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bioRxiv · 10.64898/2026.08.28.747797

In-cell structural analysis reveals a distinctive chloroplast ribosome in Chlamydomonas reinhardtii

Zhang, P.·Hou, Z.·Shen, Y.·Zhang, Z.·Lu, P.·Katzourakis, A.
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Abstract

Chloroplast ribosomes synthesize plastid-encoded components of photosynthetic machinery, yet their structure and organization remain poorly understood. We combined cryo-focused ion beam milling, cryo-electron tomography and subtomogram averaging to determine native chloroplast ribosomes in Chlamydomonas reinhardtii. The 4.4-4.9 [A] structure revealed a large arch-like extension on the small subunit (SSU). Comparisons with bacterial and plant chloroplast ribosomes, supported by proteomics, AlphaFold3 predictions and a recent atomic model, indicate that the arch is formed by insertions and extensions in SSU proteins. Classification resolved active, thylakoid-associated ribosomes with density adjacent to the nascent peptide exit and an arch-moved state enriched among thylakoid-associated particles, with coordinated displacement of the arch and beak. Phylogenetic analysis revealed an evolutionary mosaic: the uS3c insertion is broadly distributed across Chlorophyceae, whereas the uS2c insertion, uS5c and PSRP7 are concentrated in Chlamydomonadales, with PSRP7 also in Sphaeropleales. Nuclear-encoded components were recruited stepwise onto a plastid-encoded scaffold, with all four under comparable purifying selection. These findings link a lineage-specific SSU extension to ribosome dynamics, thylakoid association and evolution, highlighting the value of in-cell structural analysis.

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Zhang, P., Hou, Z., Shen, Y., Zhang, Z., Lu, P., Katzourakis, A.. 2026-09-03. In-cell structural analysis reveals a distinctive chloroplast ribosome in Chlamydomonas reinhardtii. https://doi.org/10.64898/2026.08.28.747797

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