Organization, Structure, and Assembly of α-Carboxysomes Determined by Electron Cryotomography of Intact Cells

Iancu, Cristina V. and Morris, Dylan M. and Dou, Zhicheng and Heinhorst, Sabine and Cannon, Gordon C. and Jensen, Grant J. (2010) Organization, Structure, and Assembly of α-Carboxysomes Determined by Electron Cryotomography of Intact Cells. Journal of Molecular Biology, 396 (1). pp. 105-117. ISSN 0022-2836 http://resolver.caltech.edu/CaltechAUTHORS:20100308-111110207

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	PDF (Supplementary Figure 1. Carboxysomes in dividing cells: a) table listing the numbers of carboxysomes in each set of daughter cells; b) histogram showing the distribution of the difference in numbers of carboxysomes in nascent daughter cells.) - Supplemental Material Restricted to Repository administrators only See Usage Policy. 51Kb
	PDF (Supplementary Figure 2. Carboxysomes showing gaps in the internal RuBisCO lattice. Panel 1 is from an H. neapolitanus cell. Panels 2-6 are from cells of T. intermedia. Scale bar is 50 nm. ) - Supplemental Material Restricted to Repository administrators only See Usage Policy. 167Kb
	PDF (Supplementary Figure 3. Additional EELS images of H. neapolitanus cells. Each pair of panels shows the zero-loss 2D projection image (left) and the 2D-projection electron energy loss spectrographic image focused on phosphorous (right) from the same cell. ) - Supplemental Material Restricted to Repository administrators only See Usage Policy. 1960Kb
	PDF (Supplementary Figure 4. Additional tomographic slices of T. intermedia cells: a) T. intermedia cell showing abundance of carboxysomes, clusters of carboxysomes, and physical associations between carboxysomes and a polyP granule (note the lattice emanating) - Supplemental Material Restricted to Repository administrators only See Usage Policy. 551Kb
	PDF (Supplementary Figure 5. Additional tomographic slices of T. crunogena cells: a) T. crunogena cell containing over 80 carboxysomes (not all are visible in the tomographic slice shown) (note the physical associations between carboxysomes and polyP granules,) - Supplemental Material Restricted to Repository administrators only See Usage Policy. 765Kb
	PDF (Supplementary Figure 6. Additional internal granules. Scale bar is 50 nm. ) - Supplemental Material Restricted to Repository administrators only See Usage Policy. 424Kb
	PDF (Supplementary Figure 7. Additional partial carboxysomes. Scale bar is 50 nm. ) - Supplemental Material Restricted to Repository administrators only See Usage Policy. 828Kb
	Video (QuickTime) (Supplementary Movie 1. Aligned tilt-series of the H. neapolitanus cell depicted in Fig. 1A. ) - Supplemental Material Restricted to Repository administrators only See Usage Policy. 15Mb
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	Video (QuickTime) (Supplementary Movie 4. Tomographic reconstruction of the T. crunogena cell depicted in Fig. 1C) - Supplemental Material Restricted to Repository administrators only See Usage Policy. 19Mb

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Abstract

Carboxysomes are polyhedral inclusion bodies that play a key role in autotrophic metabolism in many bacteria. Using electron cryotomography, we examined carboxysomes in their native states within intact cells of three chemolithoautotrophic bacteria. We found that carboxysomes generally cluster into distinct groups within the cytoplasm, often in the immediate vicinity of polyphosphate granules, and a regular lattice of density frequently connects granules to nearby carboxysomes. Small granular bodies were also seen within carboxysomes. These observations suggest a functional relationship between carboxysomes and polyphosphate granules. Carboxysomes exhibited greater size, shape, and compositional variability in cells than in purified preparations. Finally, we observed carboxysomes in various stages of assembly, as well as filamentous structures that we attribute to misassembled shell protein. Surprisingly, no more than one partial carboxysome was ever observed per cell. Based on these observations, we propose a model for carboxysome assembly in which the shell and the internal RuBisCO (ribulose-1,5-bisphosphate carboxylase/oxygenase) lattice form simultaneously, likely guided by specific interactions between shell proteins and RuBisCOs.

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Article

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© 2009 Elsevier Ltd. Received 18 June 2009; received in revised form 6 November 2009; accepted 9 November 2009; available online 17 November 2009. Edited by W. Baumeister. This work was supported in part by the National Institutes of Health through grants R01 AI067548 and P50 GM082545 to G.J.J., the Department of Energy through grant DE-FG02-04ER63785 to G.J.J., the National Science Foundation through grant MCB-0818680 to G.C.C. and S.H., the Beckman Institute at Caltech, and gifts to Caltech from the Gordon and Betty Moore Foundation and Agouron Institute. We are indebted to Dr. Raj Menon for culturing T. crunogena cells, Dr. Eric Williams for providing purified H. neapolitanus carboxysomes and initial batches of H. neapolitanus cells, Dr. H. Jane Ding for providing computational help with generation of phosphorus elemental maps, Dr. William F. Tivol for providing technical assistance in setting up the spectroscopic imaging experiments, and Dr. Elizabeth R. Wright for freezing grids and imaging several T. intermedia cells.

Funders:

Funding Agency	Grant Number
NIH	R01 AI067548
NIH	P50 GM082545
Department of Energy	DE-FG02-04ER63785
NSF	MCB-0818680
Beckman Institute at Caltech	UNSPECIFIED
Gordon and Betty Moore Foundation	UNSPECIFIED
Agouron Institute	UNSPECIFIED

Subject Keywords:

H. neapolitanus; bacterial microcompartment; carboxysome; cryo-EM; electron tomography

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CaltechAUTHORS:20100308-111110207

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http://resolver.caltech.edu/CaltechAUTHORS:20100308-111110207

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ID Code:

17694

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CaltechAUTHORS

Deposited By:

Jason Perez

Deposited On:

08 Mar 2010 22:38

Last Modified:

26 Dec 2012 11:49

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