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Silencing of mitochondrial Lon protease deeply impairs mitochondrial proteome and function in colon cancer cells
Author(s) -
Gibellini Lara,
Pinti Marcello,
Boraldi Federica,
Giorgio Valentina,
Bernardi Paolo,
Bartolomeo Regina,
Nasi Milena,
De Biasi Sara,
Missiroli Sonia,
Carnevale Gianluca,
Losi Lorena,
Tesei Anna,
Pinton Paolo,
Quaglino Daniela,
Cossarizza Andrea
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.14-255869
Subject(s) - mitochondrion , gene silencing , oxidative phosphorylation , microbiology and biotechnology , oxidative stress , biology , chemistry , mitochondrial matrix , mitochondrial dna , biochemistry , gene , cytosol , enzyme
Lon is a nuclear‐encoded, mitochondrial protease that assists protein folding, degrades oxidized/damaged proteins, and participates in maintaining mtDNA levels. Here we show that Lon is up‐regulated in several human cancers and that its silencing in RKO colon cancer cells causes profound alterations of mitochondrial proteome and function, and cell death. We silenced Lon in RKO cells by constitutive or inducible expression of Lon shRNA. Lon‐silenced cells displayed altered levels of 39 mitochondrial proteins (26% related to stress response, 14.8% to ribosome assembly, 12.7% to oxidative phosphorylation, 8.5% to Krebs cycle, 6.3% to β‐oxidation, and 14.7% to crista integrity, ketone body catabolism, and mtDNA maintenance), low levels of mtDNA transcripts, and reduced levels of oxidative phosphorylation complexes (with >90% reduction of complex I). Oxygen consumption rate decreased 7.5‐fold in basal conditions, and ATP synthesis dropped from 0.25 ± 0.04 to 0.03 ± 0.001 nmol/mg proteins, in the presence of 2‐deoxy‐D‐glucose. Hydrogen peroxide and mitochondrial superoxide anion levels increased by 3‐ and 1.3‐fold, respectively. Mitochondria appeared fragmented, heterogeneous in size and shape, with dilated cristae, vacuoles, and electrondense inclusions. The triterpenoid 2‐cyano‐3,12‐dioxooleana‐1,9,‐dien‐28‐oic acid, a Lon inhibitor, partially mimics Lon silencing. In summary, Lon is essential for maintaining mitochondrial shape and function, and for survival of RKO cells.—Gibellini, L., Pinti, M., Boraldi, F., Giorgio, V., Bernardi, P., Bartolomeo, R., Nasi, M., De Biasi, S., Missiroli, S., Carnevale, G., Losi, L., Tesei, A., Pinton, P., Quaglino, D., Cossarizza, A., Silencing of mitochondrial Lon protease deeply impairs mitochondrial proteome and function in colon cancer cells. FASEB J. 28, 5122–5135 (2014). www.fasebj.org