Subcellular Proteomics

As proteomics technologies are reaching a plateau in the number of proteins that can beresolvedanddetected, pre-fractionationstepshavebecomeessentialtoincreasethe depth of proteomic analysis. So far, many pre-fractionation steps have been based on chromatography methods where the proteins are separated according to their indiv- ual physicochemical properties. Subcellular fractionation methods proved to be very potentproteinpre-fractionationsteps: theyallowtherepresentationoflowabundance proteins, andtheycanbecombinedwithchromatographysteps. Moreover, asthei- latedsubcellularcomponentsalsorepresentfunctionalunits, subcellularfractionation allows the proteomic analysis of protein subsets that are functionally related in a b- logically relevant manner. The first three sections of this volume deal with different levels of subcellular organization that also correspond to specific methodological approaches. In his keynote chapter, Thierry Rabilloud superbly introduces the first section with athoroughdefinitionofmembraneproteomicswherehepinpointskeytheoreticaland practicalissuesofthisfield, therebysettingthestageforthenextcontributions.Miguet et al. address the first key issue: the quality of the membrane preparation; they int- duce and validate a microparticle strategy for plasma membrane purification. Zahedi et al. deal with the second major issue, the resolving of the hydrophobic proteins found in biological membrane samples, which they solve through two-dimensional BAC/SDS-PAGE gel electrophoresis. To close the first section, Foster and Chan review the proteomics of lipid rafts, membrane structures that are involved in int- cellular trafficking and signal transduction. They describe a clever validation scheme based on the sensitivity of lipid rafts to cholesterol disruption. Acentral theme in the second section on organelle subproteomes is the variability of their composition and how it can be exploited and interpreted. Kavanagh et