Plant catalases: New insights into targeting and activity

Reactive oxygen species (ROS) play a key role in both stress responses and developmental pathways in all living organisms. The level of ROS molecules is controlled by different antioxidant enzymes as well as non-enzymatic antioxidants. Peroxisomal catalase is considered as one of the key antioxidant enzymes in the detoxification of hydrogen peroxide (H2O2), which is generated in peroxisomes as a result of oxidases involved in many processes but especially photorespiration and β-oxidation. In Arabidopsis, catalase exists in three isoforms of which the catalase 2 (CAT2) is the most crucial for photorespiration. The cat2-1 mutant is a photorespiratory mutant. Catalase is targeted to peroxisomes by peroxisomal targeting signal type 1(PTS1) import receptor PEX5, but its carboxy-terminal sequence does not match the normal PTS1 consensus. In Arabidopsis, there is an alternative splicing event that removes the last 18 amino acids of CAT2, and previous studies indicated importance of some amino acids in this region, in particular the last 3 amino acids, for peroxisomal targeting. Transgenic lines in which the cat2-1 mutant was transformed separately with the wild type CAT2 gene, the alternative splice variant, and a version that has a consensus PTS1 sequence ARL at the C terminus, all under the control of the native CAT2 promoter, were selected. Physiological and biochemical consequences of introducing CAT2 variants with modified C-termini into cat2-1 mutant background were studied by growing the transgenic lines under short day and long day conditions. The wild type phenotype was largely restored in all of the lines transformed with CAT2 variants, together with catalase activities and redox state. Subcellular fractionation suggests that CAT2 is targeted to the peroxisomes in all cases. To confirm that, the subcellular localisation of CAT2 variants was examined using a self-assembling split superfolder green fluorescent protein (sfGFPOPT) system. My results demonstrate that the last 18 amino acids of the CAT2 sequence are required for the targeting of CAT2 to the nucleus but dispensable for peroxisomal targeting, catalase activity and redox homeostasis. These findings provide fundamental new insights into catalase targeting and pave the way for exploration of the mechanism of catalase targeting to the nucleus and its role in non-infected plants.