Despite the importance of phosphorylation-dependent signaling to cellular physiology, a comprehensive understanding of the phosphorylation networks and pathways governing cell signaling in higher eukaryotes is still lacking. Here, we developed a combined bioinformatics and protein microarray-based strategy to construct a high-resolution map of the human phosphorylation networks. To accomplish this task, we performed phosphorylation reactions with 289 human kinases on protein microarrays composed of 4,191 unique, full-length human proteins. We identified 3,656 kinase-substrate relationships that are likely to be physiologically relevant. We also determined phosphorylation motifs for 230 kinases and used these data to explore the complexity and diversity of motifs in the various kinase families. By combining these data with other resources, we obtained a high-resolution phosphorylation network, which connects the specific phosphorylation sites present in substrates with their upstream kinases.
In addition, we also developed several web tools to faciliate the research in this field.