Introduction 1 1 1 1996 2000 1992 2003 1 Fig. 1 a b c arrowhead points to a coated bud d arrows  2001 2006 1 1 1 Table 1 Assessment of published endoplasmic reticulum proteomics datasets Organelle Tissue/cells Species ER purification Mass spectrometry Proteins detected Reference NE Neuroblastoma N2a cells Mouse NE preparation (Triton-X-100 treatment) 2D-BAC gels, MALDI MS 148 2001 NE Liver Mouse Substractive proteomics (NE fraction-MM fraction) MudPIT, LCQ-Deca ion-trap MS, Tandem MS 566 2003 NE Liver Rat Nuclear pore complex fraction (enriched in nucleoporins)  1D gels, MALDI-QqTOF MS, Tandem MS  94 2002 NE Saccharomyces cerevisiae Yeast Purified nuclear pore complex HPLC, 1D gels, MALDI-TOF MS, Tandem MS 174 2000 a Liver b Membrane proteins 1D and 2D gels, MALDI-TOF MS  2002 a Liver c j 2D gels, MALDI-Q-TOF MS 39 2005 d Liver Mouse PCP-fraction co-sedimenting with calnexin LC, linear ion-trap Fourier transform MS, Tandem MS  229 2006 RER Saccharomyces cerevisiae Yeast Purified ribosomes Multidimensional LC, LCQ ion-trap MS, Tandem MS 95 1999 RER Liver Mouse e 2D gels, MALDI-TOF MS, Tandem MS 141 2003 RER Pancreas Dog Ribosome-associated membrane protein Blue Native gels, LCQ ion- trap MS, Tandem MS 30 2005 RER  Liver Rat  j e, f 1D gels, LC, QTOF-2 MS, Tandem MS 787 2006 SER  Liver Rat  j e e, g 1D gels, LC, QTOF-2 MS, Tandem MS 998 2006 h i Human j 1D gels, LC, Tandem MS 24 2004 d Liver Mouse PCP-fraction co-sedimenting with p115 LC, linear ion-trap Fourier transform MS, Tandem MS 220 2006 BAC n ERGIC G6Pase HPLC IB LC MALDI MM MS MudPIT PCP RM SM TOF a b c d e f g h i j Protein synthesis and secretion The ER is a key organelle of the secretion pathway involved in the synthesis of both proteins and lipids destined for multiple sites within and without the cell. Ribosomal proteins 1999 Saccharomyces cerevisiae 2004 2006 1995 2006 2006 1996 2006 Proteins involved in RNA metabolism 2006 2006 2006 2002 2000 1996 Proteins mediating targeting, co-translational translocation, and processing of nascent polypeptide chains 1975 1993 1996 1999 2003 2005 2002 2006 2005 2005 2007 Proteins involved in glycosylation and the calnexin cycle 2005 N 1999 N 2006 Ubiquitin metabolizing enzymes N 2006 2003 2006 2006 2006 2003 2006 2006 2002 2006 2006 Biosynthetic cargo 2006 2002 2006 2003 2006 Proteins involved in cargo exit and membrane traffic 1975 2004 2006 2006 2006 1996 1999 2000 2006 2004 2006 2003 1996 ER chaperones 1975 2007 2003 2006 2006 2006 2006 2005 2003 Calcium-handling proteins 2+ 2+ 2+ 2+ 2+ 2003 2006 2+ 2+ 3 2+ 1997 Enzymes of lipid and glucose metabolism 2007 2005 2006 2006 2003 2006 2006 1994 2006 2006 2006 2006 2006 2001 Proteins of detoxification and drug protein targets of the ER 2007 2006 2002 2006 2002 2007 Ubiquitin metabolizing enzymes 2005 2005 1995 2004 2006 1996 2006 1995 2004 Proteins involved in antigen processing 2007 2006 2007 2006 2006 2006 1995 2001 2002 2006 2005 2003 2003 Cytoskeletal proteins 2005 1998 1994 1988 1994 2006 2006 2004 2006 2005 1997 2006 2004 2005 2006 1998 2005 2006 2006 2006 2005 2006 1998 Proteins involved in signaling 2006 2004 2004 2003 2004 2000 2003 2003 2007 2006 2006 2005 2006 2002 2007 2000 2003 Phosphoproteins associated with the ER 2000 2006 2006 2005 2005 2005 2006 2006 2002 2006 2006 2005 Conclusions and future perspectives From available proteomics data parts of which are supported by electron microscope protein localization studies some tentative conclusions can be drawn about the relative segregation of proteins and molecular machines in the subcompartments of the ER. CLIMP-63 and ribosomes are enriched in rER. Reticulon, enzymes involved in ubiquitination, the proteasome, some cytoskeletal proteins, proteins involved in antigen processing, and coat proteins are enriched in sER. In contrast, proteins that appear equally distributed between rER and sER include proteins of the translocon, biosynthetic cargo, chaperones, proteins of detoxification, and proteins involved in lipid and glucose metabolism. Because proteomics of ER subcompartments is carried out using subcellular fractions, there are limitations that have to be considered when trying to interpret the results of the protein analysis. For example, the relative purity of the fractions will vary and proteins of fragments of contaminating organelles will be present in the fraction and be identified in the analysis. Moreover molecular dynamics at the cytosolic and luminal surfaces of the ER have to be taken into consideration. The composition of the ER is subject to change based on molecular interactions occurring on both cytosolic and luminal sides of the ER membrane. On the cytosolic side of the ER interaction with the cytoskeleton (to change the location, shape or size of the ER) or interaction with signaling proteins (to activate specific signaling pathways, e.g., apoptosis) leads to transient associations between the ER and cytosolic proteins. On the luminal side of the ER interactions between newly synthesized proteins and chaperones can vary under specific conditions (for example during ER stress) and will affect the overall molecular composition. Up-regulation of proteins involved in detoxification may occur under exposure to toxic chemicals and affect ER composition. Thus depending on the cell physiology protein associations with the ER may vary. Dynamics of protein interactions in organelles are often controlled by posttranslational modifications including phosphorylation. Understanding such modifications is key to understanding site-specific protein function. Proteomics studies of the subcompartments of the ER have lead to insights into the function of the different compartments of this organelle and new paradigms. However data obtained using proteomics analysis should be complimented by cytological techniques to confirm the localization of the proteins in the ER subcompartments and molecular biology should be used to modulate protein expression to examine the function. The combination of these approaches not only will yield new information about the proteins but they will also expand knowledge on the protein families to which they belong and/or protein complexes of which they are part of.