title: Characterization of intracellular aggresomes by fluorescent microscopy.
abstract: Correct folding of newly synthesized proteins is essential to cellular homeostasis and cells have evolved sophisticated means to fold and modify proteins. When misfolding occurs, the misfolded proteins often expose normally buried hydrophobic domains, causing localized aggregation. Individual small aggregates appear to be transported towards the microtubule-organizing center and there coalesce to form larger aggregates called aggresomes. Both cytoplasmic and nuclear proteins can form aggresomes. The study of aggresomes has progressed rapidly because numerous human diseases such as Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, various myopathies, and prion disease are characterized by the formation of aggresomes. Importantly, aggresomes sequester many cellular proteins and the pathology of aggresomal disease is at least partially caused by the deregulation of cellular components. Thus, it is essential to identify and characterize the composition of aggresomes formed by different proteins. However, most protein aggregates are insoluble even in buffers with high concentration of detergent, which makes them very difficult to analyze by biochemical approaches. An alternative approach that has been used successfully is the in situ characterization of protein components within aggresomes by immunofluorescent microscopy. Here, we provide detailed protocols to study the characteristic features of aggresomes by fluorescent microscopy.
authors.affiliation: Department of Cell, Developmental and Integrative Biology, Gregory Fleming James Cystic Fibrosis Research Center, University of Alabama at Birmingham, MCLM 331, 1918 University Boulevard, Birmingham, AL, 35294, USA, lianwufu@uab.edu.
authors.forename: Lianwu
authors.initials: L
authors.lastname: Fu
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Sztul
mesh_terms: D003593:Cytoplasm / Q000378:metabolism; D006801:Humans; D008856:Microscopy, Fluorescence / Q000379:methods; D022101:Microtubule-Organizing Center / Q000378:metabolism; D066329:Protein Aggregates / Q000502:physiology*; D017510:Protein Folding; D011506:Proteins / Q000378:metabolism*
chemical_list: D066329:Protein Aggregates; D011506:Proteins