Gastroenterology
Abstract
Variants in the UNC45A cochaperone have been recently associated with a syndrome combining diarrhea, cholestasis, deafness, and bone fragility. Yet the mechanism underlying intestinal failure in UNC45A deficiency remains unclear. Here, biallelic variants in UNC45A were identified by next-generation sequencing in 6 patients with congenital diarrhea. Corroborating in silico prediction, variants either abolished UNC45A expression or altered protein conformation. Myosin VB was identified by mass spectrometry as client of the UNC45A chaperone and was found misfolded in UNC45AKO Caco-2 cells. In keeping with impaired myosin VB function, UNC45AKO Caco-2 cells showed abnormal epithelial morphogenesis that was restored by full-length UNC45A, but not by mutant alleles. Patients and UNC45AKO 3D organoids displayed altered luminal development and microvillus inclusions, while 2D cultures revealed Rab11 and apical transporter mislocalization as well as sparse and disorganized microvilli. All those features resembled the subcellular abnormalities observed in duodenal biopsies from patients with microvillus inclusion disease. Finally, microvillus inclusions and shortened microvilli were evidenced in enterocytes from unc45a-deficient zebrafish. Taken together, our results provide evidence that UNC45A plays an essential role in epithelial morphogenesis through its cochaperone function of myosin VB and that UNC45A loss causes a variant of microvillus inclusion disease.
Authors
Rémi Duclaux-Loras, Corinne Lebreton, Jérémy Berthelet, Fabienne Charbit-Henrion, Ophelie Nicolle, Céline Revenu de Courtils, Stephanie Waich, Taras Valovka, Anis Khiat, Marion Rabant, Caroline Racine, Ida Chiara Guerrera, Júlia Baptista, Maxime M. Mahe, Michael W. Hess, Béatrice Durel, Nathalie Lefort, Céline Banal, Mélanie Parisot, Cecile Talbotec, Florence Lacaille, Emmanuelle Ecochard-Dugelay, Arzu Meltem Demir, Georg F. Vogel, Laurence Faivre, Astor Rodrigues, Darren Fowler, Andreas R. Janecke, Thomas Müller, Lukas A. Huber, Fernando Rodrigues-Lima, Frank M. Ruemmele, Holm H. Uhlig, Filippo Del Bene, Grégoire Michaux, Nadine Cerf-Bensussan, Marianna Parlato
Abstract
SMAD4, a mediator of TGF-β signaling, plays an important role in T cells to prevent inflammatory bowel disease (IBD). However, the precise mechanisms underlying this control remain elusive. Using both genetic and epigenetic approaches, we revealed an unexpected mechanism by which SMAD4 prevents naive CD8+ T cells from becoming pathogenic for the gut. Prior to the engagement of the TGF-β receptor, SMAD4 restrains the epigenetic, transcriptional, and functional landscape of the TGF-β signature in naive CD8+ T cells. Mechanistically, prior to TGF-β signaling, SMAD4 binds to promoters and enhancers of several TGF-β target genes, and by regulating histone deacetylation, suppresses their expression. Consequently, regardless of a TGF-β signal, SMAD4 limits the expression of TGF-β negative feedback loop genes, such as Smad7 and Ski, and likely conditions CD8+ T cells for the immunoregulatory effects of TGF-β. In addition, SMAD4 ablation conferred naive CD8+ T cells with both a superior survival capacity, by enhancing their response to IL-7, as well as an enhanced capacity to be retained within the intestinal epithelium, by promoting the expression of Itgae, which encodes the integrin CD103. Accumulation, epithelial retention, and escape from TGF-β control elicited chronic microbiota-driven CD8+ T cell activation in the gut. Hence, in a TGF-β–independent manner, SMAD4 imprints a program that preconditions naive CD8+ T cell fate, preventing IBD.
Authors
Ramdane Igalouzene, Hector Hernandez-Vargas, Nicolas Benech, Alexandre Guyennon, David Bauché, Célia Barrachina, Emeric Dubois, Julien C. Marie, Saïdi M’Homa Soudja
Abstract
Enhanced de novo lipogenesis mediated by sterol regulatory element-binding proteins (SREBPs) is thought to be involved in nonalcoholic steatohepatitis (NASH) pathogenesis. In this study, we assessed the impact of SREBP inhibition on NASH and liver cancer development in murine models. Unexpectedly, SREBP inhibition via deletion of the SREBP cleavage-activating protein (SCAP) in the liver exacerbated liver injury, fibrosis, and carcinogenesis, despite markedly reduced hepatic steatosis. These phenotypes were ameliorated by restoring SREBP function. Transcriptome and lipidome analyses revealed that SCAP–SREBP pathway inhibition altered the fatty acid (FA) composition of phosphatidylcholines due to both impaired FA synthesis and disorganized FA incorporation into phosphatidylcholine via lysophosphatidylcholine acyltransferase 3 (LPCAT3) downregulation, which led to endoplasmic reticulum (ER) stress and hepatocyte injury. Supplementation of phosphatidylcholines significantly improved liver injury and ER stress induced by SCAP deletion. The activity of SCAP-SREBP-LPCAT3 axis was found inversely associated with liver fibrosis severity in human NASH. SREBP inhibition also cooperated with impaired autophagy to trigger liver injury. Thus, excessively strong and broad lipogenesis inhibition was counterproductive for NASH therapy, which will have important clinical implications in NASH treatment.
Authors
Satoshi Kawamura, Yuki Matsushita, Shigeyuki Kurosaki, Mizuki Tange, Naoto Fujiwara, Yuki Hayata, Yoku Hayakawa, Nobumi Suzuki, Masahiro Hata, Mayo Tsuboi, Takahiro Kishikawa, Hiroto Kinoshita, Takuma Nakatsuka, Masaya Sato, Yotaro Kudo, Yujin Hoshida, Atsushi Umemura, Akiko Eguchi, Tsuneo Ikenoue, Yoshihiro Hirata, Motonari Uesugi, Ryosuke Tateishi, Keisuke Tateishi, Mitsuhiro Fujishiro, Kazuhiko Koike, Hayato Nakagawa
Abstract
Gastric carcinogenesis is mediated by complex interactions among Helicobacter pylori, host, and environmental factors. We now demonstrate that H. pylori augments gastric injury in INS-GAS mice under iron deficient conditions. Mechanistically, these phenotypes were not driven by alterations in the gastric microbiota; however, discovery-based and targeted metabolomics revealed that bile acids were significantly altered in H. pylori-infected mice with iron deficiency, with significant upregulation of deoxycholic acid (DCA), a carcinogenic bile acid. Severity of gastric injury was further augmented when H. pylori-infected mice were treated with DCA, and, in vitro, DCA increased translocation of the H. pylori oncoprotein CagA into host cells. Conversely, bile acid sequestration attenuated H. pylori-induced injury under conditions of iron deficiency. To translate these findings into human populations, the association between bile acid-sequestrant use and gastric cancer risk was evaluated in a large human cohort. Among 416,885 individuals, a significant dose-dependent reduction in risk was associated with cumulative bile acid-sequestrant use. Further, expression of the bile acid receptor TGR5 paralleled the severity of carcinogenic lesions in humans. These data demonstrate that increased H. pylori-induced injury within the context of iron deficiency is tightly linked to altered bile acid metabolism, which may promote gastric carcinogenesis.
Authors
Jennifer M. Noto, M. Blanca Piazuelo, Shailja C. Shah, Judith Romero-Gallo, Jessica L. Hart, Chao Di, James D. Carmichael, Alberto G. Delgado, Alese E. Halvorson, Robert A. Greevy, Jr., Lydia E. Wroblewski, Ayushi Sharma, Annabelle B. Newton, Margaret M. Allaman, Keith T. Wilson, M. Kay Washington, M. Wade Calcutt, Kevin L. Schey, Bethany P. Cummings, Charles R. Flynn, Joseph P. Zackular, Richard M. Peek, Jr.
Abstract
De novo and acquired resistance are major impediments to the efficacy of conventional and targeted cancer therapy. In unselected gastric cancer (GC) patients with advanced disease, trials combining chemotherapy and an anti-EGFR monoclonal antibody have been largely unsuccessful. In an effort to identify biomarkers of resistance so as to better select patients for such trials, we screened the secretome of chemotherapy-treated human GC cell lines. We found that levels of CGA, the α-subunit of glycoprotein hormones, were markedly increased in the conditioned media of chemoresistant GC cells, and CGA immunoreactivity was enhanced in GC tissues that progressed on chemotherapy. CGA levels in plasma increased in GC patients who received chemotherapy, and this increase was correlated with reduced responsiveness to chemotherapy and poor survival. Mechanistically, secreted CGA was found to bind to EGFR and activate EGFR signaling, thereby conferring a survival advantage to GC cells. N-glycosylation of CGA at Asn52 and Asn78 is required for its stability, secretion, and interaction with EGFR. GATA2 was found to activate CGA transcription, whose increase, in turn, induced the expression and phosphorylation of GATA2 in an EGFR-dependent manner, forming a positive feedback circuit that was initiated by GATA2 autoregulation upon sublethal exposure to chemotherapy. Based on this circuit, combination strategies involving anti-EGFR therapies or targeting CGA with microRNAs (miR-708-3p and miR-761) restored chemotherapy sensitivity. These findings identify a clinically actionable CGA/EGFR/GATA2 circuit and highlight CGA as a predictive biomarker and therapeutic target in chemoresistant GC.
Authors
Tianyu Cao, Yuanyuan Lu, Qi Wang, Hongqiang Qin, Hongwei Li, Hao Guo, Minghui Ge, Sarah E. Glass, Bhuminder Singh, Wenyao Zhang, Jiaqiang Dong, Feng Du, Airong Qian, Ye Tian, Xin Wang, Cunxi Li, Kaichun Wu, Daiming Fan, Yongzhan Nie, Robert J. Coffey, Xiaodi Zhao
Abstract
The striatin-interacting phosphatase and kinase (STRIPAK) complexes integrate extracellular stimuli to result in intracellular activities. Previously, we discovered STRIPAK to be a key machinery responsible for loss of the Hippo tumor suppressor signal in cancer. Here, we identified the Hippo-STRIPAK complex to be an essential player for the control of DNA double-strand break (DSB) repair and genomic stability. Specifically, the MST1/2 kinases were found, independent of the classical Hippo signaling, to directly phosphorylate ZMYND8 and hence result in suppression of DNA repair in the nucleus. In response to genotoxic stress, the cGAS-STING pathway was determined to relay nuclear DNA damage signals to the dynamic assembly of Hippo-STRIPAK via a TBK1-induced structural stabilization of the SIKE1-SLMAP arm. As such, STRIPAK-mediated MST1/2 inactivation was found to increase the DSB repair capacity of cancer cells and to endow these cells with resistance to radio/chemotherapy and PARP inhibition. Importantly, targeting the STRIPAK assembly with each of three distinct peptide inhibitors efficiently recovered the kinase activity of MST1/2 to suppress DNA repair and re-sensitize cancer cells to PARPi in both animal and patient-derived tumor models. Overall, our findings not only uncovered a previously unrecognized role for STRIPAK in modulating DSB repair, but also provided translational implications of co-targeting STRIPAK and PARP for a new type of synthetic lethality anti-cancer therapy.
Authors
Liwei An, Zhifa Cao, Pingping Nie, Hui Zhang, Zhenzhu Tong, Fan Chen, Yang Tang, Yi Han, Wenjia Wang, Zhangting Zhao, Qingya Zhao, Yuqin Yang, Yuanzhi Xu, Gemin Fang, Lei Shi, Huixiong Xu, Haiqing Ma, Shi Jiao, Zhaocai Zhou
Abstract
Gastrointestinal motility disorders involve alterations to the structure and/or function of the enteric nervous system (ENS) but the causal mechanisms remain unresolved in most cases. Homeostasis and disease in the ENS are processes that are regulated by enteric glia. Signaling mediated through type I lysophosphatidic acid receptors (LPAR1) has recently emerged as an important mechanism that contributes to disease, in part, through effects on peripheral glial survival and function. Enteric glia express LPAR1 but its role in ENS function and motility disorders is unknown. We used a combination of genetic, immunohistochemical, calcium imaging, and in vivo pharmacological approaches to investigate the role of LPAR1 in enteric glia. LPAR1 was enriched in enteric glia in mice and humans and LPA stimulated intracellular calcium responses in enteric glia, subsequently recruiting activity in a subpopulation of myenteric neurons. Blocking LPAR1 in vivo with AM966 attenuated gastrointestinal motility in mice and produced marked enteric neuro- and gliopathy. Samples from humans with chronic intestinal pseudo-obstruction (CIPO), a severe motility disorder, showed reduced glial LPAR1 expression in the colon and ileum. These data suggest that enteric glial LPAR1 signaling regulates gastrointestinal motility through enteric glia and could contribute to severe motility disorders in humans such as CIPO.
Authors
Mohammad M. Ahmadzai, Jonathon L. McClain, Christine Dharshika, Luisa Seguella, Fiorella Giancola, Roberto De Giorgio, Brian D. Gulbransen
Abstract
Anti-TNF antibodies are effective for treating patients with inflammatory bowel disease (IBD), but many patients fail to respond to anti-TNF therapy, highlighting the importance of TNF-independent disease. We previously demonstrated that acute deletion of two IBD susceptibility genes, A20 (Tnfaip3) and Abin-1 (Tnip1), in intestinal epithelial cells (IECs) sensitized mice to both TNF-dependent and TNF-independent death. Here we show that TNF-independent IEC death after A20 and Abin-1 deletion was rescued by germ-free derivation or deletion of MyD88, while deletion of Trif provided only partial protection. Combined deletion of Ripk3 and Casp8, which inhibits both apoptotic and necroptotic death, completely protected against death after acute deletion of A20 and Abin-1 in IECs. A20 and Abin-1-deficient IECs were sensitized to TNF-independent, TNFR-1-mediated death in response to lymphotoxin alpha (LT⍺) homotrimers. Blockade of LT⍺ in vivo reduced weight loss and improved survival when combined with partial deletion of MyD88. Biopsies of inflamed colon mucosa from patients with IBD exhibited increased LTA and IL1B expression, including a subset of patients with active colitis on anti-TNF therapy. These data show that microbial signals, MyD88, and LT⍺ all contribute to TNF-independent intestinal injury.
Authors
Iulia Rusu, Elvira Mennillo, Jared L. Bain, Zhongmei Li, Xiaofei Sun, Kimberly M. Ly, Yenny Y. Rosli, Mohammad Naser, Zunqiu Wang, Rommel Advincula, Philip Achacoso, Ling Shao, Bahram Razani, Ophir D. Klein, Alexander Marson, Jessie A. Turnbaugh, Peter J. Turnbaugh, Barbara A. Malynn, Averil Ma, Michael G. Kattah
Abstract
Functional gastrointestinal disorders (FGIDs) have prominent sex differences in incidence, symptoms, and treatment response that are not well understood. Androgens are steroid hormones present at much higher levels in males than females and could be involved in these differences. In adults with irritable bowel syndrome (IBS), a FGID that affects 5-10% of the population worldwide, we found that free testosterone levels were lower than those in healthy controls and inversely correlated with symptom severity. To determine how this diminished androgen signaling could contribute to bowel dysfunction, we depleted gonadal androgens in adult mice and found that this caused a profound deficit in gastrointestinal transit. Restoring a single androgen hormone was sufficient to rescue this deficit, suggesting that circulating androgens are essential for normal bowel motility in vivo. To determine the site of action, we probed androgen receptor expression in the intestine and discovered, unexpectedly, that a large subset of enteric neurons became androgen-responsive upon puberty. Androgen signaling to these neurons was required for normal colonic motility in adult mice. Taken together, these observations establish a role for gonadal androgens in the neural regulation of bowel function and link altered androgen levels with a common digestive disorder.
Authors
Daniella Rastelli, Ariel Robinson, Valentina N. Lagomarsino, Lynley T. Matthews, Rafla Hassan, Kristina Perez, William Dan, Peter D. Yim, Madison Mixer, Aleksandra Prochera, Amy Shepherd, Liang Sun, Kathryn Hall, Sarah Ballou, Anthony Lembo, Judy Nee, Meenakshi Rao
Abstract
Bitter taste receptors (TAS2R) serve as warning sensors in the lingual system against ingestion of potential poisonous food. Here, we investigated the functional role of TAS2Rs in the human gut and focused on their potential to trigger an additional host defense pathway in the intestine. Human jejunal crypts, especially from obese subjects, responded to bitter agonists by inducing the release of antimicrobial peptides (α-defensin 5 and REG3A) but also regulated the expression of other innate immune factors (mucins, chemokines) that affected E. coli growth. The effect of aloin on E. coli growth and on the release of the mucus glycoprotein CLCA1, identified via proteomics, was affected by TAS2R43 amino acid/deletion polymorphisms and thus confirmed a role for TAS2R43. RNA sequencing uncovered that denatonium benzoate induced an NRF2-mediated nutrient stress response and an unfolded protein response that increased the expression of the mitokine GDF15 but also ADM2 and the LDLR, genes that are involved in anorectic signaling and lipid homeostasis. To conclude, TAS2Rs in the intestine provide a promising target for treating diseases that involve disturbances in the innate immune system and in body weight control. Polymorphisms in TAS2Rs may be valuable genetic markers to predict therapeutic responses.
Authors
Kathrin I. Liszt, Qiaoling Wang, Mona Farhadipour, Anneleen Segers, Theo Thijs, Linda Nys, Ellen Deleus, Bart Van der Schueren, Christopher Gerner, Benjamin Neuditschko, Laurens J. Ceulemans, Matthias Lannoo, Jan Tack, Inge Depoortere
Copyright © 2022 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)