Pieroni 2017 MiP2017
Toward the standardization of mitochondrial proteomics. Pieroni_Presentation
The mitochondrial Human Proteome Project aims at understanding the function of the mitochondrial proteome and its crosstalk with the proteome of other organelles. Being able to choose a suitable and validated enrichment protocol of functional mitochondria, based on the specific needs of the downstream proteomics analysis, would greatly help the researchers in the field.
Nineteen labs were involved. Ten cell lines were selected. Three methods for mitochondrial isolation were chosen (differential centrifugation, sucrose gradient separation, a commercial kit). Samples prepared with two different methods from the same cell line reached the same mass spectrometry lab. Proteins were selected if observed in at least two out of three biological replicates and further annotated as mitochondrial proteins (Mitocarta and IMPI). Consensus lists were mapped on the reference network for the functional mitochondrial proteome, a network composed of 6592 nodes (1361 of them labelled as “mitochondrial” in the neXtProt database and 5231 “mitochondrial associated” interactors in the IM X consortium) and 16350 edges.
The overall number of identified total and mitochondrial proteins was not significantly dependent on the enrichment protocol but specific line to line differences were observed. More than 80% of the identified proteins resulted to be nodes of the network representing the functional mitochondrial proteome, but with a different ability in co-isolating mitochondria-associated structures foreach enrichment protocol/cell line pair.
No enrichment method is preferable as such. However, a given protocol would be best suited for a certain cell type and for specific enrichment purposes (e.g., true mitochondrial proteins vs. mitochondria-associated proteins).
• Bioblast editor: Kandolf G
Preparation: Isolated mitochondria
- Pieroni L(1), Alberio T(2), Ronci M(3), Roncada P(4), Urbani A(5), Fasano M(2)
- the Italian mt-HPP Consortium.
- Santa Lucia Foundation, Rome
- Univ Insubria, Busto Arsizio
- Univ Chieti-Pescara, Chieti
- “L. Spallanzani” Inst, Rivolta d’Adda
- Catholic Univ Sacred Hearth, Rome; Italy. - [email protected]
- Alberio T, Pieroni L, Ronci M, Banfi C, Bongarzone I, Bottoni P, Brioschi M, Caterino M, Chinello C, Cormio A, Cozzolino F, Cunsolo V, Fontana S, Garavaglia B, Giusti L, Greco V, Lucacchini A, Maffioli E, Magni F, Monteleone F, Monti M, Monti V, Musicco C, Petrosillo G, Porcelli V, Saletti R, Scatena R, Soggiu A, Tedeschi G, Zilocchi M, Roncada P, Urbani A, Fasano M (2017) Toward the standardization of mitochondrial proteomics. The Italian mt-HPP Initiative. J Proteome Res 16:4319-29.