Final Report Summary - PLANT-MEMORY (Role of Pc-G and Trx-G genes in the cellular memory of extrinsic signals in plants) PROJECT OBJECTIVES:The funding period of the project “Plant-Memory” has been recently concluded. Central subject of the project was the “cellular memory of extrinsic signals” in plants, i.e. the epigenetic inheritance of extrinsic, environmental cues like abiotic and biotic stress. Due to the emerging importance of stress adaptation in plants, the findings of Plant-Memory will likely be highly relevant beyond basic research. Understanding stress memory of plants has significant direct and indirect implications for applied sciences. As it becomes more and more clear that global warming causes huge damage in the agriculture of Europe, research in this area is likely to lead to new stress adaption strategies and so, in a long-term perspective, to better crop yields, and increased food quality and safety.As Polycomb-group (Pc-G) proteins, which are modifying chromatin and are involved in setting the repressive histone modification Histone H3 Lysine 27 trimethylation (H3K27me3), and Trithorax-group (Trx-G) proteins, which are involved in setting the active histone mark H3K4me3, are central epigenetic regulators of developmental processes and target stress responsive genes, we assumed that they may also be essential for memorizing environmental cues. The five main objectives of the project were: (1) Identification of stress-induced H3K27me3 of Pc-G target genes, (2) Transcriptional profiling of Pc-G depletion before and after stress, (3) Bioinformatic analysis of the whole-genome ChIP and transcription data - Identification of the stress-specific Pc-G target genes, (4) Downstream characterisation of stress-specific Pc-G target genes, and (5) Investigation of a putative trans-meristematic stress memory. Originally, UV light was chosen as the experimental stress inducer because it seemed easy to handle and UV-B constitutes a natural stress factor that is currently increasing in importance due to the depletion of the atmospheric ozone layer and global warming. Nevertheless, the researcher also analysed the priming effects of drugs and the role of phytopathogens on memory effects in the plant in the context of systemic acquired resistance (SAR). SAR was studied as after encountering biotic stress (e.g. the pathogen Pseudomonas syringae pv maculicola (Psm)), plants often memorise the previous challenge and their defence genes react faster and stronger during a secondary stress. Even if the infection is localized, plants often acquire systemic immunity to further infections in tissue distal from the infection site. Thus, SAR comprises a natural memory phenomenon in plants . Interestingly, UV stress signalling and SAR have somame common target genes. WORK PERFORMANCE, MAIN RESULTS AND THEIR POTENTIAL IMPACT AND USE:In the first six month of the project, the researcher tested numerous UV-B treatments and experimental procedures which now resulted in the identification of optimal conditions for the experiments. Using the RNA sequencing (seq) technology, the researcher was able to identify 231 candidate genes showing significant changes of mRNA expression in response to UV-B treatment and enhanced expression by a second treatment (memory effect) (1). The primary bioinformatic analysis was carried out by the researcher himself on a CLC-genomics workbench that was available at the Max Planck Institute for Plant Breeding Research in Cologne (3). The researcher started with the downstream characterisation of the candidates, which are Pc-G target genes (4). The functional relevance of the Pc-G proteins was tested comparing responses of Arabidopsis seedlings when Pc-G activity was severely depleted before and after stress induction using iCLF (inducible CLF-GR clf swn, line with flexible Pc-G activity), emf2-10 vrn2-1 double and clf single Pc-G mutants (2). The UV-B resistance of plants after initial priming by UV-B or drugs was investigated (1,4,5). Furthermore, the researcher studied the change of chromatin modifications at the loci of key regulators of SAR. Plant-Memory has produced several research results of high quality which subsequently will be published in international, peer-reviewed journals. The key findings are summarised below::- The Researcher identified several genes, which are primed by UV-B treatment, using literature and databases, single gene approaches but also whole genome approaches (RNA seq) (1,2). These candidate genes encode (amongst others) transcription factors as well as kinases, which themselves regulate likely downstream target genes. Notable progress has been made on the downstream analysis of the candidates (2,4) but the characterisation is not finished yet.- In cooperation with other laboratory members, the researcher investigated the role of a new chromatin mark in stress memory, which function was completely unknown in plants. The cooperation partners were able to demonstrate that the global abundance of the novel mark strongly increases in the plants after treatment with UV-B or a known priming drug.- In cooperation with the laboratory of Prof Jürgen Zeier (HHU Duesseldorf), the researcher investigated changes of the chromatin marks at the loci of central players of SAR. The ChIP results suggest that repression mark levels at the investigated SAR response genes do not change in systemic leaves in response to the Psm treatment. Interestingly, systemic leaves of Psm treated plants, show increased levels of an active mark at SAR response genes. However, further investigation will be needed to test whether it only reflects increased expression levels or contributes to real memory effects. However, some histone methylases and demethylases mutants show weaker SAR response suggesting a mechanistic role of histone modifications in the SAR response.Publications, regarding these three main results, are scheduled within the next six to 18 months.On the long term, the findings of Plant-Memory and its spin-off projects will therefore likely contribute directly and indirectly to several objectives of the European Research Area such as Life Sciences, Genomics and Biotechnology for Health, Food Quality and Safety, and Sustainable Development.ADDRESS OF THE PROJECT PUBLIC WEBSITE: http://www.entwicklungsepigenetik.hhu.de/unser-team/ralf-mueller-xing/plant-memory-english.html
