Analysis of human cytomegalovirus secondary envelopment by advanced electron microscopy. 2013

Martin Schauflinger, and Clarissa Villinger, and Thomas Mertens, and Paul Walther, and Jens von Einem
Institute of Virology, University Medical Center Ulm, Albert-Einstein-Allee, 89081 Ulm, Germany.

Electron microscopy (EM) allows visualization of viruses in fixed cells with high resolution. High-pressure freezing for sample fixation in combination with freeze substitution and embedding in resin improves significantly the preservation of cellular structures and specifically of membranes. This advancement allows better visualization of human cytomegalovirus (HCMV) morphogenesis occurring at membranes. To obtain comprehensive information on viral phenotypes from ultrastructural images it is important to also quantify morphological phenotypes. This again can be much refined by three-dimensional visualization after serial sectioning. For elucidation of dynamic processes three-dimensional tomography is extremely helpful. We analysed interaction of HCMV particles with host cell membranes during final envelopment. Both wild-type virus and a viral mutant with impaired envelopment were analysed in fibroblasts, but also using in vivo relevant human endothelial cells and macrophages. The quantification of the EM data showed similar ultrastructural phenotypes regarding the envelopment efficiency in the different cell types indicating similar mechanisms in late stages of virus morphogenesis. Furthermore, thorough analysis of the viral assembly complex (AC) - a virus-induced cytosolic structure - by using three-dimensional visualization techniques combined with a quantitative analysis revealed that the events of final envelopment are equally distributed within the AC irrespective of different local membrane composition.

UI MeSH Term Description Entries
D008264 Macrophages The relatively long-lived phagocytic cell of mammalian tissues that are derived from blood MONOCYTES. Main types are PERITONEAL MACROPHAGES; ALVEOLAR MACROPHAGES; HISTIOCYTES; KUPFFER CELLS of the liver; and OSTEOCLASTS. They may further differentiate within chronic inflammatory lesions to EPITHELIOID CELLS or may fuse to form FOREIGN BODY GIANT CELLS or LANGHANS GIANT CELLS. (from The Dictionary of Cell Biology, Lackie and Dow, 3rd ed.) Bone Marrow-Derived Macrophages,Monocyte-Derived Macrophages,Macrophage,Macrophages, Monocyte-Derived,Bone Marrow Derived Macrophages,Bone Marrow-Derived Macrophage,Macrophage, Bone Marrow-Derived,Macrophage, Monocyte-Derived,Macrophages, Bone Marrow-Derived,Macrophages, Monocyte Derived,Monocyte Derived Macrophages,Monocyte-Derived Macrophage
D008854 Microscopy, Electron Microscopy using an electron beam, instead of light, to visualize the sample, thereby allowing much greater magnification. The interactions of ELECTRONS with specimens are used to provide information about the fine structure of that specimen. In TRANSMISSION ELECTRON MICROSCOPY the reactions of the electrons that are transmitted through the specimen are imaged. In SCANNING ELECTRON MICROSCOPY an electron beam falls at a non-normal angle on the specimen and the image is derived from the reactions occurring above the plane of the specimen. Electron Microscopy
D008867 Microtomy The technique of using a microtome to cut thin or ultrathin sections of tissues embedded in a supporting substance. The microtome is an instrument that hold a steel, glass or diamond knife in clamps at an angle to the blocks of prepared tissues, which it cuts in sections of equal thickness. Thin Sectioning,Ultramicrotomy,Sectioning, Thin,Sectionings, Thin,Thin Sectionings
D009154 Mutation Any detectable and heritable change in the genetic material that causes a change in the GENOTYPE and which is transmitted to daughter cells and to succeeding generations. Mutations
D002478 Cells, Cultured Cells propagated in vitro in special media conducive to their growth. Cultured cells are used to study developmental, morphologic, metabolic, physiologic, and genetic processes, among others. Cultured Cells,Cell, Cultured,Cultured Cell
D003587 Cytomegalovirus A genus of the family HERPESVIRIDAE, subfamily BETAHERPESVIRINAE, infecting the salivary glands, liver, spleen, lungs, eyes, and other organs, in which they produce characteristically enlarged cells with intranuclear inclusions. Infection with Cytomegalovirus is also seen as an opportunistic infection in AIDS. Herpesvirus 5, Human,Human Herpesvirus 5,Salivary Gland Viruses,HHV 5,Herpesvirus 5 (beta), Human,Cytomegaloviruses,Salivary Gland Virus,Virus, Salivary Gland,Viruses, Salivary Gland
D005347 Fibroblasts Connective tissue cells which secrete an extracellular matrix rich in collagen and other macromolecules. Fibroblast
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D014764 Viral Proteins Proteins found in any species of virus. Gene Products, Viral,Viral Gene Products,Viral Gene Proteins,Viral Protein,Protein, Viral,Proteins, Viral
D016610 Tissue Embedding The technique of placing cells or tissue in a supporting medium so that thin sections can be cut using a microtome. The medium can be paraffin wax (PARAFFIN EMBEDDING) or plastics (PLASTIC EMBEDDING) such as epoxy resins. Embedding, Tissue

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