The rcsA Gene from Erwinia amylovora: Identification, Nucleotide Sequence, and Regulation of Exopolysaccharide Biosynthesis. Frank Bernhard. Department of Plant Pathology, The Ohio State University, Columbus 43210-1087 U.S.A. Karl Poetter(1), Klaus Geider(2), and David L. Coplin(1). (1)Department of Plant Pathology, The Ohio State University, Columbus 43210-1087 U.S.A., and (2)Max-Planck-Institut für Medizinische Forschung, Abteilung Biophysik, D-6900 Heidelberg, Federal Republic of Germany.. MPMI 3:429-437. Accepted 27 July 1990. Copyright 1990 The American Phytopathological Society.

RcsA is a positive activator of extracellular polysaccharide synthesis in the Enterobacteriaceae. A cosmid clone containing the rcsA gene from Erwinia amylovora was identified by its ability to restore mucoidy to an E. stewartii rcsA mutant. The rcsA gene was subcloned on a 2.2-kilobase HindIII-PstI fragment that hybridized with an E. stewartii rcsA probe and complemented E. stewartii and Escherichia coli rcsA mutants. In addition, the cloned E. amylovora rcsA gene stimulated expression of cps::lac fusions in E. coli and E. stewartii. The rcsA region was sequenced, and one open reading frame of 211 amino acids was found. The predicted protein sequence specified by this open reading frame was 55% homologous with that of the Klebsiella pneumoniae RcsA protein. Highly conserved regions in the 3’ and 5’ ends of the two proteins were observed. An E. amylovora rcsA mutant was constructed by Tn5 mutagenesis of the cloned gene followed by recombination of the mutation into the chromosome of wildtype strain Ea1/79. The synthesis of both amylovorin and levan was reduced by more than 90% in this mutant, indicating common regulation of the two polysaccharides by rcsA. Virulence of the rcsA mutant on immature pear fruit was diminished but not completely abolished.