Identification of thioredoxin-dependent regulatory cysteines in Chlamydomonas reinhardtii using the reductome approach.

ABSTRACT: Thiol-based redox post-translational modifications have emerged as important mechanisms of signaling and regulation in all organisms. In this context, the small ubiquitous oxido-reductase thioredoxin (Trx) plays a key role by controlling the thiol-disulfide status of target proteins. Recent redox proteomic studies revealed hundreds of proteins regulated by glutathionylation and nitrosylation in the unicellular green alga Chlamydomonas reinhardtii while much less is known about the thioredoxin interactome, called thioredoxome hereafter, in this photosynthetic model organism. Using a quantitative large scale proteomic analysis based on the in vitro reconstitution of the enzymatic thioredoxin system (NADPH, NTR, Trx) within an acellular extract and followed by an isotopic labeling with cleavable Isotopic-Coded Affinity Tag reagents, we identified 1052 Trx-targeted cysteines spread over 602 proteins. These Trx-targets participate in a wide variety of metabolic pathways and cellular processes. This approach, combined with the affinity purification approach study (PXD006097) broadens not only the redox regulation to new enzymes involved in metabolic pathways already known to be regulated by thioredoxins (carbon, lipid and energy metabolism) but also shed light on cellular processes for which data supporting a putative redox regulation in these processes are scarce (aromatic amino-acid biosynthesis, nuclear transport,…). Moreover, by comparing this thioredoxome with proteomic data for glutathionylation and nitrosylation at the protein and cysteine levels, this work confirms the existence of a complex redox regulation network in Chlamydomonas and provides evidence of a tremendous selectivity of redox post-translational modifications for specific cysteines residues.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Chlamydomonas Reinhardtii

TISSUE(S): Photosynthetic Cell

SUBMITTER: Christophe H. MARCHAND  

LAB HEAD: Stephane Lemaire

PROVIDER: PXD006116 | Pride | 2017-07-28

REPOSITORIES: Pride