Peroxisome Dynamics in Cultured Mammalian Cells

• Published in: Traffic (Copenhagen, Denmark) Vol. 10; no. 11; pp. 1722 - 1733
• Main Authors: Huybrechts, Sofie J, Van Veldhoven, Paul P, Brees, Chantal, Mannaerts, Guy P, Los, Georgyi V, Fransen, Marc
• Format: Journal Article
• Language: English
• Published: Oxford, UK Oxford, UK : Blackwell Publishing Ltd 01-11-2009; Blackwell Publishing Ltd
• Subjects: Animals; autophagy; Autophagy - genetics; Biotinylation; Cell Fusion; Cells, Cultured; CHO Cells; Cricetinae; Cricetulus; de novo synthesis; Fluorescent Dyes - metabolism; Green Fluorescent Proteins - metabolism; in vivo pulse-labeling; Indoles - metabolism; Ligands; Mammals - genetics; Mammals - metabolism; Membrane Proteins - genetics; Membrane Proteins - metabolism; non-symmetric fission; peroxisomes; Peroxisomes - metabolism; Plasmids; protein trafficking; Protein Transport; Transfection; Xanthenes - metabolism
• ISSN: 1398-9219; 1600-0854
• DOI: 10.1111/j.1600-0854.2009.00970.x

Despite the identification and characterization of various proteins that are essential for peroxisome biogenesis, the origin and the turnover of peroxisomes are still unresolved critical issues. In this study, we used the HaloTag technology as a new approach to examine peroxisome dynamics in cultured mammalian cells. This technology is based on the formation of a covalent bond between the HaloTag protein-a mutated bacterial dehalogenase which is fused to the protein of interest-and a synthetic haloalkane ligand that contains a fluorophore or affinity tag. By using cell-permeable ligands of distinct fluorescence, it is possible to image distinct pools of newly synthesized proteins, generated from a single genetic HaloTag-containing construct, at different wavelengths. Here, we show that peroxisomes display an age-related heterogeneity with respect to their capacity to incorporate newly synthesized proteins. We also demonstrate that these organelles do not exchange their protein content. In addition, we present evidence that the matrix protein content of pre-existing peroxisomes is not evenly distributed over new organelles. Finally, we show that peroxisomes in cultured mammalian cells, under basal growth conditions, have a half-life of approximately 2 days and are mainly degraded by an autophagy-related mechanism. The implications of these findings are discussed.