Plastid Proteome Assembly without Toc159: Photosynthetic Protein Import and Accumulation of N-Acetylated Plastid Precursor Proteins  

Abstract

Import of nuclear-encoded precursor proteins from the cytosol is an essential step in chloroplast biogenesis that is mediated by protein translocon complexes at the inner and outer envelope membrane (TOC). Toc159 is thought to be the main receptor for photosynthetic proteins, but lacking a large-scale systems approach, this hypothesis has only been tested for a handful of photosynthetic and nonphotosynthetic proteins. To assess Toc159 precursor specificity, we quantitatively analyzed the accumulation of plastid proteins in two mutant lines deficient in this receptor. Parallel genomewide transcript profiling allowed us to discern the consequences of impaired protein import from systemic transcriptional responses that contribute to the loss of photosynthetic capacity. On this basis, we defined putative Toc159-independent and Toc159-dependent precursor proteins. Many photosynthetic proteins accumulate in Toc159-deficient plastids, and, surprisingly, several distinct metabolic pathways are negatively affected by Toc159 depletion. Lack of Toc159 furthermore affects several proteins that accumulate as unprocessed N-acetylated precursor proteins outside of plastids. Together, our data show an unexpected client protein promiscuity of Toc159 that requires a far more differentiated view of Toc159 receptor function and regulation of plastid protein import, in which cytosolic Met removal followed by N-terminal acetylation of precursors emerges as an additional regulatory step.

Protein Abundance Changes and Ubiquitylation Targets Identified after Inhibition of the Proteasome with Syringolin A

“…If the ubiquitylated plastid-and mitochondria-targeted proteins are the cytosolic precursors, we would expect to find peptides from their transit sequences. We therefore used the length of the transit sequences predicted by TransitP and ChloroP (63) and determined the most N-terminal localized peptide for these proteins (37). We detected a peptide located in the transit peptide sequence for 10.3% of the chloroplastand 31.6% of the mitochondria-targeted proteins.…”

Posttranslational Modifications of FERREDOXIN-NADP+ OXIDOREDUCTASE in Arabidopsis Chloroplasts

“…N a -acetylation refers to the addition of an acetyl group to the a-amino group of the N-terminal amino acid of proteins. Cotranslationally occurring N a -acetylation of preprotein transit peptides has been shown to be crucial for efficient translocation into the plastids (Pesaresi et al, 2003;Bischof et al, 2011). In addition to the preproteins, many mature nucleus-encoded chloroplast proteins are targets of posttranslational N a -acetylation inside the chloroplast (Zybailov et al, 2008;Bienvenut et al, 2011Bienvenut et al, , 2012, and a number of chloroplast-encoded proteins, such as D1, D2, CP43, RUBISCO LARGE SUBUNIT, and ATP SYNTHASE « SUBUNIT (AtpE), have also been shown to be N a -acetylated (Michel et al, 1988;Mulligan et al, 1988;Zybailov et al, 2008;Hoshiyasu et al, 2013).…”

Posttranslational Modifications of Chloroplast Proteins: An Emerging Field

“…N a -acetylation of preproteins is required for organelle import, as plants lacking the homolog of NATC show disturbed structure and function of photosynthetic machinery (Pesaresi et al, 2003). Moreover, in plants lacking TOC159 (subunit of TRANSLOCON OF OUTER ENVELOPE MEMBRANE OF CHLOROPLAST), the N a -acetylated chloroplast preproteins accumulate in the cytosol (Bischof et al, 2011). The second type of N a -acetylation occurs when nuclear-encoded proteins are posttranslationally acetylated inside the chloroplast after transit peptide excision.…”