For a long time it has been assumed that the decay of RNA in eukaryotes is mainly carried out by exoribonucleases, which is in contrast to bacteria, where endoribonucleases are well documented to initiate RNA degradation. In recent years, several as yet unknown endonucleases have been described, which has changed our view on eukaryotic RNA metabolism. Most importantly, it was shown that the primary eukaryotic 3′ –> 5′ exonuclease, the exosome complex has the ability to endonucleolytically cleave its physiological RNA substrates, and novel endonucleases involved in both nuclear and cytoplasmic RNA surveillance pathways were discovered concurrently. In addition, endoribonucleases responsible for long-known processing steps in the maturation pathways of various RNA classes were recently identified. Moreover, one of the most intensely studied RNA decay pathways–RNAi–is controlled and stimulated by the action of different endonucleases. Furthermore, endoribonucleolytic cleavages executed by various enzymes are also the hallmark of RNA degradation and processing in plant chloroplasts. Finally, multiple context-specific endoribonucleases control qualitative and/or quantitative changes of selected transcripts under particular conditions in different eukaryotic organisms. The aim of this review is to discuss the impact of all of these discoveries on our current understanding of eukaryotic RNA metabolism.