Subcellular localization and nitric oxide-scavenging activity of plant hemoglobins

Abstract

Symbio􀆟c hemoglobins provide O2 to N2-fixing bacteria within legume nodules. However, the func􀆟ons of nonsymbio􀆟c hemoglobins or phytoglobins (Glbs) are less defined. Three Glb classes can be dis􀆟nguished based on phylogene􀆟c and biochemical analyses and may coexist in plant 􀆟ssues: class 1 Glbs have extreme O2 affinity and are induced by hypoxia; class 2 Glbs have moderate O2 affinity and are precursors of leghemoglobins; and class 3 have low O2 affinity and high sequence homology with bacterial truncated hemoglobins. Immunogold labeling combined with confocal microscopy of Glbs tagged with GFP at the C-terminus was used to determine the subcellular localiza􀆟ons of Glbs in the model plants Arabidopsis and Lotus japonicus. To this end, we used overexpressing and knockout or silenced lines of Arabidopsis, performed quan􀆟ta􀆟ve immunolabeling, and monitored the GFP-tagged proteins in leaf cells and protoplasts.