title: Imaging and quantification of amyloid fibrillation in the cell nucleus.
abstract: Xenobiotics, as well as intrinsic processes such as cellular aging, contribute to an environment that constantly challenges nuclear organization and function. While it becomes increasingly clear that proteasome-dependent proteolysis is a major player, the topology and molecular mechanisms of nuclear protein homeostasis remain largely unknown. We have shown previously that (1) proteasome-dependent protein degradation is organized in focal microenvironments throughout the nucleoplasm and (2) heavy metals as well as nanoparticles induce nuclear protein fibrillation with amyloid characteristics. Here, we describe methods to characterize the landscape of intranuclear amyloid on the global and local level in different systems such as cultures of mammalian cells and the soil nematode Caenorhabditis elegans. Application of discrete mathematics to imaging data is introduced as a tool to develop pattern recognition of intracellular protein fibrillation. Since stepwise fibrillation of otherwise soluble proteins to insoluble amyloid-like protein aggregates is a hallmark of neurodegenerative protein-misfolding disorders including Alzheimer's disease, CAG repeat diseases, and the prion encephalopathies, investigation of intracellular amyloid may likewise aid to a better understanding of the pathomechanisms involved. We consider aggregate profiling as an important experimental approach to determine if nuclear amyloid has toxic or protective roles in various disease processes.
authors.affiliation: IUF - Leibniz Research Institute of Environmental Medicine at Heinrich-Heine-University Duesseldorf, Auf'm Hennekamp 50, Duesseldorf, 40225, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Arnhold
authors.forename: Andrea
authors.initials: A
authors.lastname: Scharf
authors.forename: Anna
authors.initials: A
authors.lastname: von Mikecz
mesh_terms: D000682:Amyloid / Q000737:chemistry*; D000818:Animals; D052160:Benzothiazoles; D017173:Caenorhabditis elegans / Q000166:cytology / Q000187:drug effects; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism*; D003224:Congo Red / Q000378:metabolism; D006801:Humans; D008856:Microscopy, Fluorescence; D057054:Molecular Imaging / Q000379:methods*; D053758:Nanoparticles; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D012822:Silicon Dioxide / Q000737:chemistry / Q000494:pharmacology; D013194:Staining and Labeling; D013844:Thiazoles / Q000378:metabolism; D016707:Tissue Fixation
chemical_list: D000682:Amyloid; D052160:Benzothiazoles; D013844:Thiazoles; C009462:thioflavin T; D003224:Congo Red; D012822:Silicon Dioxide