Iron homeostasis in root nodules

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Janneke Balk leads a research group studying the biogenesis of metal proteins in plants, algae and fungi. She has previously identified several proteins that are involved in the assembly of iron-sulfur cofactors. As one of her core projects, Dr Balk will investigate metal homeostasis during nodule development, in both the plant and bacterial partners of the symbiotic interaction. Several key enzymes required for biological nitrogen fixation require large amounts of iron and molybdenum, but little is know how these metals are acquired and correctly distributed.

Objectives
1. Confirm that the host plant provides the metals, which has, surprisingly, not been demonstrated thus far. Or, in other words, we need to exclude that the bacteria bring a hoard of iron and molybdenum with them when invading the plant roots. We will measure the metal content of free-living Rhizobia and Rhizobia at different stages of the colonization process (exposure to root exudates, entering the root hair cells, migration to the cortex, early nodules and N2-fixing nodules), using a number of techniques such as mass spectrometry, colorimetric assays and in-situ Fe staining.
2. Identify plant and bacterial genes that are up-regulated in nodules AND are putatively involved in metal homeostasis. Transcriptomics data sets of pea nodules, before and after inoculation, are available in collaboration with Phil Poole.
3. Functional characterization of selected genes identified in (2), using (i) a genetic strategy and (ii) recombinant protein techniques. Rhizobium genes will be deleted and the strains tested for their ability to infect the host plant and/or fix nitrogen. The plant genes will be studied using gene silencing or available insertion mutants. The same bacterial and plant genes will be expressed in E. coli, purified, and their biochemical function studied in-vitro.

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