Proteomic analysis of lipid raft-enriched membranes isolated from internal organelles.
The mitochondria-associated membrane (MAM) is a sub-region of the endoplasmic reticulum (ER) that facilitates crosstalk between the ER and mitochondria. The MAM actively influences vital cellular processes including Ca(2+) signaling and protein folding. Detergent-resistant microdomains (DRMs) may localize proteins to the mitochondria/MAM interface to coordinate these events. However, the protein composition of DRMs isolated from this region is not known. Lipid-raft enriched DRMs were isolated from a combined mitochondria/MAM sample and analyzed using two-dimensional reversed-phased tandem mass spectrometry. Strict post-acquisition filtering of the acquired data led to the confident identification 250 DRM proteins. The majority (58%) of the identified proteins are bona fide mitochondrial or ER proteins according to Gene Ontology annotation. Additionally, 74% of the proteins have previously been noted as MAM-resident or -associated proteins. Furthermore, ∼20% of the identified proteins have a documented association with lipid rafts. Most importantly, known internal LR marker proteins (inositol 1,4,5-trisphosphate receptor type 3, erlin-2, and voltage-dependent anion channel 1) were detected as well as most of the components of the mitochondrial/MAM-localized Ca(2+) signaling complex. Our study provides the basis for future work probing how the protein activities at the mitochondrion/MAM interface are dependent upon the integrity of these internal lipid-raft-like domains.
Brown University, Department of Chemistry, Providence, RI 02912, USA.
Chloe N
CN
Poston
Ellen
E
Duong
Yuan
Y
Cao
Carthene R
CR
Bazemore-Walker
D000818:Animals; D002460:Cell Line; D003902:Detergents / Q000737:chemistry; D021962:Membrane Microdomains / Q000737:chemistry / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D051336:Mitochondrial Membranes / Q000737:chemistry / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D040901:Proteomics; D051381:Rats
D003902:Detergents; D008565:Membrane Proteins; D024101:Mitochondrial Proteins