Knocking Out the Gene RLS1 Induces Hypersensitivity to Oxidative Stress and Premature Leaf Senescence in Rice

Improving a plant's level of tolerance to oxidative stress can frequently also enhance its tolerance to several other abiotic stresses. Here, a screen of a japonica type rice T-DNA insertion mutant library identified a highly oxidative stress-sensitive mutant. The line exhibited premature leaf senescence, starting at the three-leaf stage, and the symptoms were particularly severe from the five-leaf stage onwards. The leaves progressively lost chlorophyll, suffered protein degradation and were compromised with respect to their photosynthetic activity; their leaf mesophyll and bulliform cells became shrunken, and several senescence-associated genes (SAGs), senescence-associated transcription factor genes (SATFs) and autophagy-related genes (ATGs) were progressively up-regulated. The product of the gene inactivated by the mutation, identified via positional cloning, was putatively a ubiquitin-conjugating enzyme. The gene was denoted here as RLS1 (reactive oxygen species-sensitive leaf senescence1). The phenotype of plants in which RLS1 was knocked down using RNA interference was comparable to that of the rls1 mutant. A comparative analysis of the knock-out line and the wild type leaves showed that the former accumulated more hydrogen peroxide and more malondialdehyde, expressed a heightened level of superoxide dismutase activity and a decreased level of catalase activity, and exhibited an altered transcriptional profile with respect to several SAGs, SATFs and ATGs, and that these effects were magnified when the plants were exposed to oxidative stress. The product of RLS1 is presumed to be a critical component of the rice oxidative stress response and is involved in ROS (reactive oxygen species)-mediated leaf senescence.