ABSTRACT
Autophagy (cellular self-eating) is a highly regulated degradation process of the eukaryotic cell during which parts of the cytoplasm are delivered into, and broken down within, the lysosomal compartment. The process serves as a main route for the elimination of superfluous and damaged cellular constituents, thereby mediating macromolecular and organellar turnover. In addition to maintaining cellular homeostasis, autophagy is involved in various other cellular and developmental processes by degrading specific regulatory proteins, and contributing to the clearance of intracellular pathogens. The physiological roles and pathological involvement of autophagy can be effectively studied in divergent eukaryotic model systems ranging from yeast to mice. Such a tractable animal modelapplied only recently for autophagy researchis the zebrafish Danio rerio, which also facilitates the analysis of more specific biological processes such as tissue regeneration. In this chapter, we overview the main methods and tools that are used to monitor autophagic structures and to assay autophagic responses in this vertebrate organism. We place emphasis on genetic (functional) approaches applied for exploring novel cellular and developmental roles of the autophagic process.
Subject(s)
Autophagy , Zebrafish/metabolism , Animals , Blotting, Western/methods , CRISPR-Cas Systems , Gene Expression Regulation , Gene Silencing , Microscopy, Electron, Transmission/methods , Microscopy, Fluorescence/methods , Mutagenesis , Polymerase Chain Reaction/methods , RNA, Messenger/genetics , Zebrafish/genetics , Zebrafish Proteins/genetics , Zebrafish Proteins/metabolismSubject(s)
Creatinine/urine , Diphosphonates/adverse effects , Hypocalcemia/chemically induced , Imidazoles/adverse effects , Immunosuppressive Agents/therapeutic use , Multiple Myeloma/drug therapy , Thalidomide/therapeutic use , Aged , Diphosphonates/therapeutic use , Female , Humans , Hypocalcemia/urine , Imidazoles/therapeutic use , Male , Multiple Myeloma/urine , Zoledronic AcidABSTRACT
CD39 is a human lymphoid cell activation antigen, (also referred to E-ATPDase or apyrase) that hydrolyzes extracellular ATP and ADP. Although it has been widely studied, its physiological role, however, still remains unclear. This ectonucleotidase generally is said to be evenly distributed in the membrane of the cells. However, we observed that in cell types which possess caveolae, specialised membrane invaginations involved in signalling, CD39 is preferentially targeted to these membrane microdomains. Since all molecules involved in signalling (eNOS, G-proteins, receptors) which are targeted to the caveolae undergo posttranslational modifications (e.g., palmitoylation) we hypothesize the same to be the case for CD39. Furthermore, its presence in the caveolae supports its participation in signalling events.
