarcinoma (PDAC) due to its immunosuppressive tumor-microenvironment (TME). KRAS-mutations contribute to the inflammatory TME and therapeutic resistance by upregulating IL-8 via MAPK pathways. Thus, this study attempted to overcome the resistance to anti-PD-1 antibody by targeting downstream KRAS-effectors. The study found that the resistance to anti-PD-1 antibody can be overcome through MEK1/2-inhibition. The combination of anti-PD-1 antibody and MEK inhibitor displayed antitumor activity in Krasmut (mutated) KPC mouse tumors, but not KrasWT (wild-type) Panc02 tumors. The combination of anti-PD-1 antibody and MEK inhibitor induced recruitment of tumor-associated neutrophils (TANs) via CXCR2, an IL-8 receptor, and increased memory CD8+ T cells and IFNgamma production in treatment-sensitive tumors. However, larger tumors still resisted to the combination of anti-PD-1 antibody and MEK inhibitor likely due to hypoxia/necrosis-induced NETosis and associated paucity of CD8+ T cells. The subsequent addition of anti-CXCR2 antibody overcame this resistance by blocking TAN-infiltration to hypoxic/necrotic areas. Consistently, a risk-score based on the NETosis-MAPK signaling interaction is significantly associated with poorer survival in human PDACs. This study thus provides a new venue for overcoming resistance to strategies targeting KRAS signaling.
Authors
Brian Herbst, Alex B. Blair, Yiming Li, Elizabeth M. Jaffee, Lei Zheng
No posts were found with this tag.
Copyright © 2026 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)