Proteins are fundamental constituents of all living organisms. Mammalian genomes contain thousands of protein coding genes (~20,000 in human). A great majority of proteins in eukaryotic cells are modified by different posttranslational modification (PTMs). PTMs are key constituents of cell signaling networks, comprising some of the most sophisticated control switches in the cell. Given the vast complexity and their all essential role in controlling biology of living cells, analysis of proteins and PTMs is one of the most exiting areas of research in biology. Recent developments in high-resolution mass spectrometry (MS), computational proteomics, and optimized PTM-enrichment strategies have greatly facilitated unbiased (non-hypothesis driven) analysis of posttranslational modifications. We have applied these approaches to investigate dynamics of three major PTMs - phosphorylation, acetylation and ubiquitylation. Large-scale analysis of these PTMs provided systems view of signaling networks, and revealed their extensive involvement in diverse biological processes. I will discuss these recent developments in the MS- based PTM analysis, and highlight their applications in quantification of posttranslational modifications in cell signaling networks.