Genomics and Transcript Profiling
The Central Infrastructure Group Genomics and Transcript Profiling led by Dr. Dirk Hincha provides on the one hand services in high-throughput gene expression and enzyme activity analysis for the whole institute. On the other hand, the group also performs scientific research on plant abiotic stress tolerance, in the areas of freezing, drought, desiccation, heat and salt stress, using mainly Arabidopsis thaliana, rice and potato as the experimental species.
Molecular mechanisms of Arabidopsis cold acclimation, deacclimation and cold memory
As part of the CRC 973 "Priming and Memory of Organismic Responses to Stress" (www.sfb973.de) that is funded by the German Research Foundation (DFG) we are exploring how plants can remember a previous cold exposure and then react differently to a second cold stress. We are interested, on the one hand, in the "memory phase", i.e. the time after cold acclimated (primed) plants are transferred back to warm growth conditions and partially or completely lose their freezing tolerance. On the other hand, we are characterizing the responses of plants to a second (triggering) cold treatment, after they have "forgotten" the first cold exposure. The figure shows a schematic representation of the experimental approaches taken in this project. Gene expression studies provide us with a selection of candidate transcription factors and other regulatory proteins that are related to such memory functions. The role of these candidates is tested using gene knock-out plants and functional characterization of confirmed candidates is performed using a combination of physiological, biochemical and genetic approaches.
The role of LEA (late embryogenesis abundant) proteins in cellular dehydration tolerance
LEA proteins are synthesized late during embryogenesis in plant seed development. Structurally related proteins have also been found in vegetative plant tissues under various stress conditions and in desiccation tolerant invertebrates and microbes. They thus seem to be universal dehydration stress proteins in living cells, with the exception of vertebrates. Their biological functions in stress tolerance, however, are unclear. Most of these proteins have no stable secondary structure in solution, i.e. they belong to the large group of intrinsically disordered proteins (IDPs). We are concentrating our efforts on the largest LEA protein family (Pfam LEA_4) that has 18 members. We produce proteins of interest recombinantly in E. coli and use mainly fluorescence, far-UV circular dichroism (CD) and Fourier-transform infrared (FTIR) spectroscopy for structural and functional characterization. To study the interactions with membranes, we use artificial lipid vesicles (liposomes) that can be produced with a variety of user-defined lipid compositions. Using these approaches we have shown that two of our candidate proteins (COR15A and B) stabilize membranes during freezing through a folding and binding mechanism that is triggered by molecular crwoding. In addition, we use genetic approaches such as RNA interference, overexpression or GFP tagging to study the function and localization of the proteins in planta.
Natural genetic diversity and molecular mechanisms of high night temperature tolerance in rice
Global warming has an increasing influence on the productivity of crop plants. In the past century a stronger increase in daily minimum compared to maximum temperatures has resulted in an asymmetric global warming. However, only a few studies have so far investigated the influence of high night temperatures (HNT) on crop physiology. We are analyzing the effects of HNT on different rice cultivars. A clear distinction between tolerant and sensitive cultivars was found based on leaf chlorosis estimates. HNT also results in a high degree of flower sterility and subsequent reduction in grain yield. We are investigating the molecular consequences of HNT and the basis of the varying tolerance using a wide range of physiological approaches in addition to metabolite, enzyme activity and gene expression profiling. In addition, we are using quantitative trait locus (QTL) mapping to identify genomic regions of rice that are responsible for HNT tolerance in particular cultivars.
Molecular mechanisms of tolerance in rice under combined heat and drought stress
Rice is predominantly grown in the tropics and sub-tropics where temperatures are high throughout the growth season. As long as plants have sufficient water supply, they can limit shoot temperature by transpirational cooling. However, when water becomes limiting, drought conditions are invariably accompanied by heat stress. Heat and in particular combined heat and drought stress leads to massive yield reduction, in part due to pollen sterility. However, different rice cultivars show different susceptibility to yield loss under such conditions. We have preformed heat and drought stress experiments in fields in the Philippines in collaboration with the International Rice Research Institute (http://irri.org). The upper picture in the figure shows a well-watered control plot, while the bottom picture shows a drought-stress plot. Samples from these plants, including tolerant and sensitive cultivars, were transferred to the MPI and are currently analyzed using transcriptomic and metabolomic approaches to understand the physiological basis of heat and drought stress tolerance and to identify molecular markers that could be used by breeders to select for more stress tolerant varieties in the future.