Biological Sciences Department Plant sensing and responding to stress Plant Stress Physiology (PSP)   Dr. Khalid Alsharafa

ROS; reactive oxygen species • Introduction High light Light / Dark Pathogens Temperature Herbivorous Folding Drought Fluctuations in their growth environment Metabolic responses : inhibition of protein synthesis energy metabolism Molecular responses : involve interaction and cross talk with many molecules pathways These responses depend on the signals that emgating from the stimuli Heavy metals Salinity ROS ROS; reactive oxygen species

• Enzymatic / non enzymatic routes Gadjev et al. (2008)

• Scavenging of ROS in plants Developmental signals Abiotic stress Biotic stress Development PCD Stress acclimation ROS strategies Lower the rate of ROS production suppressing ROS-producing reactions different stress avoidance strategies Scavenge of ROS anti-oxidants (Carotenoids, Tocopherols, Glutathione, Ascorbate, Flavonoids) anti-oxidative enzymes (SOD, AsA-GSH cycle, Catalase, Peroxiredoxins, GPX, Apx) Mittler (2002), Mittler et al. (2004)

Water-water cycle Modified after Dietz, 2007 PSI PSII Fd e- NAD(P)+   PSII Fd e- NAD(P)+ NAD(P)H tSOD tAPX PrxQ O2 H2O H2O2 Mehler reaction O2.- PrxIIE sAPX sSOD DHA MDHA Ascorbate MDHAR NADP+ DHAR GSSG GSH GR 2CPA Modified after Dietz, 2007

Genotype X Environment Genome Genomics Genotype X Environment Translatome Transcriptome Transcriptomics Translatomics Microarrays Proteome 2D Gels + MS LC-MSMSn Proteomics Analysis Metabolome NMR, LC-MS Metabolomics

Abiotic stress, severity, Duration (min’s……..) Stimulus Abiotic stress, severity, Duration (min’s……..) LHCB PSII PQH2 cytb6/f PSI O-2 H2O2 NADPH 1O*2 Heat Kinases, phosphoralses GSH Thr JA ?? ATP EX Sensing Chl NPQ Transition state Signals Retrograde Signals Responding Gene regulation Transcription Post-transcription Translation Ribosome TFs AAAAAAAAA TFs Nucleus Candidate genes Microarray DNA metabolism Protein modification ?? AAAAAAAAA photosynthesis Transport + = _ Stress response Transcription miRNA AAAAAAAAA Stability Slicing Regulation Mechanism AAAAAAAAA AAAAAAAAA Translation Steady state Nucleo run on AAAAAAAAA AAAAAAAAA AAAAAAAA 3´ m7G uORF CPE AAUAAA 5´ 5´ UTR 3´ UTR Coding region AAAAAAAAA

• Aims of the my talk - Monitoring plants acclimation response for biotic and different exposure times of different abiotic stresses - Detection of different retrograde signalling in response to abiotic stresses - Analysis of post-transcriptional regulation at the level of enzyme activity in response to abiotic stress dose and exposure duration

- Light shift processes • Results HL exposure (0min, 10min, 30min, 60min, 180min, 360min)   Three weeks under NL Ten days under NL Ten days under LL - Light shift processes NL; Normal Light LL ; Low Light HL; High Light (~80 µmol photons * m-2 * s-1) (~80 µmol photons * m-2 * s-1) (~8 µmol photons * m-2 * s-1) (~800 µmol photons * m-2 * s-1) Morphogenic differences such as elongated petioles, small leaves and curling leaves

- Mineral deficiency leaves tip burn

- Biotic stress leaf tip burn Al-sharafa and Al-limoun (2015) Table Soil electric conductivity (EC) and pH; Data were represent means ± SD, n=3. Soil character EC (μS/cm) pH Soil depth 20cm 368±1.4 8.1±0.14 40cm 311.33±2.6 8.053±0.0094 60cm 420±0.47 8.03±0.008 Al-sharafa and Al-limoun (2015)

Figure : Chlorophyll a and b contents in L. esculentum Mill. leaves in Chlorophyll content under different mineral deficiencies Figure : Chlorophyll a and b contents in L. esculentum Mill. leaves in response to specific mineral deficiencies in comparison to complete nutrients solution grown plants at time dependent exposure (A) 48h, (B) 72h and (C) 96h (*P≤ 0.05). no indications of chlorophyll breakdown Al-sharafa (2017)

Detection of different retrograde signalling Reactive oxygen species detection Hydrogen peroxide production Elevation depend on the specific mineral deficiency Al-sharafa (2017) means ± S.D; n= 4-9, *p<0.05

Production level in parallel with H2O2 level Lipid peroxidation level Production level in parallel with H2O2 level Al-sharafa (2017) means ± S.D; n= 4-9, *p<0.05

Quantification of antioxidants Anthocyanin content Production level in parallel with H2O2 level Al-sharafa (2017) means ± S.D; n=3, *p<0.05,

Production level in parallel with H2O2 level Carotenoids content Production level in parallel with H2O2 level Al-sharafa (2017) means ± S.D; n=3, *p<0.05,

Production level in parallel with H2O2 level Total phenolic compounds content Production level in parallel with H2O2 level Al-sharafa (2017) means ± S.D; n=3, *p<0.05,

• Conclusions The involvement of sophisticated and tightly regulated stress-sensing mechanisms, efficient signal transduction pathways, efflux or compartmentation of toxic ion systems and key detoxification strategies, illustrate how fascinating is the story of plant ion homeostasis under environmental stress.

Thank you for your attention….

• References Alsharafa K, Vogel MO, Oelze M-L, Moore M, Stingl N, König K, Friedman H, Mueller MJ, Dietz K-J. 2014 Kinetics of retrograde signalling initiation in the high light response of Arabidopsis thaliana. Phil. Trans. R. Soc. B 369: 20130424. Alsharafa KY. 2017. Mineral deficiencies influence on tomato leaves: pigments, hydrogen peroxide and total phenolic compounds contents. Plant Omics. Alsharafa KY. 2017. Mineral deficiencies effect on resistance-related enzymes activities in tomato leaves. In press.