1.7.3S
ECOLOGICAL AND MOLECULAR ASPECTS OF INDOLE-3-ACETIC ACID PRODUCTION IN AN EPIPHYTIC STRAIN OF ERWINIA HERBICOLA

MT BRANDL and SE LINDOW

Plant and Microbial Biology Deptartment, 111 Koshland Hall, University of California, Berkeley, CA 94720, USA

Erwinia herbicola is a successful colonizer of aerial plant surfaces which commonly produces the plant growth regulator indole-3-acetic acid (IAA) in culture supplemented with tryptophan. IAA-producing strains of E. herbicola have been implicated in pear fruit russetting, presumably by altering the normal development of epidermal cell layers. Unlike auxin production in gall-forming pathogenic bacteria, the role of IAA biosynthesis in epiphytic bacteria remains unclear. E. herbicola 299R, a strain isolated from pear in California, synthesizes IAA via the indolepyruvate pathway, contrary to gall-forming bacteria which produce IAA through indoleacetamide. ipdC, a gene involved in the indolepyruvate pathway, was cloned from strain 299R. Insertional mutagenesis of ipdC significantly reduced the ability of this strain to produce IAA.

The study of the transcriptional activity of ipdC-inaZ fusions showed that ipdC is plant-inducible and that its expression increased as much as 73-fold under conditions of low water availability in culture. Induction of ipdC on plants and in vitro was reduced 10-fold in a sigma38 mutant of strain 299R, suggesting that IAA production in E. herbicola may be regulated by signals related to environmental stresses in the phyllosphere. Insertional interruption of rpoS also reduced the ability of strain 299R to colonize plant surfaces and to survive many stresses in vitro. Epiphytic fitness studies conducted in the greenhouse and in the field revealed that strain 299R achieved higher population sizes than its isogenic IAA- strain. The increasing ratio of the populations of strain 299R to its IAA- mutant during periods of active growth on plant tissue indicate that IAA production conferred a selective advantage to the parental strain, possibly by modification of its microhabitat. We hypothesize that epiphytic bacteria benefit from the production and secretion of IAA by increasing the permeability of plant cells and thus triggering the release of nutrients in their vicinity. Transcriptional fusions of a sucrose-responsive promoter to the green fluorescent protein (GFP) reporter gene are being used to study the differential sensing of nutrients in strain 299R and its IAA- mutant in situ on plant surfaces. This 'biological sensor' should reveal if higher amounts of sucrose are available to IAA-secreting bacterial cells during colonization of plant tissue. The distribution of the expression of ipdC-GFP fusions is also being investigated to determine the relative production of IAA in strain 299R in various microsites on the phylloplane. These studies should provide insight into the ecological significance of IAA biosynthesis in epiphytic bacteria.