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Journal of Bacteriology, September 2007, p. 6512-6520, Vol. 189, No. 18
0021-9193/07/$08.00+0     doi:10.1128/JB.00273-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Unstable Escherichia coli L Forms Revisited: Growth Requires Peptidoglycan Synthesis{triangledown}

Danièle Joseleau-Petit,1 Jean-Claude Liébart,1 Juan A. Ayala,2 and Richard D'Ari1*

Institut Jacques Monod (C.N.R.S., Université Paris 6, Université Paris 7), 75251 Paris Cedex 05, France,1 Centro de Biología Molecular Severo Ochoa CSIC-UAM, Campus de Cantoblanco, 28049 Madrid, Spain2

Received 22 February 2007/ Accepted 8 June 2007

Growing bacterial L forms are reputed to lack peptidoglycan, although cell division is normally inseparable from septal peptidoglycan synthesis. To explore which cell division functions L forms use, we established a protocol for quantitatively converting a culture of a wild-type Escherichia coli K-12 strain overnight to a growing L-form-like state by use of the ß-lactam cefsulodin, a specific inhibitor of penicillin-binding proteins (PBPs) 1A and 1B. In rich hypertonic medium containing cefsulodin, all cells are spherical and osmosensitive, like classical L forms. Surprisingly, however, mutant studies showed that colony formation requires D-glutamate, diaminopimelate, and MurA activity, all of which are specific to peptidoglycan synthesis. High-performance liquid chromatography analysis confirmed that these L-form-like cells contain peptidoglycan, with 7% of the normal amount. Moreover, the ß-lactam piperacillin, a specific inhibitor of the cell division protein PBP 3, rapidly blocks the cell division of these L-form-like cells. Similarly, penicillin-induced L-form-like cells, which grow only within the agar layers of rich hypertonic plates, also require D-glutamate, diaminopimelate, and MurA activity. These results strongly suggest that cefsulodin- and penicillin-induced L-form-like cells of E. coli—and possibly all L forms—have residual peptidoglycan synthesis which is essential for their growth, probably being required for cell division.

* Corresponding author. Mailing address: Institut Jacques Monod, 2 place Jussieu, 75251 Paris Cedex 05, France. Phone: 331 4427 6943. Fax: 331 4427 5716. E-mail: dari{at}ijm.jussieu.fr

{triangledown} Published ahead of print on 22 June 2007.

Journal of Bacteriology, September 2007, p. 6512-6520, Vol. 189, No. 18
0021-9193/07/$08.00+0     doi:10.1128/JB.00273-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



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