Production of Reactive Oxygen Intermediates (O2˙−, H2O2, and ˙OH) by Maize Roots and Their Role in Wall Loosening and Elongation Growth

Cell extension in the growing zone of plant roots typically takes place with a maximum local growth rate of 50% length increase per hour. The biochemical mechanism of this dramatic growth process is still poorly understood. Here we test the hypothesis that the wall-loosening reaction controlling root elongation is effected by the production of reactive oxygen intermediates, initiated by a NAD(P)H oxidase-catalyzed formation of superoxide radicals (O₂^˙−) at the plasma membrane and culminating in the generation of polysaccharide-cleaving hydroxyl radicals (^˙OH) by cell wall peroxidase. The following results were obtained using primary roots of maize (Zea mays) seedlings as experimental material. (1) Production of O₂^˙−, H₂O₂, and ^˙OH can be demonstrated in the growing zone using specific histochemical assays and electron paramagnetic resonance spectroscopy. (2) Auxin-induced inhibition of growth is accompanied by a reduction of O₂^˙− production. (3) Experimental generation of ^˙OH in the cell walls with the Fenton reaction causes wall loosening (cell wall creep), specifically in the growing zone. Alternatively, wall loosening can be induced by ^˙OH produced by endogenous cell wall peroxidase in the presence of NADH and H₂O₂. (4) Inhibition of endogenous ^˙OH formation by O₂^˙− or ^˙OH scavengers, or inhibitors of NAD(P)H oxidase or peroxidase activity, suppress elongation growth. These results show that juvenile root cells transiently express the ability to generate ^˙OH, and to respond to ^˙OH by wall loosening, in passing through the growing zone. Moreover, inhibitor studies indicate that ^˙OH formation is essential for normal root growth.