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Journal of Bacteriology, September 2006, p. 6059, Vol. 188, No. 17
0021-9193/06/$08.00+0     doi:10.1128/JB.00353-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

DIALOG

Rebuttal: Study of the Deinococcus radiodurans Nucleoid

Department of Organic Chemistry,¹ Chemical Services, The Weizmann Institute of Science, Rehovot 76100, Israel²

In their dialog article (2), Eltsov and Dubochet highlight the unique benefits associated with the cryoelectron microscopy of vitreous sections (CEMOVIS) technique, which allows for the visualization of biological specimens in their native, unstained states, thus providing a means for elucidating the fine structures of intracellular components.

We agree with the authors that CEMOVIS has several appealing advantages but argue that, like any other imaging method, it has its own intrinsic shortcomings. Perhaps the most serious drawback of this technique is the large compression of the specimen in the direction of the cutting, which falls in the range of 30 to 50% and is particularly severe for thin sections (4). The dramatic outcomes of this compression are indeed evident when the distances between the S layer and the cytoplasmic membrane along and perpendicular to the direction of the cutting are compared (see Fig. 1 in reference 2). When compressed by 30 to 50%, bona fide ring-like structures will inevitably reveal rod-like or completely unstructured morphologies. We consequently claim that this compression, along with the fact that DNA cannot be specifically stained in CEMOVIS preparations, renders this technique inadequate for assessing the global structure of bacterial genomes.

Eltsov and Dubochet speculate that the ring-like structures observed in our electron microscopy studies (3) as well as in fluorescence studies of various members of the Deinococcaceae family (6) might result from DNA wrapping around intracellular granules. Notably, even if that were the case, DNA spooling around a granule would result in a tight DNA alignment and hence restricted diffusion. Granules are, however, seldom observed in exponentially growing Deinococcus radiodurans cells and practically never observed in stationary-state cells.

Eltsov and Dubochet claim that DNA repair in D. radiodurans is based upon physiological, rather than structural, factors. At the heart of our model lies the notion that following genome fragmentation into more than 150 fragments (formally resulting in 150! different pathways for religation) and in the absence of an intact DNA template, a structural support for accurate DNA repair is a sine qua non. Restricted diffusion of DNA fragments effected in tightly packed DNA structures would allow for efficient and, to a high extent, error-free repair processes through nonhomologous end joining. The identification of a ligase that specifically promotes DNA end joining in D. radiodurans (5) strongly supports this notion. We agree with Cox and Battista (1), who recently suggested that restricted diffusion does not necessarily depend upon a particular DNA morphology but can be effected by any mode of DNA condensation. We note, however, that ring-like DNA conformations represent a particularly stable mode of DNA packaging that is encountered not only in members of the Deinococcaceae family but also in other resistant bacterial species such as Bacillus subtilis spores (3).

	FOOTNOTES

 
* Corresponding author. Mailing address: Department of Organic Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel. Phone: 972-8-9342003. Fax: 972-8-9344142. E-mail: avi.minsky{at}weizmann.ac.il.

REFERENCES

1. Cox, M. M., and J. R. Battista. 2005. Deinococcus radiodurans—the consummate survivor. Nat. Rev. Microbiol. 3:882-892.[CrossRef][Medline]
2. Eltsov, M., and J. Dubochet. 2006. A study of the Deinococcus radiodurans nucleoid by cryoelectron microscopy of vitreous sections: supplementary comments. J. Bacteriol. 188:6053-6058.[Free Full Text]
3. Englander, J., E. Klein, V. Brumfeld, A. K. Shama, A. J. Doherty, and A. Minsky. 2004. DNA toroids: framework DNA repair in Deinococcus radiodurans and in germinating bacterial spores. J. Bacteriol. 186:5973-5977.[Free Full Text]
4. Hsieh, C. E., A. Leith, C. A. Mannella, J. Frank, and M. Marko. 2006. Towards high-resolution three-dimensional imaging of native mammalian tissue: electron tomography of frozen-hydrated rat liver sections. J. Struct. Biol. 153:1-13.[CrossRef][Medline]
5. Narumi, I., K. Satoh, S. Cui, T. Funayama, S. Kitayama, and H. Watanabe. 2004. PprA: a novel protein from Deinococcus radiodurans that stimulates DNA ligation. Mol. Microbiol. 54:278-285.[CrossRef][Medline]
6. Zimmerman, J. M., and J. R. Battista. 2005. A ring-like nucleoid is not necessary for radioresistance in the Deinococcaceae. BMC Microbiol. 5:17-27.[CrossRef][Medline]

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