Reactive oxygen species (ROS) play a key signaling role in plants and are controlled in cells by a complex network of ROS metabolizing enzymes found in several different cellular compartments. To study how different ROS signals, generated in different cellular compartments, are integrated in cells, we generated a double mutant lacking thylakoid ascorbate peroxidase (tylapx) and cytosolic ascorbate peroxidase1 (apx1). Our analysis suggests that two different signals are generated in plants lacking cytosolic APX1 or tylAPX. The lack of a chloroplastic hydrogen peroxide removal enzyme triggers a specific signal in cells that results in enhanced tolerance to heat stress, whereas the lack of a cytosolic hydrogen peroxide removal enzyme triggers a different signal, which results in stunted growth and enhanced sensitivity to oxidative stress. When the two signals are coactivated in cells (i.e. tylapx/apx1), a new response is detected, suggesting that the integration of the two different signals results in a new signal that manifests in late flowering, low protein oxidation during light stress, and enhanced accumulation of anthocyanins. Our results demonstrate a high degree of plasticity in ROS signaling in Arabidopsis (Arabidopsis thaliana) and suggest the existence of redundant pathways for ROS protection that compensate for the lack of classical ROS removal enzymes such as cytosolic and chloroplastic APXs. Further investigation of the enhanced heat tolerance in plants lacking tylAPX, using mutants deficient in chloroplast-to-nuclei retrograde signaling, suggests the existence of a chloroplast-generated stress signal that enhances basal thermotolerance in plants.Reactive oxygen species (ROS) control in-plant processes such as growth, development, stomata signaling, and biotic and abiotic stress responses (Torres and

Eukaryotic organisms evolved under aerobic conditions subjecting nuclear DNA to damage provoked by reactive oxygen species (ROS). Although ROS are thought to be a major cause of DNA damage, little is known about the molecular mechanisms protecting nuclear DNA from oxidative stress. Here we show that protection of nuclear DNA in plants requires a coordinated function of ROS-scavenging pathways residing in the cytosol and peroxisomes, demonstrating that nuclear ROS scavengers such as peroxiredoxin and glutathione are insufficient to safeguard DNA integrity. Both catalase (CAT2) and cytosolic ascorbate peroxidase (APX1) play a key role in protecting the plant genome against photorespiratory-dependent H 2 O 2 -induced DNA damage. In apx1/ cat2 double-mutant plants, a DNA damage response is activated, suppressing growth via a WEE1 kinase-dependent cell-cycle checkpoint. This response is correlated with enhanced tolerance to oxidative stress, DNA stress-causing agents, and inhibited programmed cell death.Arabidopsis | stress tolerance | hydrogen peroxide R eactive oxygen species (ROS) are toxic molecules continuously produced in cells during aerobic metabolism. In plants ROS are produced mainly in peroxisomes during photorespiration, in chloroplasts during photosynthesis, and in mitochondria during respiration (1, 2). Unless detoxified by specialized enzymes and low molecular antioxidants, ROS can lead to protein, lipid, and DNA oxidation and to cell death (1, 2). Plants contain a large network of genes encoding different pathways involved in ROS scavenging and production, with a key role in managing the overall steady-state level of ROS in cells (2). Similar to genotoxic agents or ionizing radiation, ROS-derived DNA oxidation leads to altered bases and damaged sugar residues, resulting in DNA single-and double-strand breaks (3, 4). Strand breaks trigger a DNA damage response (DDR) by inducing the expression of molecular markers associated with DNA damage repair, such as poly(ADP ribose) polymerase (PARP), RAD51, and BREAST CANCER (BRCA) family members (5-8). Upon DNA stress, the ataxia telangiectasia-mutated (ATM) and the ataxia telangiectasia and Rad3-related (ATR) signaling kinases are activated and lead, via the WEE1 serine/ threonine kinase, to a transient cell-cycle arrest that allows cells to repair DNA before proceeding into mitosis (9). Although oxidative DNA base damage has been shown to initiate a DDR in mammalian and yeast cells (10, 11), reports in plants on either the sources of oxidative stress that cause DNA damage or the subsequent induction of a DDR directly through ROS remain scarce (3,12,13). Until now, DNA damage and DDR in plants were studied mainly in response to exogenously applied DNAdamaging agents such as bleomycin and hydroxyurea or ionizing irradiation (9, 14, 15).Protection against damage caused by ROS traditionally has been attributed to enzymes with ROS-detoxifying activities (16), and mutants lacking a particular ROS-scavenging enzyme were considered more sensitive to oxidative stress ...

Over 13% of all genes in the Arabidopsis thaliana genome encode for proteins classified as having a completely unknown function, with the function of >30% of the Arabidopsis proteome poorly characterized. Although empirical data in the form of mRNA and proteome profiling experiments suggest that many of these proteins play an important role in different biological processes, their functional characterization remains one of the major challenges in modern biology. To expand the annotation of genes with unknown function involved in the response of Arabidopsis to different environmental stress conditions, we selected 1007 such genes and tested the response of their corresponding homozygous T-DNA insertional mutants to salinity, oxidative, osmotic, heat, cold and hypoxia stresses. Depending on the specific abiotic stresses tested, 12-31% of mutants had an altered stress-response phenotype. Interestingly, 832 out of 1007 mutants showed tolerance or sensitivity to more than one abiotic stress treatment, suggesting that genes of unknown function could play an important role in abiotic stress-response signaling, or general acclimation mechanisms. Further analysis of multiple stress-response phenotypes within different populations of mutants revealed interesting links between acclimation to heat, cold and oxidative stresses, as well as between sensitivity to ABA, osmotic, salinity, oxidative and hypoxia stresses. Our findings provide a significant contribution to the biological characterization of genes with unknown function in Arabidopsis and demonstrate that many of these genes play a key role in the response of plants to abiotic stresses.