Protein depletion using the arabinose promoter in Xanthomonas citri subsp. citri.
Xanthomonas citri subsp. citri (X. citri) is a plant pathogen and the etiological agent of citrus canker, a severe disease that affects all the commercially important citrus varieties, and has worldwide distribution. Citrus canker cannot be healed, and the best method known to control the spread of X. citri in the orchards is the eradication of symptomatic and asymptomatic plants in the field. However, in the state of São Paulo, Brazil, the main orange producing area in the world, control is evolving to an integrated management system (IMS) in which growers have to use less susceptible plants, windshields to prevent bacterial spread out and sprays of cupric bactericidal formulations. Our group has recently proposed alternative methods to control citrus canker, which are based on the use of chemical compounds able to disrupt vital cellular processes of X. citri. An important step in this approach is the genetic and biochemical characterization of genes/proteins that are the possible targets to be perturbed, a task not always simple when the gene/protein under investigation is essential for the organism. Here, we describe vectors carrying the arabinose promoter that enable controllable protein expression in X. citri. These vectors were used as complementation tools for the clean deletion of parB in X. citri, a widespread and conserved gene involved in the essential process of bacterial chromosome segregation. Overexpression or depletion of ParB led to increased cell size, which is probably a resultant of delayed chromosome segregation with subsequent retard of cell division. However, ParB is not essential in X. citri, and in its absence the bacterium was fully competent to colonize the host citrus and cause disease. The arabinose expression vectors described here are valuable tools for protein expression, and especially, to assist in the deletion of essential genes in X. citri.
Depto. Bioquímica e Microbiologia, Instituto de Biociências, Universidade Estadual Paulista, Av. 24A, 1515, Rio Claro, SP 13506-900, Brazil.
Lilian A
LA
Lacerda
Depto. Bioquímica e Microbiologia, Instituto de Biociências, Universidade Estadual Paulista, Av. 24A, 1515, Rio Claro, SP 13506-900, Brazil.
Lucia B
LB
Cavalca
Centro de Citricultura Sylvio Moreira, Rodovia Anhangüera, km 158, Caixa Postal 04, Cordeirópolis, SP 13490-970, Brazil.
Paula M M
PMM
Martins
Depto. de Estatística, Matemática Aplicada e Computação, Instituto de Geociências e Ciências Exatas, Universidade Estadual Paulista, Av. 24A, 1515, Rio Claro, SP 13506-900, Brazil.
José S
JS
Govone
Centro de Estudos de Insetos Sociais, Instituto de Biociências, Universidade Estadual Paulista, Av. 24A, 1515, Rio Claro, SP 13506-900, Brazil.
Maurício
M
Bacci
Depto. Bioquímica e Microbiologia, Instituto de Biociências, Universidade Estadual Paulista, Av. 24A, 1515, Rio Claro, SP 13506-900, Brazil. Electronic address: henrique.ferreira@linacre.oxon.org.
Henrique
H
Ferreira
D001089:Arabinose / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002455:Cell Division; D020090:Chromosome Segregation; D002876:Chromosomes, Bacterial / Q000378:metabolism / Q000648:ultrastructure; D002957:Citrus / Q000382:microbiology*; D003001:Cloning, Molecular; D019915:DNA Primase / Q000172:deficiency* / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D055786:Gene Knockout Techniques; D010935:Plant Diseases / Q000382:microbiology*; D018515:Plant Leaves / Q000382:microbiology; D010957:Plasmids / Q000737:chemistry / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014774:Virulence; D014974:Xanthomonas / Q000235:genetics / Q000254:growth & development / Q000472:pathogenicity*
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D001089:Arabinose; D019915:DNA Primase
Bacterial gene knockout; Chromosome segregation; Citrus canker