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Sachin Kumar, Marie-Dominique Filippi
Transmission electron microscopy (TEM) provides details of the cellular organization and ultrastructure. However, TEM analysis of rare cell populations, especially cells in suspension such as hematopoietic stem cells (HSCs), remains limited due to the requirement of a high cell number during sample preparation. There are a few cytospin or monolayer approaches for TEM analysis from scarce samples, but these approaches fail to get significant quantitative data from the limited number of cells. Here, an alternative and novel approach for sample preparation in TEM studies is described for rare cell populations that enables quantitative analysis...
October 12, 2016: Journal of Visualized Experiments: JoVE
Sang-Ho Kwon, Sekyung Oh, Marisa Nacke, Keith E Mostov, Joshua H Lipschutz
Exosomes, 40-150 nm extracellular vesicles, transport biological macromolecules that mediate intercellular communications. While exosomes are known to originate from maturation of endosomes into multivesicular endosomes (MVEs; also known as multivesicular bodies, MVBs) with subsequent fusion of the MVEs with the plasma membrane, it remains unclear how cargos are selected for exosomal release. Using an inducible expression system for the exosome cargo protein GPRC5B and following its trafficking trajectory, we show here that newly synthesized GPRC5B protein accumulates in the Golgi complex prior to its release into exosomes...
October 20, 2016: Journal of Biological Chemistry
Min Wu, Jinyan Huang, Jianming Zhang, Cyril Benes, Bo Jiao, Ruibao Ren
RAS oncogenic mutations are common in human cancers, but RAS proteins have been difficult to target. We sought to identify pharmacological agents to block the RAS oncogenic signaling by a distinct mechanism. Since the biological activity of RAS proteins rely upon lipid modifications and RAS regulates lipid metabolisms in cancer cells, we screened a bioactive lipid library using a RAS specific cell viability assay. We report the discovery of a new class of inhibitors for RAS transformation. Compounds in the class represented by endocannabinoid N-arachidonoyl dopamine (NADA) can induce cell oncosis, independent of its ability to engage cannabinoid receptors...
October 19, 2016: Molecular Cancer Therapeutics
Mariana Pavel, David C Rubinsztein
Autophagy (literally "self-eating") is an evolutionarily conserved degradation process where cytoplasmic components are engulfed by vesicles called autophagosomes, which are then delivered to lysosomes, where their contents are degraded. Under stress conditions, such as starvation or oxidative stress, autophagy is upregulated in order to degrade macromolecules and restore the nutrient balance. The source of membranes that participate in the initial formation of phagophores is still incompletely understood and many intracellular structures have been shown to act as lipid donors, including the endoplasmic reticulum, Golgi, nucleus, mitochondria and the plasma membrane...
October 18, 2016: FEBS Journal
Shilpi Yadav, Jonathan K Williamson, Maria A Aronova, Andrew A Prince, Irina D Pokrovskaya, Richard D Leapman, Brian Storrie
Platelets are small, anucleate cell fragments that are central to hemostasis, thrombosis, and inflammation. They are derived from megakaryocytes from which they inherit their organelles. As platelets can synthesize proteins and contain many of the enzymes of the secretory pathway, one might expect all mature human platelets to contain a stacked Golgi apparatus, the central organelle of the secretory pathway. By thin section electron microscopy, stacked membranes resembling the stacked Golgi compartment in megakaryocytes and other nucleated cells can be detected in both proplatelets and platelets...
October 18, 2016: Platelets
Wen Du, Maoge Zhou, Wei Zhao, Dongwan Cheng, Lifen Wang, Jingze Lu, Eli Song, Wei Feng, Yanhong Xue, Pingyong Xu, Tao Xu
Secretory granules, also known as dense core vesicles, are generated at the trans-Golgi network and undergo several maturation steps, including homotypic fusion of immature secretory granules (ISGs), and processing of prehormones to yield active peptides. The molecular mechanisms governing secretory granule maturation are largely unknown. Here, we investigate a highly conserved protein named HID-1 in a mouse model. A conditional knockout of HID-1 in pancreatic β cells leads to glucose intolerance and a remarkable increase in the serum proinsulin/insulin ratio caused by defective proinsulin processing...
October 18, 2016: ELife
Michael Saul, Petra Majdak, Samuel Perez, Matthew Reilly, Theodore Garland, Justin S Rhodes
Though exercise is critical for health, many lack the motivation to exercise, and it is unclear how motivation might be increased. To uncover the molecular underpinnings of increased motivation for exercise, we analyzed the transcriptome of the striatum in four mouse lines selectively bred for high voluntary wheel running and four non-selected control lines. The striatum was dissected and RNA was extracted and sequenced from four individuals of each line. We found multiple genes and gene systems with strong relationships to both selection and running history over the previous 6 days...
October 17, 2016: Genes, Brain, and Behavior
Carol A Casey, Ganapati Bhat, Melissa S Holzapfel, Armen Petrosyan
BACKGROUND: It is known that ethanol (EtOH) and its metabolites have a negative effect on protein glycosylation. The fragmentation of the Golgi apparatus induced by alteration of the structure of largest Golgi matrix protein, giantin, is the major consequence of damaging effects of EtOH-metabolism on the Golgi; however, the link between this and abnormal glycosylation remains unknown. Because previously we have shown that Golgi morphology dictates glycosylation, we examined the effect EtOH administration has on function of Golgi residential enzymes involved in N-glycosylation...
October 17, 2016: Alcoholism, Clinical and Experimental Research
Bin Lin, Heng Yu, Yuting Lin, Chao Cai, Huoquan Lu, Xinbo Zhu
The aim of the present study was to assess the neuroprotective effects of tetrahydrocurcumin (THC) in a mouse model of cerebral ischemia/reperfusion (I/R) injury, and to investigate the involvement of Golgi reassembly and stacking protein 65 (GRASP65) and the extracellular signal‑regulated kinase (ERK) signaling pathway. Cerebral I/R injury was induced using the Pulsinelli four‑vessel occlusion method. After 5 min of reperfusion, mice received THC (5, 10 or 25 mg/kg) or saline by intraperitoneal injection...
October 6, 2016: Molecular Medicine Reports
Jesús M Gómez-Salinero, Marina M López-Olañeta, Paula Ortiz-Sánchez, Javier Larrasa-Alonso, Alberto Gatto, Leanne E Felkin, Paul J R Barton, Inmaculada Navarro-Lérida, Miguel Ángel Del Pozo, Pablo García-Pavía, Balaji Sundararaman, Giovanna Giovinazo, Gene W Yeo, Enrique Lara-Pezzi
Embryonic stem cells (ESC) have the potential to generate all the cell lineages that form the body. However, the molecular mechanisms underlying ESC differentiation and especially the role of alternative splicing in this process remain poorly understood. Here, we show that the alternative splicing regulator MBNL1 promotes generation of the atypical calcineurin Aβ variant CnAβ1 in mouse ESCs (mESC). CnAβ1 has a unique C-terminal domain that drives its localization mainly to the Golgi apparatus by interacting with Cog8...
September 28, 2016: Cell Chemical Biology
Manuela Viola, Kathrin Brüggemann, Evgenia Karousou, Ilaria Caon, Elena Caravà, Davide Vigetti, Burkhard Greve, Christian Stock, Giancarlo De Luca, Alberto Passi, Martin Götte
Proteoglycans and glycosaminoglycans modulate numerous cellular processes relevant to tumour progression, including cell proliferation, cell-matrix interactions, cell motility and invasive growth. Among the glycosaminoglycans with a well-documented role in tumour progression are heparan sulphate, chondroitin/dermatan sulphate and hyaluronic acid/hyaluronan. While the mode of biosynthesis differs for sulphated glycosaminoglycans, which are synthesised in the ER and Golgi compartments, and hyaluronan, which is synthesized at the plasma membrane, these polysaccharides partially compete for common substrates...
October 15, 2016: Glycoconjugate Journal
Mohammad A Al-Qudah, Ahmed Al-Dwairi
Neurotrophins are secreted proteins that are synthesized as pre-pro-neurotrophins on the rough endoplasmic reticulum, which are subsequently processed and then secreted as mature proteins. During synthesis, neurotrophins are sorted in the trans-Golgi apparatus into 2 pathways of secretion; the constitutive and the regulated pathways. Neurotrophins in the constitutive pathway are secreted cautiously without any trigger, while in the regulated pathway of secretion an external stimulus elevates the calcium concentration intracellularly leading to neurotrophin release...
October 2016: Neurosciences: the Official Journal of the Pan Arab Union of Neurological Sciences
Longbin Zheng, Ying Li, Xuesong Li, Jiayuan Kou, Zhaoyu Zhong, Yueqing Jiang, Zhongni Liu, Ye Tian, Liming Yang
BACKGROUND/AIMS: Sonodynamic therapy (SDT) is considered a new approach for the treatment of atherosclerosis. We previously confirmed that hydroxyl acetylated curcumin (HAC) was a sonosensitizer. In this study, we investigated the mechanism of THP-1 macrophage apoptosis and autophagy induced by HAC mediated SDT (HAC-SDT). METHODS: Cell viability was measured using a CCK-8 assay. Laser scanning confocal microscopy was used to measure the levels of intracellular reactive oxygen species (ROS), sub-cellular HAC localization, BAX and cytochrome C translocation, LC3 expression, monodansylcadaverine staining and Dil-labeled oxidized low density lipoprotein (Dil-ox-LDL) uptake...
October 17, 2016: Cellular Physiology and Biochemistry
Tomokazu Kimizu, Yukitoshi Takahashi, Taikan Oboshi, Asako Horino, Takayoshi Koike, Shinsaku Yoshitomi, Tatsuo Mori, Tokito Yamaguchi, Hiroko Ikeda, Nobuhiko Okamoto, Mitsuko Nakashima, Hirotomo Saitsu, Mitsuhiro Kato, Naomichi Matsumoto, Katsumi Imai
INTRODUCTION: Mutations of SLC35A2 that encodes Golgi-localized Uridine diphosphate (UDP)-galactose transporter at Xp11.23 lead to congenital disorders of glycosylation (CDG). Although patients with CDG generally have diverse systemic symptoms, patients with a SLC35A2 mutation manifest predominantly disorders of the central nervous system (CNS). CASE REPORT: A female infant aged 12months was referred to our center because of intractable seizures. The patient was born with birth weight of 3228g after 40weeks of unremarkable gestation...
October 12, 2016: Brain & Development
Yusuke Matsuda, Noriko Kimura, Takanobu Yoshimoto, Yoshihiro Sekiguchi, Junzo Tomoishi, Ichiro Kasahara, Yoshihito Hara, Yoshihiro Ogawa
Pheochromocytomas and paragangliomas, which exclusively produce dopamine, are very rare. Herein, we report for the first time a Japanese case of an exclusively dopamine-producing paraganglioma accompanied by detailed immunohistochemical analyses. A 70-year-old Japanese woman was referred to our hospital for functional examination of her left retroperitoneal mass. Her adrenal functions were normal, except for excessive dopamine secretion. After the tumorectomy, her dopamine level normalized. The histopathological diagnosis of the tumor was paraganglioma; this was confirmed by positive immunostaining of chromogranin A (CgA), tyrosine hydroxylase (TH), dopamine β-hydroxylase (DBH), and succinate dehydrogenase gene subunit B (SDHB)...
October 14, 2016: Endocrine Pathology
Ye Cao, Kai Guan, Xiang He, Congwen Wei, Zirui Zheng, Yanhong Zhang, Shengli Ma, Hui Zhong, Wei Shi
The Yersinia outer protein J (YopJ) plays a pivotal role in evading the host immune response and establishes a persistent infection in host cells after bacterial infection. YopJ is a cysteine protease and can act as a deubiquitinating enzyme that deubiquitinates several targets in multiple signaling pathways. Stimulator of interferon genes (STING) is a critical adapter for the induction of interferon regulatory factor 3 (IRF3) phosphorylation and subsequent production of the cytokines in response to nucleic acids in the cytoplasm...
October 11, 2016: Biochimica et Biophysica Acta
Z Schultzhaus, T B Johnson, B D Shaw
Cell growth necessitates extensive membrane remodeling events including vesicle fusion or fission, processes that are regulated by coat proteins. The hyphal cells of filamentous fungi concentrate both exocytosis and endocytosis at the apex. This investigation focuses on clathrin in Aspergillus nidulans, with the aim of understanding its role in membrane remodeling in growing hyphae. We examined clathrin heavy chain (ClaH-GFP) which localized to three distinct subcellular structures: late Golgi (trans-Golgi equivalents of filamentous fungi), which are concentrated just behind the hyphal tip but are intermittently present throughout all hyphal cells; the region of concentrated endocytosis just behind the hyphal apex (the "endocytic collar"); and small, rapidly moving puncta that were seen trafficking long distances in nearly all hyphal compartments...
October 14, 2016: Molecular Microbiology
Graziella Anselmo Joanitti, Rayane Ganassin, Mosar Rodrigues Corrêa, João Paulo Figueiró Longo, Cheng-Shi Jiang, Jinsong Gu, Sandra Milena Leal Pinto, Maria de Fátima Menezes Almeida Dos Santos, Ricardo Bentes de Azevedo, Luis Alexandre Muehlmann
Nanotechnology has provided powerful tools to improve the chemotherapy of cancer. Different nanostructures have been developed which deliver the anticancer drugs more selectively to tumor than to healthy tissues. The result has generally been the increase in efficacy and safety of classical anticancer drugs. In recent years, several studies have focused not only on the delivery of anticancer drugs to tumors, but also on delivering the drugs to specific organelles of cancer cells. Endoplasmic reticulum, Golgi apparatus, lysosomes, mitochondria, and nucleus have been the targets of different nanostructured drug delivery systems developed with the goal of circumventing drug-resistance, increasing drug efficacy, and so on...
October 13, 2016: Mini Reviews in Medicinal Chemistry
Julien Karim Malet, Pascale Cossart, David Ribet
Bacterial pathogens can interfere during infection with host cell organelles, such as mitochondria, the endoplasmic reticulum-Golgi system or nuclei. As important cellular functions are often compartmentalized in these organelles, their targeting allows pathogens to manipulate key host functions during infection. Here, we identify lysosomes as a new class of organelles targeted by the pathogenic bacterium Listeria monocytogenes. We demonstrate that extracellular Listeria, via secretion of the pore-forming toxin Listeriolysin O (LLO), alter lysosomal integrity in epithelial cells, but not in macrophages...
October 14, 2016: Cellular Microbiology
Karin Förster-Fromme, Sarah Schneider, Georg A Sprenger, Christoph Albermann
OBJECTIVES: To investigate the translocation of nucleotide-activated sugars from the cytosol across a membrane into the endoplasmatic reticulum or the Golgi apparatus which is an important step in the synthesis of glycoproteins and glycolipids in eukaryotes. RESULTS: The heterologous expression of the recombinant and codon-adapted human GDP-L-fucose antiporter gene SLC35C1 (encoding an N-terminal OmpA-signal sequence) led to a functional transporter protein located in the cytoplasmic membrane of Escherichia coli...
October 13, 2016: Biotechnology Letters
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