Prion protein with Y145STOP mutation induces mitochondria-mediated apoptosis and PrP-containing deposits in vitro.

Abstract

A pathogenic truncation of an amber mutation at codon 145 (Y145STOP) in Gerstmann-Straussler-Scheinker disease (GSS) was investigated through the real-time imaging in living cells, by utilizing GFP-PrP constructs. GFP-PrP(1-144) exhibited an aberrant localization to mitochondria in mouse neuroblastoma neuro2a (N2a) and HpL3-4 cells, a hippocampal cell line established from prnp gene-ablated mice, whereas full-length GFP-PrP did not. The aberrant mitochondrial localization was also confirmed by Western blot analysis. Since GFP-PrP(1-121), as previously reported, and full-length GFP-PrP do not exhibit such mitochondrial localization, the mitochondrial localization of GFP-PrP(1-144) requires not only PrP residues 121-144 (in human sequence) but also COOH-terminal truncation in the current experimental condition. Subsequently, the GFP-PrP(1-144) induced a change in the mitochondrial innermembrane potential (DeltaPsi(m)), release of cytochrome c from the intermembrane space into the cytosol, and DNA fragmentation in these cells. Non-fluorescent PrP(1-144) also induced the DNA fragmentation in N2a and HpL3-4 cells after the proteasomal inhibition. These data may provide clues as to the molecular mechanism of the neurotoxic property of Y145STOP mutation. Furthermore, immunoelectron microscopy revealed numerous electron-dense deposits in mitochondria clusters of GFP-PrP(1-144)-transfected N2a cells, whereas no deposit was detected in the cells transfected with full-length GFP-PrP. Co-localization of GFP/PrP-immunogold particles with porin-immunogold particles as a mitochondrial marker was observed in such electron-dense vesicular foci, resembling those found in autophagic vacuoles forming secondary lysosomes. Whether such electron-dense deposits may serve as a seed for the growth of amyloid plaques, a characteristic feature of GSS with Y145STOP, awaits further investigations.

• Authors: Naomi S Hachiya, Kota Watanabe, Makiko Y Kawabata, Akiko Jozuka, Yoshimichi Kozuka, Yuji Sakasegawa, Kiyotoshi Kaneko
• Journal: Biochemical and biophysical research communications (Biochem Biophys Res Commun) Vol. 327 Issue 3 Pg. 894-9 (Feb 18 2005) ISSN: 0006-291X [Print] United States
• PMID: 15649429 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Codon
• Neuroprotective Agents
• Prions
• Cytochromes c
• DNA

Topics

• Animals
• Apoptosis
• Blotting, Western
• Codon (genetics)
• Cytochromes c (metabolism)
• Cytosol (metabolism)
• DNA (genetics, metabolism)
• Gerstmann-Straussler-Scheinker Disease (genetics, metabolism)
• Lysosomes (metabolism)
• Mice
• Mitochondria (physiology)
• Mutation
• Neuroblastoma (metabolism)
• Neurons (cytology, metabolism)
• Neuroprotective Agents (pharmacology)
• Prions (chemistry, genetics, metabolism, physiology)