Quantitative proteomics analysis of the Arg/N-end rule pathway of targeted degradation in Arabidopsis roots

ABSTRACT: According to the Arg/N-end rule pathway, proteins with basic N-termini are targeted for degradation by the Arabidopsis thaliana E3 ligase, PROTEOLYSIS6 (PRT6). Proteins can also become PRT6 substrates following post-translational arginylation by arginyltransferases ATE1 and 2. Here, we undertook a quantitative proteomics study of Arg/N-end rule mutants, ate1/2 and prt6, to investigate the impact of this pathway on the root proteome. Tandem mass tag (TMT) labelling identified a small number of proteins with increased abundance in the mutants, some of which represent downstream targets of transcription factors known to be N-end rule substrates. Isolation of N-terminal peptides using terminal amine isotope labelling of samples (TAILS) combined with triple dimethyl labelling identified 1465 unique N-termini. Stabilising residues were over-represented among the free neo-N-termini, but destabilising residues were not markedly enriched in N-end rule mutants. The majority of free neo-N-termini were revealed following cleavage of organellar targeting signals, thus compartmentation may account in part for the presence of destabilising residues in the wild type N-terminome. Our data suggest that PRT6 does not have a marked impact on the global proteome of Arabidopsis roots and is likely involved in the controlled degradation of relatively few regulatory proteins.

INSTRUMENT(S):

ORGANISM(S): Arabidopsis Thaliana (mouse-ear Cress)

TISSUE(S): Plant Cell, Root

SUBMITTER: Hongtao Zhang  

LAB HEAD: Frederica L. Theodoulou

PROVIDER: PXD001719 | Pride | 2015-03-26

REPOSITORIES: Pride