Hybrid cyanobacterial-tobacco Rubisco supports autotrophic growth and pre-carboxysomal aggregation

Orr, Douglas and Worrall, Dawn and Lin, Myat T. and Carmo-Silva, Elizabete and Hanson, Maureen R. and Parry, Martin (2020) Hybrid cyanobacterial-tobacco Rubisco supports autotrophic growth and pre-carboxysomal aggregation. Plant Physiology, 182. pp. 807-818. ISSN 0032-0889

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Abstract

Much of the research aimed at improving photosynthesis and crop productivity attempts to overcome shortcomings of the primary CO2 fixing enzyme Rubisco. Cyanobacteria utilize a CO2 concentrating mechanism (CCM), which encapsulates Rubisco with poor specificity but a relatively fast catalytic rate within a carboxysome micro-compartment. Alongside the active transport of bicarbonate into the cell, and localization of carbonic anhydrase within the carboxysome shell with Rubisco, cyanobacteria are able to overcome the limitations of Rubisco via localization within a high CO2 environment. As part of ongoing efforts to engineer a β-cyanobacterial CCM into land plants, we investigated the potential for Rubisco large subunits (LSU) from the β-cyanobacteria Synechococcus elongatus (Se) to form aggregated Rubisco complexes with the carboxysome linker protein CcmM35 within tobacco (Nicotiana tabacum) chloroplasts. Transplastomic plants were produced that lacked cognate SeRubisco small subunits (SSU) and expressed SeLSU in place of tobacco LSU, with and without CcmM35. Plants were able to form a hybrid enzyme utilizing tobacco SSU and the SeLSU, allowing slow autotrophic growth in high CO2. CcmM35 was able to form large Rubisco aggregates with the SeLSU, and these incorporated small amounts of native tobacco SSU. Plants lacking the SeSSU showed delayed growth, poor photosynthetic capacity and significantly reduced Rubisco activity compared to both wild-type tobacco and lines expressing the SeSSU. These results demonstrate the ability of the SeLSU and CcmM35 to form large aggregates without the cognate SeSSU in planta, harboring active Rubisco that enables plant growth, albeit at a much slower pace than plants expressing the cognate SeSSU.