Regulation of expression and interactions between glutamyl-tRNA reductase and glutamate-1-semialdehyde aminotransferase in Chlamydomonas reinhardtii

5-Aminolevulinic acid (ALA) is the first committed precursor in the tetrapyrrole biosynthesis pathway. In plants, algae, and most bacteria, ALA is generated from glutamate. First, glutamyl-tRNA synthetase activates glutamate by ligating it to tRNAGlu. Activated glutamate is converted to glutamate 1-semialdehyde (GSA) by glutamyl-tRNA reductase (GTR). Finally, GSA is rearranged to ALA by GSA aminotransferase (GSAT).; In the green alga Chlamydomonas reinhardtii, GTR and GSAT were only found in the chloroplasts. The levels of both proteins and mRNAs are equally abundant in cells growing in continuous dark or light. In synchronized cells, GTR and GSAT were present at all phases of the cycle. The GTR mRNA level increased in the light and peaked about 2 fold at 2 h into the light phase, and GTR protein levels also increased 2 fold at 4--6 h into the light phase. In contrast, the GSAT mRNA level increased several fold at 2 h into the light phase and the level of GSAT protein remained constant in the light and dark phases. Under all growth conditions, the cells contained significantly more GSAT than GTR. Our results indicate that the rate of chlorophyll synthesis in C. reinhardtii is not directly controlled by the expression levels of the mRNAs for GTR or GSAT, or by the cellular abundance of these enzymes.; GTR and GSAT form a complex that allows channeling of GSA between the enzymes. In vitro results indicate that GTR and GSAT enzymes specifically interact. In vivo results also demonstrate that GTR and GSAT are components of a high molecular-mass complex. There is also competition between wild-type and mutant GSAT for binding to GTR and channeling GSA from GTR to GSAT. Further evidence supporting kinetic interaction of GTR and GSAT is the observation that both wild-type and mutant GSAT stimulate glutamyl-tRNA-dependent NADPH oxidation by GTR.