Cell surface labeling and mass spectrometry reveal diversity of cell surface markers and signaling molecules expressed in undifferentiated mouse embryonic stem cells.

Although interactions between cell surface proteins and extracellular ligands are key to initiating embryonic stem cell differentiation to specific cell lineages, the plasma membrane protein components of these cells are largely unknown. We describe here a group of proteins expressed on the surface of the undifferentiated mouse embryonic stem cell line D3. These proteins were identified using a combination of cell surface labeling with biotin, subcellular fractionation of plasma membranes, and mass spectrometry-based protein identification technology.

Large-scale identification of proteins expressed in mouse embryonic stem cells.

A protein subset expressed in the mouse embryonic stem (ES) cell line, E14-1, was characterized by mass spectrometry-based protein identification technology and data analysis. In total, 1790 proteins including 365 potential nuclear and 260 membrane proteins were identified from tryptic digests of total cell lysates. The subset contained a variety of proteins in terms of physicochemical characteristics, subcellular localization, and biological function as defined by Gene Ontology annotation groups.

Analysis of small human proteins reveals the translation of upstream open reading frames of mRNAs.

To find novel short coding sequences from accumulated full-length cDNA sequences, proteomic analysis of small proteins expressed in human leukemia K562 cells was performed using high-resolution nanoflow liquid chromatography coupled with electrospray ionization tandem mass spectrometry. Our analysis led to the identification of 54 proteins not more than 100 amino acids in length, including four novel ones. These novel short coding sequences were all located upstream of the longest open reading frame (ORF) of the corresponding cDNA.

Mammalian phospholipase Czeta induces oocyte activation from the sperm perinuclear matrix.

Mammalian sperm-borne oocyte activating factor (SOAF) induces oocyte activation from a compartment that engages the oocyte cytoplasm, but it is not known how. A SOAF-containing extract (SE) was solubilized from the submembrane perinuclear matrix, a domain that enters the egg. SE initiated activation sufficient for full development.

Phosphorylation-dependent interaction of kinesin light chain 2 and the 14-3-3 protein.

The protein 14-3-3 is a key regulator in a cell signaling pathway mediated by protein phosphorylation. To identify the cellular targets of this protein systematically, we have employed a proteomic approach: protein components pulled down from PC12 cells stably expressing a myc-tagged 14-3-3eta isoform were analyzed by means of SDS-PAGE and mass spectrometry. This procedure allowed us to identify more than 30 proteins that include various known and unknown targets of the 14-3-3 protein.