2012 Annual Report
1a.Objectives (from AD-416):
Identify promoters and ECF sigma factors that control expression of known and suspected virulence factors.
Characterize the subset of the transcriptome related to growth in defined medium, the induction of virulence factors, and response to iron bioavailability.
Elucidate mechanisms leading to iron-dependent expression of operons encoding virulence factors and regulators.
1b.Approach (from AD-416):
Research will employ an interdisciplinary approach involving computational biology and laboratory methods for high-throughput functional genomics and genetics. Expression studies including the use of microarrays and high-throughput reporter screens will be used to characterize the components and behavior of virulence-related pathways, especially those related to iron homeostasis. Mutants in key regulatory proteins and gene reporter systems will be used to elucidate regulatory interactions. Computational methods will be used to identify regulatory motifs, detect statistically significant correlations in gene expression, and model selected pathways. We aggressively integrate laboratory and computational approaches to genome-scale problems in order to design and implement the most effective experiments and analytical methods.
This project terminated in February 2012, (see 1907-21000-035-00D for current/ongoing activities), and during this reporting period we focused on bringing this project to conclusion. This involved writing manuscripts, and finishing the development of datasets for future projects. Pursuant to Objective 2A, we finished the development of a novel method for capturing the start of bacterial transcripts using high-throughput sequencing technologies. A paper describing this method was published. Pursuant to Objective 1, we continued our study of the iron starvation (IS) extracytoplasmic function (ECF) sigma factors in DC3000. These sigma factors, together with the Fur repressor, are critical to regulating DC3000’s uptake of extracellular iron. A paper describing one of these sigma factors, PSPTO_1203, was published.
Filiatrault, M.J., Stodghill, P., Myers, C.R., Bronstein, P.A., Butcher, B.G., Lam, H., Grills, G., Schweitzer, P., Wang, W., Schneider, D.J., Cartinhour, S.W. 2011. Genome-wide identification of transcriptional start sites in the plant pathogen Pseudomonas syringae pv. tomato str. DC3000. PLoS One. 6(12). Available: http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029335.
Swingle, B.M., Markel, E.J., Maciak, C.L., Butcher, B., Myers, C., Stodghill, P., Cartinhour, S.W., Bao, Z. 2011. An ECF sigma factor mediated cell surface signaling system in Pseudomonas syringae pv. tomato DC3000 regulates gene expression in response to heterologous siderophores. Journal of Bacteriology. 193:5775-5783.
Schechter, L.M., Valenta, J.C., Schneider, D.J., Collmer, A., Sakk, E. 2012. Functional and computational analysis of amino acid patterns predictive of type III secretion system substrates in Pseudomonas syringae. PLoS One. 7(4):1-13.
Bocsanczy, A., Schneider, D.J., Declerck, G.A., Cartinhour, S.W., Beer, S.V. 2011. HOPX1 Ea (Eop3) in Erwinia Amylovora functions as an avirulence gene in apple and is regulated by HRPL. Journal of Bacteriology. 194:553-560.