IN INOCULANTS Nodulaid - 17th International Nitrogen Fixation ...

17 th International Congress on Nitrogen Fixation
Fremantle, Western Australia
27 November – 1 December 2011
Session Details: Tuesday 29 November 2011
Authors:
The symbiosome membrane (SM) is the interface between the plant and bacteria in the legume-rhizobium
symbiosis. This plant-derived membrane separates the symbiotic form of the rhizobia, from the plant cytoplasm
and controls both the type and amount of solutes exchanged between the two partners. In exchange for nitrogen
fixed by the bacteroid, the plant provides reduced carbon, probably in the form of malate, to the bacteroids, as
well as other solutes including iron, sulphate, zinc and possibly amino acids. We are investigating transport
proteins on the SM that regulate this exchange in soybean nodules using a proteomic approach and nodule
transcriptome data. Candidates that potentially encode malate, iron and amino acids are a major focus.
Proteomic analysis of the SM involved trypsin digestion of total SM proteins, separation of peptide fragments by
liquid chromotography (LC), and analysis with tandem mass spectrometry (MS/MS). Of the 71 proteins identified
in all three biological replicates, 33 are predicted to be integral membrane proteins. A number of proteins known
to be associated with the SM, such as nodulin 26 (an aquaporin), ATPase, symbiotic sulphur transporter (sst1),
Rab7, nodulins 24 and 53, were identified. Many of the identified SM proteins are highly expressed in nodules
compared to other tissues (Libault et al. 2010; Severin et al. 2010). We are currently assessing the importance of
some integral membrane proteins of unknown function through an RNAi approach and confirming the location of
proteins on the SM using reporter gene fusions. Candidate transporters confirmed to be localised to the SM will
be functionally characterised in heterologous expression systems.
We have also characterised a transporter, GmYSL1, with homology to Yellow Stripe Like (YSL) proteins that are
implicated in iron transport in other plants. The gene complements growth of the yeast iron transport deficient
mutant, fet3fet4, on media containing low concentrations of ferric citrate. Fusion of the coding region to GFP
confirmed the localisation of this protein on the SM. Its expression in nodules is up-regulated in response to iron
and this and its location suggest it may be involved in sequestration of iron in the symbiosome.
Libault M Plant J. 2010 63:86
Severin AJ BMC Plant Biol. 2010 10:160
Concurrent Session 5 – Bacteroids & Symbiosomes
1100 - 1230
Patrick Loughlin 1 , Victoria Clarke 1 , Chi Chen 1 , Yihan Qu 1 , Joanne Castelli 3 , Martha
Ludwig 3 , David Day 2 and Penelope Smith 1
1 School of Biological Sciences, Macleay Bldg (A12), University of Sydney, Camperdown
NSW, 2006. 2 School of Biological Sciences, Flinders University, SA, Australia. 3 School of
Biomedical, Biomolecular and Chemical Sciences (M310), University of Western
Australia, Crawley WA 6009.
Presentation Title: Identification of transport proteins on the symbiosome membrane in soybean
Presentation Time: 1120 - 1140
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2011