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DIALOG

Rebuttal: Ring-Like Nucleoids and DNA Repair in Deinococcus radiodurans

Laboratoire d'Analyse Ultrastructurale, Biophore, Université de Lausanne, CH-1015 Lausanne, Switzerland

Our study of the Deinococcus radiodurans Sark nucleoid by cryoelectron microscopy of vitreous sections (CEMOVIS) revealed that DNA is not arranged in tight toroids (1). Minsky et al. expressed doubts about our results based on the claimed irrelevance of our micrographs and the strength of previous evidence in favor of the toroidal model (3). In our dialog article (2), we explain why fully hydrated, unfixed, and unstained cryosections must be evaluated with criteria other than conventional sections and why our images are trustworthy.

In their article, Minsky et al. claim that highly dense or even toroidal packing of DNA is present in D. radiodurans. They base their conclusion on the general shape of the nucleoid (3). We agree that this shape can give an indication about DNA organization, but the final proof can come only from the molecular arrangement. With CEMOVIS, the latter can be seen directly.

CEMOVIS revealed, in locally ordered DNA, bundles of D. radiodurans Sark which are not compatible with a toroidal spooling of DNA (1, 2). It further showed that the DNA packing is not very dense, with an interfilamentous distance of ca. 4.8 nm. Minsky et al. dismiss these observations with the statement that our method is irrelevant for the study of DNA arrangement (3). This claim is not supported by the presented data (1, 2), and it ignores previously published CEMOVIS studies, which clearly show DNA arrangements in DNA crystals and liquid crystals (4, 5).

Freeze-substitution is, according to Minsky et al., the optimal high-resolution method for the observation of nucleoids. Nevertheless, their study of the D. radiodurans nucleoid, as with any other study using this method, has reported neither the arrangement of individual DNA filaments nor the interfilamentous distance. This is to be expected since, during freeze-substitution, water is removed from the sample. Molecules swimming in the native sample do not fly away when water is gone!

In order to remove any uncertainty and constructively resolve the above dispute, we propose that those (up to three groups) who have experience with D. radiodurans and evidence of toroidal nucleoid test it in our laboratory (we will cover local expenses). The visitor will prepare the bacteria under adequate conditions, and we will observe them with CEMOVIS (ca. 1 to 2 days). Together we will analyze the results. In case of disagreement, we propose consulting the editor in chief of the Journal of Bacteriology.

	FOOTNOTES

 
* Corresponding author. Mailing address: Laboratoire d'Analyse Ultrastructurale, Biophore, Université de Lausanne, CH-1015 Lausanne, Switzerland. Phone: 41 21 6924289. Fax: 41 21 6924285. E-mail: Mikhail.Eltsov{at}unil.ch.

REFERENCES

1. Eltsov, M., and J. Dubochet. 2005. Fine structure of the Deinococcus radiodurans nucleoid revealed by cryoelectron microscopy of vitreous sections. J. Bacteriol. 187:8047-8054.[Abstract/Free Full Text]
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. Minsky, A., E. Shimoni, and J. Englander. 2006. Ring-like nucleoids and DNA repair through error-free nonhomologous end joining in Deinococcus radiodurans. J. Bacteriol. 188:6047-6051.[Free Full Text]
4. Richter, K., and J. Dubochet. 1990. High resolution study of DNA in vitrified sections, p. 488-489. In L. D. Peachey and D. B. Williams (ed.), Proceedings of the 12th International Congress of Electron Microscopy, vol. 1. San Francisco Press, Inc., San Francisco, Calif.
5. Sartori Blanc, N., A. Senn, A. Leforestier, F. Livolant, and J. Dubochet. 2001. DNA in human and stallion spermatozoa forms local hexagonal packing with twist and many defects. J. Struct. Biol. 134:76-81.[CrossRef][Medline]

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