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Journal of Bacteriology, November 2009, p. 6694-6700, Vol. 191, No. 21
0021-9193/09/$08.00+0     doi:10.1128/JB.00839-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Dam Methylation Controls O-Antigen Chain Length in Salmonella enterica Serovar Enteritidis by Regulating the Expression of Wzz Protein{triangledown}

Sebastián H. Sarnacki,1,2 Cristina L. Marolda,3 Mariángeles Noto Llana,1,2 Mónica N. Giacomodonato,1,2 Miguel A. Valvano,3,4* and María Cristina Cerquetti1,2*

CEFYBO-UBA-CONICET,1 Departamento de Microbiología, Parasitología e Inmunología, Facultad de Medicina, Universidad de Buenos Aires, Buenos Aires, Argentina,2 Infectious Diseases Research Group, Siebens-Drake Research Institute,3 Departments of Microbiology and Immunology and Medicine, University of Western Ontario, London, Ontario N6A 5C1, Canada4

Received 25 June 2009/ Accepted 18 August 2009

We reported previously that a Salmonella enterica serovar Enteritidis dam mutant expressing a truncated Dam protein does not agglutinate in the presence of specific antibodies against O9 polysaccharide. Here we investigate the participation of Dam in lipopolysaccharide (LPS) synthesis in Salmonella. The LPS O-antigen profiles of a dam null mutant (SE{Delta}dam) and the Salmonella serovar Enteritidis parental strain were examined by using electrophoresis and silver staining. Compared to the parental strain, SE{Delta}dam produced LPS with shorter O-antigen polysaccharide chains. Since Wzz is responsible for the chain length distribution of the O antigen, we investigated whether Dam methylation is involved in regulating wzz expression. Densitometry analysis showed that the amount of Wzz produced by SE{Delta}dam is threefold lower than the amount of Wzz produced by the parental strain. Concomitantly, the activity of the wzz promoter in SE{Delta}dam was reduced nearly 50% in logarithmic phase and 25% in stationary phase. These results were further confirmed by reverse transcription-PCR showing that wzz gene expression was threefold lower in the dam mutant than in the parental strain. Our results demonstrate that wzz gene expression is downregulated in a dam mutant, indicating that Dam methylation activates expression of this gene. This work indicates that wzz is a new target regulated by Dam methylation and demonstrates that DNA methylation not only affects the production of bacterial surface proteins but also the production of surface polysaccharides.

* Corresponding author. Mailing address for María Cristina Cerquetti: Departamento de Microbiología, Parasitología e Inmunología, Facultad de Medicina, Universidad de Buenos Aires, Paraguay 2155, 1121 Buenos Aires, Argentina. Phone: 054-11-5950-9500. Fax: 054-11-4964-2554. E-mail: ccerquetti{at}fmed.uba.ar. Mailing address for Miguel A. Valvano: Departament of Microbiology and Immunology, University of Western Ontario, Dental Science Building 3014, London, Ontario, Canada N6A 5C1. Phone: (519) 661-3427. Fax: (519) 661-3499. E-mail: mvalvano{at}uwo.ca

{triangledown} Published ahead of print on 28 August 2009.

Journal of Bacteriology, November 2009, p. 6694-6700, Vol. 191, No. 21
0021-9193/09/$08.00+0     doi:10.1128/JB.00839-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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