Research project

Metabolite allocation in developing seed and flower: non invasive imaging and modeling approaches

Seed filling parameters are associated with agronomic traits. Thus, the mechanisms underlying sucrose delivery to the developing seed are a prominent topic of crop research. While the transport route is hidden inside the seed and barely available for destructive study, our approach allows the non-invasive imaging of sucrose distribution within the seed tissues (pea: Melkus et al, 2009), monitoring of sucrose allocation within the seed (barley: Melkus et al., 2011) and conversion of sucrose to amino acids. Primary synthesis of alanine is localised to central region of starchy endosperm (Rolletschek et al., 2011). The analytical method of choice is based on high-field nuclear magnetic resonance  (NMR) and a custom made 13C ⁄ 1H double resonant coil. Development of NMR technology is a joint project with Würzburg University (Prof. P. Jakob, Germany).

 

13C Metabolic Imaging of sucrose allocation and conversion of sucrose to amino acids using geHMQC: (A)1 H-reference image of the barley caryopsis. (B) Allocation of 13C-Suc after 12 h of feeding with 100 mM 13C-Suc.(C) Ala distribution after 12 h of feeding with 100 mM 13C-Suc. See for details: [link]Rolletschek et al. 2011 

 

Using this method we were able to demonstrate in vivo effect of downregulation of Jekyll gene on seed filling in barley (Melkus et al., 2011), which was in accordance with our hypotheses about the role of Jekyll in grain development (Radchuk et al., 2006). This also shed light on the role of Jekyll in barley flowers (Radchuk et al., 2012).

Sucrose monitoring can be integrated with an in silico flux balance analysis (FBA) representing a powerful platform for non-invasive study of seed filling in crops (Rolletschek et al., 2011). Currently the region-specific metabolism in the endosperm was characterised by metabolite profiling and by flux balance analysis. In the central region of the endosperm, the TCA cycle was shifted to a non-cyclic mode, accompanied by elevated glycolytic flux and the accumulation of alanine. The 13C-alanine gradient can be used as an in vivo marker of the occurrence of hypoxia. The metabolic compartmentation within the endosperm is advantageous for both the grain's carbon and energy economy, with a prominent role being played by alanine aminotransferase. 

In general, the combination of MRI with metabolic modelling offers tremendous opportunities for the non-invasive analysis of metabolic compartmentation in plants.

Funding:  the German Federal Ministry of Education and Research, Deutsche Forschungsgemeinschaft.

Localisation of Jekyll expression in barley flowers and caryopsis using GFP-reporter, see for details Radchuk et al, 2006; 2012. Cooperation with Dr. T. Rutten, Dept. of Physiology and Cell Biology, Molecular Plant Nutrition

 

Melkus G, Rolletschek H, Radchuk R, Fuchs J, Rutten T, Wobus U, Altmann T, Jakob P, Borisjuk L. (2009). The Metabolic Role of Legume Endosperm: A Non-invasive Imaging Study. Plant Physiology 151: 1139-1154.

 

Melkus G, Rolletschek H, Fuchs J, Radchuk V, Grafahrend-Belau E, Sreenivasulu N, Rutten T, Weier D, Heinzel N, Schreiber F, Altmann T, Jakob P, Borisjuk L. (2011). Dynamic 13C/1H NMR imaging uncovers sugar allocation in the living seed. Plant Biotechnology Journal 9: 1022-1037.

 

Rolletschek H, Melkus G, Grafahrend-Belau E, Fuchs J, Heinzel N, Schreiber F, Jakob PM, Borisjuk L. (2011). Combined non-invasive imaging and modelling approaches reveal metabolic compartmentation in the barley endosperm. Plant Cell 23: 3041–3054.

 

Radchuk V, Kumlehn J, Rutten T, Sreenivasulu N, Radchuk R, Rolletschek H, Herrfurth C, Feussner I, Borisjuk L. (2012). Fertility in barley flowers depends on Jekyll functions in male and female sporophytes. New Phytologist (in press).

 

Radchuk V, Borisjuk L, Radchuka R, Steinbiss H-H, Rolletschek H, Broeders S, Wobus U. (2006). Jekyll encodes a novel protein involved in the sexual reproduction of barley. Plant Cell 18: 1652-1666.