Supplying plant peroxisomes with energy

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ATP represents the major energy currency of the cell, driving a large number of metabolic reactions. Its availability and continuous provision has a crucial impact on cellular metabolism. Since ATP synthesis is restricted to only a few cell compartments in plants, ATP has to be transported in and out of cell compartments that are unable to synthesize it. Specific carrier proteins mediate the intracellular ATP exchange and thus are crucial for multiple metabolic pathways. In our previous funding period we characterized two ATP transport proteins in Arabidopsis that are able to import ATP into peroxisomes.In this proposal we will address the following major questions to understand the function of peroxisomal ATP import: (1) Which ATP-consuming reactions depend on peroxisomal ATP? We demonstrated that Arabidopsis plants with an impaired peroxisomal ATP import were unable to metabolize their storage oil. As a consequence the mutant seedlings were arrested in their growth. Our preliminary results also indicate that peroxisomal ATP is crucial for the degradation of the toxic metabolite propionic acid. We will perform 13C-labelling experiments and targeted metabolite analyses to elucidate the involvement of peroxisomes in this less-studied pathway. (2) What is the driving force for ATP import into peroxisomes? We demonstrated that ATP is transported in exchange with ADP or AMP. Such a counter-exchange would generate a membrane potential across the peroxisomal membrane that counteracts the continuous supply of ATP into peroxisomes. We hypothesize that the inward-directed net-transfer of negative charges during ATP import is balanced by proton transport. We will test this hypothesis in this proposal. (3) How does supply and demand of ATP impact peroxisomal metabolism? We will assess the ATP pool in Arabidopsis peroxisomes in vivo by using a GFP-based ATP sensor in diverse mutant plants. In addition, we will implement an ATP-hydrolyzing enzyme in the peroxisomal matrix to intensify the ATP sink. This will allow us to get insights in dynamic changes of the ATP pool in plant peroxisomes, which will further elucidate the impact of ATP on peroxisomal metabolism.