Bilateral NSF/BIO-BBSRC Synthesis of Microcompartments in Plants for Enhanced Carbon Fixation

Abstracts are not currently available in GtR for all funded research. This is normally because the abstract was not required at the time of proposal submission, but may be because it included sensitive information such as personal details.

Experimental data and modeling indicate that increasing photosynthesis increases crop yields, provided that other constraints do not become limiting. One approach to enhancing the efficiency of photosynthesis is to introduce carbon-concentrating mechanism (CCM), which increases the CO2 substrate available for assimilation by Rubisco (ribulose-1,5-bisphosphate (RuBP) carboxylase/oxygenase) and suppresses a competing reaction with O2. The oxygenation of RuBP is also catalysed by Rubisco and initiates photorespiration, with the consequent loss of fixed carbon, consumption of energy and release of nitrogen. CCMs limit photorespiration by altering the proportion of the two gaseous substrates, CO2 and O2, in the vicinity of Rubisco. While CCMs have evolved in a variety of organisms, including certain crop species such as maize, they are lacking in many globally important, so-called C3 crop species such as soybean and wheat. We propose to apply the methods of synthetic biology to engineer a CCM derived from cyanobacteria into the chloroplast in a model C3 plant, tobacco, with potential for future application to crop plants. The cyanobacterial Beta-carboxysome will be constructed in the tobacco chloroplast through expression of component proteins from the chloroplast genome. We will optimize chloroplast vectors for modular cloning that will allow rapid design of new operons to introduce carboxysomal proteins in the proper ratio for assembly of the microcompartment. Our efforts to adjust protein levels will be guided by assays of transgene expression, electron and fluorescence microscopy, and immunolocalization. Incorporating the complete system into chloroplasts will also require introduction of a nuclear-encoded bicarbonate transporter and genetic removal of carbonic anhydrase from the chloroplast stroma. Promising transgenic lines will be assayed for carbon assimilation, stomatal conductance, Rubisco amount and activity, and growth rate and biomass accumulation.

This proposal is for fundamental research to develop new conceptual approaches relevant to ideas emerging within the international plant, systems and synthetic biology communities. The research to develop a new micro-compartment will stimulate thinking around strategies for modelling and for applications of synthetic biology in plants, especially in relation to photosynthesis, and it should strengthen methodologies relevant at many levels from cell to crop engineering. Thus, the research is expected to benefit fundamental researchers and, in the longer-term, agriculture and industry, through conceptual developments and approaches to not only improve carbon capture by plants but also for other applications (e.g. packaging bioactive molecules). The research will feed into higher education training programmes through capacity building at the postgraduate and postdoctoral levels. Additional impact is proposed through public displays and the development of teaching resources building on the background work for this proposal. Finally the research will help guide future efforts in applications to agricultural/industrial systems. The applicants have established links with industrial/technology transfer partners and research institutes to take advantage of these developments. Further details of these, and additional impacts will be found in Part 1 of the Case for Support and in the attached Impact Pathways.