Determinants of specificity in two-component signal transduction

► Two-component signaling pathways are highly specific, exhibiting minimal cross-talk. ► Specificity stems from molecular recognition, phosphatases, and substrate competition. ► Specificity residues coevolve following a duplication to insulate the new pathways. ► Avoiding cross-talk is a major selective pressure acting on two-component proteins. ► Tethering a kinase to a substrate relaxes pressure to diversify specificity residues.