NO dioxygenase- and peroxidase-like activity of Arabidopsis phytoglobin 3 and its role in Sclerotinia sclerotiorum defense

•Arabidopsis thaliana genome encodes three distinct class of phytoglobins viz. AHb1, AHb2, AHb3.•OE-AHb3-GFP transgenic lines displayed enriched accumulation of AHb3 in guard cells with both cytoplasmic and nuclear localization.•AHb3 knockout line displayed reduced root length, fresh weight and exhibit enhanced susceptibility towards S. sclerotiorum.•Enhanced NO and ROS accumulation in AHb3 knockout lines may explain the observed phenotype.

Phytoglobin 3 appears to be ubiquitous in plants, yet there has been dearth of evidence for their potent physiological functions. Previous crystallographic studies suggest a potential NO dioxygenase like activity of Arabidopsis phytoglobin 3 (AHb3). The present work examined the in vivo function of AHb3 in plant physiology and its role in biotic stress using Arabidopsis- Sclerotinia sclerotorium pathosystem. The gene was found to be ubiquitously expressed in all plant tissues, with moderately increased expression in roots. Its expression was induced upon NO, H2O2 and biotic stress. A C-terminal tagged GFP version of the wild type protein revealed its enhanced accumulation in the guard cells. AHb3-GFP was found to be partitioned majorly into the nucleus while residual amounts were present in the cytoplasm. The loss of function AHb3 mutant exhibited reduced root length and fresh weight. AHb3 knockout lines also displayed enhanced susceptibility towards the S. sclerotiorum. Interestingly, these lines displayed enhanced ROS accumulation upon pathogen challenge as suggested by DAB staining. Furthermore, enhanced/decreased NO accumulation in AHb3 knockout/overexpression lines upon treatment with multiple NO donors suggests a potent NO dioxygenase like activity for the protein. Taken together, our data indicate that AHb3 play a crucial role in regulating root length as well as in mediating defense response against S. sclerotiorum, possibly by modulating NO and ROS levels.