Comparative large scale characterization of plant versus mammal proteins reveals similar and idiosyncratic N-α-acetylation features

Abstract

N-terminal modifications play a major role in the fate of proteins in terms of activity, stability, or subcellular compartmentalization. Such modifications remain poorly described and badly characterized in proteomic studies, and only a few comparison studies among organisms have been made available so far. Recent advances in the field now allow the enrichment and selection of N-terminal peptides in the course of proteome-wide mass spectrometry analyses. These targeted approaches unravel as a result the extent and nature of the protein N-terminal modifications. Here, we aimed at studying such modifications in the model plant Arabidopsis thaliana to compare these results with those obtained from a human sample analyzed in parallel. We applied large scale analysis to compile robust conclusions on both data sets. Our data show strong convergence of the characterized modifications especially for protein N-terminal methionine excision, co-translational N-α-acetylation, or N-myristoylation between animal and plant kingdoms. Because of the convergence of both the substrates and the N-α-acetylation machinery, it was possible to identify the N-acetyltransferases involved in such modifications for a small number of model plants. Finally, a high proportion of nuclear-encoded chloroplast proteins feature post-translational N-α-acetylation of the mature protein after removal of the transit peptide. Unlike animals, plants feature in a dedicated pathway for post-translational acetylation of organelle-targeted proteins. The corresponding machinery is yet to be discovered.

Fig. 1.

Phylogram of the catalytic subunits of the main Nats, i.e. NAA10 (NatA), NAA20 (NatB), NAA30 (NatC), NAA40 (NatD), and NAA50 (NatE), for various species. The orthologous sequences of the human genes were extracted using BLAST searches against D. rerio (DANRO), D. melanogaster (DRONE), P. trichocarpa (POPTR), C. reinhardtii (CHLRE), V. vinifera (VITVI), M. truncatula (MEDTR), and A. thaliana (ARATH) and have been added, as well as the S. solfataricus ortholog of NAA10 and RIMI from E. coli.

Fig. 2.

WebLogo of the N-terminome (sequence at positions 2–6) of A. thaliana translated genes product (TAIR10) (A) and the experimentally characterized NAAed proteins (B) (n = 777). Similar representation have been produced for the human subset of (C) Uniprot/Swiss-Prot protein (human subset) versus (D) experimentally characterized NAAed proteins (n = 648).

Fig. 3.

Sequence logos of the cTP cleavage sites related to the experimentally characterized dNAAed chloroplastic proteins. A, proteins with a single cTP cleavage site (n = 170). B, proteins with multiple cTP cleavage sites (n = 85). Each protein is represented with as many sequences as experimental cleavage sites characterized. C, IceLogo for the comparison of the sequences used in the two previous sequence logos, i.e. single (A) versus multiple (B) cleavage sites. cTP cleavage site occurs between residues −1 and +1.

Fig. 4.

Sequence logo of the cTP cleavage sites related to all experimentally characterized nuclear encoded chloroplastic proteins (n = 255) characterized with dNAA (cTP site position within 20–200) (A), chloroplast inner integral envelope proteins (n = 36) (B), plastid stroma proteins (n = 73) (C), and thylakoid proteins (n = 27) (D). cTP cleavage site occurs between residues −1 and +1.