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Westrich, L. D.

Publications and source records attributed to Westrich, L. D..

2 recordsLinked to original sources

OHP2 is not required for psbA translation in Chlamydomonas

In land plants and cyanobacteria, co-translational association of chlorophyll (Chl) to the nascent D1 polypeptide, a reaction center protein of photosystem II (PSII), requires a Chl binding complex consisting of a short-chain dehydrogenase (HCF244/Ycf39) and One-Helix Proteins of the LHC superfamily (OHP1 and OHP2 in chloroplasts). Here, we show that an ohp2 mutant of the green alga Chlamydomonas reinhardtii fails to accumulate core PSII subunits, in particular D1. Extragenic suppressors arise at high frequency, suggesting the existence of another route for Chl association to PSII. The ohp2 mutant can be complemented by the Arabidopsis ortholog. In contrast to land plants, where psbA translation is prevented in the absence of OHP2, ribosome profiling experiments show that the Chlamydomonas mutant translates the psbA transcript over its full length. Pulse labelling suggests that D1 is degraded during or immediately after translation. The translation of other PSII subunits is affected by assembly-controlled translational regulation (the CES process). Proteomics show that HCF244, a translation factor which associates with and is stabilized by OHP2 in land plants, still partly accumulates in the Chlamydomonas ohp2 mutant, explaining the persistence of psbA translation. Several Chl biosynthesis enzymes overaccumulate in the mutant membranes. Partial inactivation of the D1-degrading FtsH protease restores a low level of PSII activity in an ohp2 background, but not photoautotrophy. Taken together, our data suggest that OHP2 is not required for psbAD1 translation in Chlamydomonas, but necessary for its stabilization.

plant biology↗

The proteomic inventory reveals the chloroplast ribosome as nexus within a diverse protein network

Chloroplast gene expression is tightly regulated and majorly controlled on the level of protein synthesis. Fine-tuning of translation is vital for plant development, acclimation to environmental challenges and for the assembly of major protein complexes such as the photosynthesis machinery. However, many regulatory mediators and the interaction network of chloroplast ribosomes are not known to date. We report here on a deep proteomic analysis of the plastidic ribosome interaction network in Chlamydomonas reinhardtii cells. Affinity-purification of ribosomes was achieved via endogenous affinity tagging of the chloroplast-encoded protein Rpl5, yielding a specific enrichment of >650 chloroplast-localized proteins. The ribosome interaction network was validated for several proteins and provides a new source of mainly conserved factors directly linking translation with central processes such as protein folding, photosystem biogenesis, redox control, RNA maturation, energy and metabolite homeostasis. Our approach provided the first evidence for the existence of a plastidic co-translational acting N-acetyltransferase (cpNAT1). Expression of tagged cpNAT1 confirmed its ribosome-association, and we demonstrated the ability of cpNAT1 to acetylate substrate proteins at their N-terminus. Our dataset establishes that the chloroplast protein synthesis machinery acts as nexus in a highly choreographed, spatially interconnected protein network and underscores its wide-ranging regulatory potential during gene expression. ONE-SENTENCE SUMMARYAffinity purification of Chlamydomonas reinhardtii chloroplast ribosomes and subsequent proteomic analysis revealed a broad spectrum of interactors ranging from global translation control to specific pathways.

biochemistry↗