Mechanisms underlying motor neuron degeneration in amyotrophic lateral sclerosis (ALS) are yet unclear. Specific deletion of the ER-component membralin in astrocytes manifested postnatal motor defects and lethality in mice, causing the accumulation of extracellular glutamate through reducing the glutamate transporter EAAT2. Restoring EAAT2 levels in membralin-KO astrocytes limited astrocyte-dependent excitotoxicity in motor neurons. Transcriptomic profiles from mouse astrocytic membralin-KO motor cortex indicated significant perturbation in KEGG pathway components related to ALS, including downregulation of Eaat2 and upregulation of Tnfrsf1a. Changes in gene expression with membralin deletion also overlapped with mouse ALS models and reactive astrocytes. Our results show that activation of the TNF receptor (TNFR1) NFκB pathway known to suppress Eaat2 transcription was upregulated with membralin deletion. Further, reduced membralin and EAAT2 levels correlated with disease progression in spinal cord from SOD1-mutant mouse models, and reductions in membralin/EAAT2 were observed in human ALS spinal cord. Importantly, overexpression of membralin in SOD1G93A astrocytes decreased TNFR1 levels and increased EAAT2 expression, and improved motor neuron survival. Importantly, upregulation of membralin in SOD1G93A mice significantly prolonged mouse survival. Our study provided a mechanism for ALS pathogenesis where membralin limited glutamatergic neurotoxicity, suggesting that modulating membralin had potential in ALS therapy.
Lu-Lin Jiang, Bing Zhu, Yingjun Zhao, Xiaoguang Li, Tongfei Liu, Juan Pina-Crespo, Lisa Zhou, Wenxi Xu, Maria J. Rodriguez, Haiyang Yu, Don W. Cleveland, John Ravits, Sandrine Da Cruz, Tao Long, Dongxian Zhang, Timothy Y. Huang, Huaxi Xu
Gliomas account for approximately 80% of primary malignant tumors in the central nervous system. Despite aggressive therapy, the prognosis of patients remains extremely poor. Glioma stem cells (GSCs), considered a potential target of therapy for their crucial role in therapeutic resistance and tumor recurrence, are believed to be key factors in the disappointing outcome. Here, we took advantage of GSCs as the cell model to perform high-throughput drug screening, and the old antibiotic clofoctol was identified as the most effective compound, showing reduction of colony formation and induction of apoptosis of GSCs. Moreover, growth of tumors was obviously inhibited in vivo after clofoctol treatment especially in primary patient-derived xenografts and transgenic xenografts. The anticancer mechanisms demonstrated by analysis of related downstream genes and discovery of the targeted binding protein revealed that clofoctol exhibited the inhibition of GSCs by upregulation of Krüppel-like factor 13 (KLF13), a tumor suppressor gene, through clofoctol’s targeted binding protein, Upstream of N-ras (UNR). Collectively, these data demonstrate that induction of KLF13 expression suppressed growth of gliomas and provide a potential therapy for gliomas targeting GSCs. Importantly, our results also identify the RNA-binding protein UNR as a drug target.
Yan Hu, Meilian Zhang, Ningyu Tian, Dengke Li, Fan Wu, Peishan Hu, Zhixing Wang, Liping Wang, Wei Hao, Jingting Kang, Bin Yin, Zhi Zheng, Tao Jiang, Jiangang Yuan, Boqin Qiang, Wei Han, Xiaozhong Peng
Lactation induces bone loss to provide sufficient calcium in the milk, a process that involves osteoclastic bone resorption but also osteocytes and perilacunar resorption. The exact mechanisms by which osteocytes contribute to bone loss remain elusive. Osteocytes express genes required in osteoclasts for bone resorption, including cathepsin K (Ctsk), and lactation elevates their expression. We show that Ctsk deletion in osteocytes prevented the increase in osteocyte lacunar area seen during lactation, as well as the effects of lactation to increase osteoclast numbers and decrease trabecular bone volume, cortical thickness, and mechanical properties. In addition, we show that Ctsk deletion in osteocytes increased bone parathyroid hormone–related peptide (PTHrP) and prevented the decrease in serum parathyroid hormone (PTH) induced by lactation, but amplified the increase in serum 1,25-dyhydroxyvitamin D [1,25(OH)2D]. The net result of these changes is to maintain serum and milk calcium levels in the normal range, ensuring normal offspring skeletal development. Our studies confirm the fundamental role of osteocytic perilacunar remodeling in physiological states of lactation and provide genetic evidence that osteocyte-derived Ctsk contributes not only to osteocyte perilacunar remodeling, but also to the regulation of PTH, PTHrP, 1,25(OH)2D, osteoclastogenesis, and bone loss in response to the high calcium demand associated with lactation.
Sutada Lotinun, Yoshihito Ishihara, Kenichi Nagano, Riku Kiviranta, Vincent T. Carpentier, Lynn Neff, Virginia Parkman, Noriko Ide, Dorothy Hu, Pamela Dann, Daniel Brooks, Mary L. Bouxsein, John Wysolmerski, Francesca Gori, Roland Baron
Lumen integrity in vascularization requires fully differentiated endothelial cells (ECs). Here, we report that endothelial-mesenchymal transitions (EndMTs) emerged in ECs of cerebral arteriovenous malformation (AVMs) and caused disruption of the lumen or lumen disorder. We show that excessive Sry-box 2 (Sox2) signaling was responsible for the EndMTs in cerebral AVMs. EC-specific suppression of Sox2 normalized endothelial differentiation and lumen formation and improved the cerebral AVMs. Epigenetic studies showed that induction of Sox2 altered the cerebral-endothelial transcriptional landscape and identified jumonji domain–containing protein 5 (JMJD5) as a direct target of Sox2. Sox2 interacted with JMJD5 to induce EndMTs in cerebral ECs. Furthermore, we utilized a high-throughput system to identify the β-adrenergic antagonist pronethalol as an inhibitor of Sox2 expression. Treatment with pronethalol stabilized endothelial differentiation and lumen formation, which limited the cerebral AVMs.
Jiayi Yao, Xiuju Wu, Daoqin Zhang, Lumin Wang, Li Zhang, Eric X. Reynolds, Carlos Hernandez, Kristina I. Boström, Yucheng Yao
Lymph nodes (LNs) filter lymph to mount effective immune responses. Small soluble lymph-borne molecules from the periphery enter the draining LNs via a reticular conduit system. Intact antibodies and other larger molecules, in contrast, are physically unable to enter the conduits, and they are thought to be transported to the LNs only within migratory DCs after proteolytic degradation. Here, we discovered that lymph-borne antibodies and other large biomolecules enter within seconds into the parenchyma of the draining LN in an intact form. Mechanistically, we found that the uptake of large molecules is a receptor-independent, fluid-phase process that takes place by dynamin-dependent vesicular transcytosis through the lymphatic endothelial cells in the subcapsular sinus of the LN. Physiologically, this pathway mediates a very fast transfer of large protein antigens from the periphery to LN-resident DCs and macrophages. We show that exploitation of the transcytosis system allows enhanced whole-organ imaging and spatially controlled lymphocyte activation by s.c. administered antibodies in vivo. Transcytosis through the floor of the subcapsular sinus thus represents what we believe to be a new physiological and targetable mode of lymph filtering.
Laura Kähäri, Ruth Fair-Mäkelä, Kaisa Auvinen, Pia Rantakari, Sirpa Jalkanen, Johanna Ivaska, Marko Salmi
Because of the less-than-robust response to therapy and impact on choice of optimal chemotherapy and prognosis, chronic kidney disease has drawn attention in the treatment of multiple myeloma, a malignant hematologic disorder that can produce significant amounts of monoclonal immunoglobulin free light chains (FLCs). These low-molecular-weight proteins are relatively freely filtered through the glomerulus and are reabsorbed by the proximal tubule. The present study demonstrated that during the process of metabolism of immunoglobulin FLCs, ROS activated the STAT1 pathway in proximal tubule epithelium. STAT1 activation served as the seminal signaling molecule that produced the proinflammatory molecule IL-1β, as well as the profibrotic agent TGF-β by this portion of the nephron. These effects occurred in vivo and were produced specifically by the generation of hydrogen peroxide by the VL domain of the light chain. To the extent that the experiments reflect the human condition, these studies offer insights into the pathogenesis of progressive kidney failure in the setting of lymphoproliferative disorders, such as multiple myeloma, that feature increased circulating levels of monoclonal immunoglobulin fragments that require metabolism by the kidney.
Wei-Zhong Ying, Xingsheng Li, Sunil Rangarajan, Wenguang Feng, Lisa M. Curtis, Paul W. Sanders
Phosphorylation of dynamin-related protein 1 (Drp1) represents an important regulatory mechanism for mitochondrial fission. Here, we established the role of Drp1 serine 600 (Drp1S600) phosphorylation in mitochondrial fission in vivo and assessed the functional consequences of targeted elimination of the Drp1S600 phosphorylation site in the progression of diabetic nephropathy (DN). We generated a knockin mouse in which S600 was mutated to alanine (Drp1S600A). We found that diabetic Drp1S600A mice exhibited improved biochemical and histological features of DN along with reduced mitochondrial fission and diminished mitochondrial ROS in vivo. Importantly, we observed that the effect of Drp1S600 phosphorylation on mitochondrial fission in the diabetic milieu was stimulus dependent but not cell type dependent. Mechanistically, we show that mitochondrial fission in high-glucose conditions occurs through concomitant binding of phosphorylated Drp1S600 with mitochondrial fission factor (MFF) and actin-related protein 3 (Arp3), ultimately leading to accumulation of F-actin and Drp1 on the mitochondria. Taken together, these findings establish the idea that a single phosphorylation site in Drp1 can regulate mitochondrial fission and progression of DN in vivo and highlight the stimulus-specific consequences of Drp1S600 phosphorylation in mitochondrial dynamics.
Daniel L. Galvan, Jianyin Long, Nathanael Green, Benny H. Chang, Jamie S. Lin, Paul Schumacker, Luan D. Truong, Paul Overbeek, Farhad R. Danesh
About 1% of all newborns are affected by congenital heart disease (CHD). Recent findings identify aberrantly functioning cilia as a possible source for CHD. Faulty cilia also prevent the development of proper left-right asymmetry and cause heterotaxy, the incorrect placement of visceral organs. Intriguingly, signaling cascades such as mTOR that influence mitochondrial biogenesis also affect ciliogenesis, and can cause heterotaxy-like phenotypes in zebrafish. Here, we identify levels of mitochondrial function as a determinant for ciliogenesis and a cause for heterotaxy. We detected reduced mitochondrial DNA content in biopsies of heterotaxy patients. Manipulation of mitochondrial function revealed a reciprocal influence on ciliogenesis and affected cilia-dependent processes in zebrafish, human fibroblasts and Tetrahymena thermophila. Exome analysis of heterotaxy patients revealed an increased burden of rare damaging variants in mitochondria-associated genes as compared with 1000 Genome controls. Knock down of such candidate genes caused cilia elongation and ciliopathy-like phenotypes in zebrafish, which could not be rescued by RNA encoding damaging rare variants identified in heterotaxy patients. Our findings suggest that ciliogenesis is coupled to the abundance and function of mitochondria. Our data further reveal disturbed mitochondrial function as an underlying cause for heterotaxy-linked CHD and provide a mechanism for unexplained phenotypes of mitochondrial disease.
Martin D. Burkhalter, Arthi Sridhar, Pedro Sampaio, Raquel Jacinto, Martina S. Burczyk, Cornelia Donow, Max Angenendt, Competence Network for Congenital Heart Defects Investigators, Maja Hempel, Paul Walther, Petra Pennekamp, Heymut Omran, Susana S. Lopes, Stephanie M. Ware, Melanie Philipp
Recent studies have demonstrated that CD4+ T cells can efficiently reject MHC-II–negative tumors. This requires indirect presentation of tumor-associated antigens on surrounding antigen-presenting cells. We hypothesized that intercellular transfer of proteins is not the sole consequence of cell death–mediated protein release, but depends on heat-shock cognate protein 70 (HSC70) and its KFERQ-like binding motif on substrate proteins. Using human Y chromosome antigen DBY, we showed that mutation of one of its 2 putative binding motifs markedly diminished T cell activation after indirect presentation and reduced protein-protein interaction with HSC70. Intercellular antigen transfer was shown to be independent of cell-cell contact, but relied on engulfment within secreted microvesicles. In vivo, alterations of the homologous KFERQ-like motif in murine DBY hampered tumor rejection, T cell activation, and migration into the tumor and substantially impaired survival. Collectively, we show that intercellular antigen transfer of DBY is tightly regulated via binding to HSC70 and that this mechanism influences recognition and rejection of MHC-II–negative tumors in vivo.
Sascha Kretschmann, Stefanie Herda, Heiko Bruns, Josefine Russ, Edith D. van der Meijden, Ursula Schlötzer-Schrehardt, Marieke Griffioen, Il-Kang Na, Andreas Mackensen, Anita N. Kremer
Hypertension is a primary risk factor for cardiovascular diseases including myocardial infarction and stroke. Major determinants of blood pressure are vasodilatory factors such as nitric oxide (NO) released from the endothelium under the influence of fluid shear stress exerted by the flowing blood. Several endothelial signaling processes mediating fluid shear stress–induced formation and release of vasodilatory factors have been described. It is, however, still poorly understood how fluid shear stress induces these endothelial responses. Here we show that the endothelial mechanosensitive cation channel PIEZO1 mediated fluid shear stress–induced release of adrenomedullin, which in turn activated its Gs-coupled receptor. The subsequent increase in cAMP levels promoted the phosphorylation of endothelial NO synthase (eNOS) at serine 633 through protein kinase A (PKA), leading to the activation of the enzyme. This Gs/PKA-mediated pathway synergized with the AKT-mediated pathways leading to eNOS phosphorylation at serine 1177. Mice with endothelium-specific deficiency of adrenomedullin, the adrenomedullin receptor, or Gαs showed reduced flow-induced eNOS activation and vasodilation and developed hypertension. Our data identify fluid shear stress–induced PIEZO1 activation as a central regulator of endothelial adrenomedullin release and establish the adrenomedullin receptor and subsequent Gs-mediated formation of cAMP as a critical endothelial mechanosignaling pathway regulating basal endothelial NO formation, vascular tone, and blood pressure.
Andras Iring, Young-June Jin, Julián Albarrán-Juárez, Mauro Siragusa, ShengPeng Wang, Péter T. Dancs, Akiko Nakayama, Sarah Tonack, Min Chen, Carsten Künne, Anna M. Sokol, Stefan Günther, Alfredo Martínez, Ingrid Fleming, Nina Wettschureck, Johannes Graumann, Lee S. Weinstein, Stefan Offermanns
No posts were found with this tag.