In-depth characterization of apoptosis N-terminome reveals a link between caspase-3 cleavage and post-translational N-terminal acetylation

Abstract

The N-termini of proteins contain information about their biochemical properties and functions. These N-termini can be processed by proteases and can undergo other co- or post-translational modifications. We have developed LATE (LysN Amino Terminal Enrichment), a method that uses selective chemical derivatization of -amines to isolate the N-terminal peptides, in order to improve N-terminome identification in conjunction with other enrichment strategies. We applied LATE alongside another N-terminomic method to study caspase-3 mediated proteolysis both in vitro and during apoptosis in cells. This has enabled us to identify many unreported caspase-3 cleavages, some of which cannot be identified by other methods. Moreover, we have found direct evidence that neo-N-termini generated by caspase-3 cleavage can be further modified by Nt-acetylation. Some of these neo-Nt-acetylation events occur in the early phase of the apoptotic process and may have a role in translation inhibition. This has provided a comprehensive overview of the caspase-3 degradome and has uncovered previously unrecognized crosstalk between post-translational Nt-acetylation and caspase proteolytic pathways.