Hanson Lab
 
Improving Photosynthesis by Engineering a Cyanbacterial Carboxysome into Chloroplasts
 

Punctate loci obtained by expression and chloroplast targeting of multiple carboxysomal proteins (Red=chlorophyll autofluorescence; Green=YFP label on a carboxysomal protein)

 

 

Image: Punctate loci obtained by expression and chloroplast targeting of multiple carboxysomal proteins (Red=chlorophyll autofluorescence; Green=YFP label on a carboxysomal protein)


 

Yields of many crops could be increased if photosynthesis could be engineered to become more efficient.   The plant carbon-fixing enzyme Rubisco (ribulose bisphosphate carboxylase/oxygenase) has poor kinetic properties with regard to its reaction with CO2, and also reacts with O2, leading to the wasteful process of photorespiration.


One possible way to enhance carbon fixation by Rubisco is to surround the enzyme with CO2,  a strategy that has evolved in cyanobacteria (1).  These microorganisms contain microcompartments named carboxysomes that encapsulate Rubisco in a protein shell and contain an enzyme,  carbonic anhydrase (CA), which converts bicarbonate ion to CO2.   A typical microcompartment is 100-200 nm in size, with a 3-4 nm proteinaceous shell.


Cyanobacteria such as Synchecoccus elongatus encode a Rubisco enzyme that is faster than land plant Rubisco, though more oxygen sensitive than plant enzymes.  Because CO2  is concentrated near Rubisco within the carboxysome, the enzyme can work efficiently to fix carbon.  We have been able to express β-cyanobacterial carboxysome shell proteins transiently in chloroplasts of tobacco leaves.  By labeling one of the shell proteins with Yellow Fluorescent Protein (YFP), we were able to use confocal fluorescence microscopy to observe punctuate loci forming inside chloroplasts.    Electron microscopy by our collaborators in Martin Parry’s group at Rothamsted Research revealed the presence of both linear structures and oval structures similar to empty microcomparments. (2)


We have replaced the tobacco RbcL gene with transgenic loci containing two or three cyanobacterial genes encoding the S. elongatus Rubisco large subunit (Se LS), small subunit (Se SS) either alone or with the putative chaperone RbcX or a gene encoding the carboxysomal protein M35. Tobacco plants could grow phototrophically despite the absence of tobacco Rubisco, if provided with elevated CO2. (3,4)

This unexpected finding was reported in a number of lay publications (see News and Events).

The gene regulatory sequences in the SeLS construct were improved from our initial SeLSX and SeLSM35 constructs, allowing greater accumulation of cyanobacterial Rubisco and improved growth. In elevated CO2, the SeLS plants could grow nearly as fast as wild-type (4).


 

 
Citations
 

1. Hanson MR, Lin MT, Carmo-Silva AE, Parry MA. 2016 Towards engineering carboxysomes into C3 plants. Plant J. 2016.

2. Lin, M. T., Occhialini, A., Andralojc, P. J., Devonshire, J., Hines, K. M., Parry, M. A., and Hanson, M. R. (2014) ß-carboxysomal proteins assemble into highly organized structures in Nicotiana chloroplasts. Plant Journal 79, 1-12.

3. Lin , M.T., A. Occhialini, P.J. Andralojc, M.A. Parry and M.R. Hanson. 2014. A faster Rubisco with potential to increase photosynthesis in crops. Nature 513:547-50.

4. Occhialini A, Lin MT, Andralojc PJ, Hanson MR, Parry MA. 2016 Transgenic tobacco plants with improved cyanobacterial Rubisco expression but no extra assembly factors grow at near wild-type rates if provided with elevated CO2. Plant Journal 85:148-60

 
Collaborators
 

Laboratory of Martin Parry at Rothamsted Research

Alessandro Occhialini, John Andralojc, Jean Devonshire

 


Laboratory of Stephen Long at University of Illinois

Justin McGrath

 

 

Laboratory of Cheryl Kerfeld at Michigan State University

Jeff Cameron, Markus Sutter

 
Project Sponsors
 

 
Maureen R. Hanson
Liberty Hyde Bailey Professor
Phone: 607-254-4833
Fax: 607-255-6249

Hanson Laboratory
Department of Molecular Biology and Genetics
321 Biotechnology Building
Cornell University
Ithaca, NY 14853
Phone: 607-254-4832

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Maureen Hanson at NCBI Pubmed   Maureen Hanson at Google Scholar