Supplemental Text 1. Role categories of genes in the GacA and RpoS regulons
An overview of the GacA and RpoS regulons of Pf-5 grown in culture and on seed
surfaces was obtained through analysis of the role categories overrepresented in each set of
regulated genes (Fig. S1).
Only the Hypothetical Proteins role category was overrepresented in the RpoS regulon of
Pf-5 grown on seed, but the Transport and Binding Proteins role category was significantly
overrepresented in the sets of genes both upregulated and downregulated in an rpoS mutant
grown in culture. Of the 27 upregulated genes in this role category, 11 belong to the Cations and
Iron Carrying Compounds subrole. These are almost exclusively related to iron transport,
including genes involved in siderophore transport and the TonB system. Six other upregulated
genes in this role category are in the pyoluteorin gene cluster. In contrast, on the downregulated
gene list, 17 of 35 genes belong to the Amino Acids, Peptides, and Amines subrole and primarily
encode amino acid transporters.
Four role categories are overrepresented in the list of genes upregulated in a gacA mutant
on seed: Biosynthesis of Cofactors, Prosthetic groups, and Carriers (16 genes), Mobile and
Extrachromosomal Element Functions (10 genes), Transport and Binding Proteins (71 genes),
and Energy Metabolism (32 genes). Overrepresentation of the Biosynthesis of Cofactors,
Prosthetic Groups, and Carriers role category is largely due to upregulation of siderophore
biosynthetic genes. Over-representation of the Mobile and Extrachromosomal Element
Functions role category is due to upregulation of 12 genes of the prophage 01 region of the Pf-5
genome (PFL_1210 to PFL_1229; Mavrodi et al., 2009), including the gene encoding the
bacteriocin LlpA1. Fifteen of the 71 upregulated genes in the Transport and Binding Proteins
role category fall into the Cations and Iron Carrying Compounds subrole and are mostly involved
in iron uptake, while 36 genes fall into the Amino Acids, Peptides, and Amines subrole, as
described in the main text of the manuscript. The overrepresentation of the Energy Metabolism
category in the genes upregulated in a gacA mutant on seed was also due largely to the Amino
Acids and Amines subrole (17 of 32 genes), suggesting that regulation of uptake and metabolism
of these compounds is particularly important to Pf-5 growing on pea seed surfaces. Interestingly,
the Energy Metabolism role category was overrepresented in the genes downregulated in a gacA
mutant in culture. This role category also encompasses seven of the 13 genes found to be
upregulated in a gacA mutant on seed surfaces and downregulated in a gacA mutant in culture,
indicating quite different regulation and/or expression of genes in this category under the two
conditions.
Two role categories, Cellular Processes and Protein Fate, in addition to Hypothetical
Proteins, were overrepresented among genes downregulated in a gacA mutant both on seed and
in culture, due largely to secondary metabolite biosynthesis and type VI secretion system (T6SS)
genes. These two role categories were not found to be significantly overreprented in the culture
data by Hassan et al., (2010) but were significant based on our reanalysis using only 1.4-fold
regulated genes and an improved annotation of the Pf-5 genome.
The Transcription role category was overrepresented significantly in the GacA regulon of
Pf-5 grown in culture but not on seed (Fig. S1), due primarily to increased transcript levels of 28
Extracytoplasmic function (ECF) sigma factors in the gacA mutant grown in culture (Hassan et
al., 2010).
References
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Inactivation of the GacA response regulator in Pseudomonas fluorescens Pf-5 has farreaching transcriptomic consequences. Environ Microbiol 12: 899-915.
Mavrodi, D.V., Loper, J.E., Paulsen, I.T., and Thomashow, L.S. (2009) Mobile genetic elements
in the genome of the beneficial rhizobacterium Pseudomonas fluorescens Pf-5. BMC
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