Classical and omic approaches to the analysis of acid stress alfalfa nodulating rhizobia

DRAGHI, W.O.; DEL PAPA, M.F.; PISTORIO, M.; MOLINARI, M.L.; LOZANO, M.J.; GIUSTI, M.A.; BOIARDI, J. L.; WATT S.; NIEHAUS, N.; PÜHLER A.; LAGARES, A.

Buenos AIres

Congreso; 1st Annual Iberoamerican Proteomics Congress; 2007

Latinoamerican Human Proteome Organisation

The poor tolerance of several rhizobia to the hydrogen ions is considered among the main factors that restrict the development of the N2-fixing symbiosis in moderately acidic soils. This circumstance reinforces the necessity to better understand the molecular basis of the rhizobial response to acidity. To advance in this direction, and to investigate the relevance of differentially expressed protein markers, we set up N-limited continuous cultures of the model strain Sinorhizoboium meli/oiti (Sme) 2011 at different pHs and characterized their proteomes. In such experiments the extracellular pH was gradually decreased 0.2-0.5 units (± 0.05) stepwise starting from a continuous culture at pH 7.4. Results showed that: strain Sme 2011 stopped growing when reaching pHs lower than 6.1 in the chemostat (thus considered the pHlimit). 2D-gel electrophoresis-MALDI-TOF proteome analysis of strain 2011 sampled from the chemostat and from batch cultures at different pHs allowed for the identification of several proteins associated to the growth under each pH condition. We recognized 8 and 22 differentially expressed proteins associated to the acid and to the neutral condition, respectively. The up-regulated proteins under acidity included: Ppi and Tig (peptidyl-prolyl isomerases, one of them ribosome-associated), SodS (superoxide dismutase), DegP1 (protease), TufA (translation factor), Mur (murein synthesis), and other proteins involved in the biosynthesis and/or transport of small molecules. Most of the acid-induced proteins were related to the support of housekeeping activities and to the recovery of protein structure/activities, denoting a clear 'reactive behavior of the cells against the operating stress condition. A very similar pattern of over/under expression was observed for the differential proteome markers at the transcriptome level, showing a highly positive correlation between the proteome and transcriptome data under our experimental conditions. Most relevant, several singleTn5 mutants affected in the identified proteome markers showed a more acid sensitive phenotype. Thus, the collected data provided experimental support to screen and compare, at a functional level, the individual contribution of acid-induced/repressed marker to the growth of Sme under acidity.