In vivo dissection of functional domains of plant glutamyl-tRNA reductase, the regulatory key enzyme of tetrapyrrole biosynthesis
Final Report Abstract
5-Aminolevulinic acid (ALA) is the first committed substrate of tetrapyrrole biosynthesis and is formed from glutamyl-tRNA by two enzymatic steps. Glutamyl-tRNA reductase (GluTR) as the first enzyme of ALA synthesis is encoded by HEMA genes and tightly regulated at the transcriptional and posttranslational levels. GluTR stability and activity are adapted to endogenous and environmental changes by several posttranslational control mechanisms and ensure rapid and instantaneous changes in the ALA synthesis rate for optimized formation of chlorophyll and heme and avoidance of accumulated metabolic intermediates. Heme and FLOURESCENT IN BLUE LIGHT (FLU) act negatively on ALA synthesis. A GluTR-binding protein (GBP) is bound to the thylakoid membrane, acts as an anchor for GluTR and is proposed to ensure a continuous ALA formation for the production of heme. GluTR1 has been also identified as a substrate for the caseinolytic protease (Clp) in plastids. Thereby, Clp subunits and GBP interact with the N-terminal domain of GluTR, counterbalance the activity of GluTR for chlorophyll and heme synthesis and ensure a controlled GluTR degradation. The C-terminus of GluTR is the FLU-binding domain and the removal of more than 22 C-terminal amino acids suppresses the GluTR-inactivation of FLU. We used the lethal Arabidopsis hema1 mutant to express different transgenes for the synthesis of different full-length and truncated GluTR variants. These lines enabled to determine the functional role of different GluTR domains. Function and activity of GluTR were determined by analysis of enzyme activities, levels of tetrapyrrole intermediates, and transcript as well as protein levels for important steps of tetrapyrrole biosynthesis.
Publications
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(2011) An Arabidopsis GluTR binding protein mediates spatial separation of 5-aminolevulinic acid synthesis in chloroplasts. Plant Cell 23: 4476-4491
Czarnecki O, Hedtke B, Melzer M, Rothbart M, Richter A, Schroter Y, Pfannschmidt T, Grimm B
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(2011) Overexpression of HEMA1 encoding glutamyl-tRNA reductase, J Plant Physiol 168: 1372-1379
Schmied J, Hedtke B and Grimm B
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(2011) Tomato fruit photosynthesis is seemingly unimportant in primary metabolism and ripening but plays a considerable role in seed development. Plant Physiol 157: 1650-1663
Lytovchenko A, Eickmeier I, Pons C, Osorio S, Szecowka M, Lehmberg K, Arrivault S, Tohge T, Pinida B, Anton MT, Hedtke B, Lu Y, Fisahn J, Bock R, Stitt M, Grimm B, Granell A and Fernie AR
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(2014). GluTR2 complements a hema1 mutant lacking glutamyl-tRNA reductase 1, but is differently regulated at the post-translational level. Plant Cell Physiol 55, 645-657
Apitz, J, Schmied J, Lehmann MJ, Hedtke B, and Grimm B
