14-3-3 genes encode a ubiquitous family of highly conserved eukaryotic proteins from fungi to humans and plants with several molecular and cellular functions. Most notably, 14-3-3 proteins bind to phosphoserine/phosphothreonine motifs in a sequence-specific manner. More than 100 14-3-3 binding partners involved in signal transduction, cell cycle regulation, apoptosis, stress responses, and malignant transformation have been identified. The 14-3-3 proteins form homodimers and heterodimers, and there is redundancy of the binding specificity and function of different 14-3-3 proteins because of their highly similar amino acid sequence and tertiary structure. 14-3-3 proteins can regulate target protein function by several mechanisms. Although the molecular and cellular functions of 14-3-3 proteins have been well studied, there have been fewer studies addressing the in vivo role of 14-3-3s. Here we review what is known about 14-3-3 proteins during eukaryotic development.