Stress-induced nucleosome dynamics in plants

Chromosomal DNA in eukaryotes is organized into chromatin with nucleosomes as basic subunits. The histones in the nucleosomal core are highly conserved proteins, with several variants that can carry specific post-translational modifications. Nucleosome position and composition are important elements of epigenetic information that, together with the genomic DNA, can be inherited during somatic and sexual propagation. DNA sequence information is copied faithfully during replication and hence represents the most conservative layer of inheritance. In contrast, arrangement, composition, and modification of the nucleosomes can vary during growth, or between tissues and individual cells, thereby affecting gene expression patterns and contributing substantially to differentiation and development. Besides these programmed changes at nucleosomes, recent data from one partner lab have shown that in plants, also extended exposure to high temperature can cause extensive and global dissociation of nucleosomes from DNA, connected with transcriptional activation of otherwise silent genes and transposons. Although these gene expression changes are transient, they result in a lasting effect on the organization of heterochromatin in nuclei of differentiated leaf cells. Reassembly of nucleosomes requires the chromatin assembly factor (CAF) complex, a well-conserved histone H3-H4 chaperone. In the context of the proposed project, we want to build on this work and investigate the molecular processes occurring during nucleosome dis- and re-association and the nature of the persistent effects on nuclear chromatin organization. To this end, we will focus in particular on the role of histone H3-H4 chaperones in the removal of old and supply of new histones as well as on the dynamics and chromatin enrichment of different H3 variants. Further, we want to investigate the extent to which the heat stress-induced changes also affect meristematic tissue. This is an important question as environmentally induced epigenetic changes are presumed to affect subsequent generations, but evidence on the molecular level is lacking. We will combine one team`s expertise in the molecular biology of histone variants, histone chaperones, and cytological analysis of nuclear organization with the know-how of the partner lab in stress application, analysis of global transcription, and nucleosome occupancy. The project is expected to provide insight into the mechanism of an interesting and so far under-investigated layer of epigenetic control and into the potential impact of adverse environmental conditions on inheritance. The project leaders have an established record of fruitful collaboration and will involve several young scientists in this work, for whom the research is an excellent training platform in a scientifically stimulating and well-equipped academic surrounding.

The length of DNA molecules exceeds the diameter of cell nuclei by several orders of magnitude. Despite the necessary compaction, DNA must be accessible for gene expression, replication and repair of damage. At the basis of the intranuclear spatial DNA organisation is its winding around spherical protein complexes, the nucleosomes, consisting of 4 x 2 histone subunits. The position of the nucleosomes on the DNA, their composition and diverse chemical modifications of the subunits represent important epigenetic information that determines the accessibility of DNA and the activity of the genes at corresponding locations. Using the model plant Arabidopsis, work in this FWF-funded project has addressed the question, whether and how heat stress influences the density and position of the nucleosomes along the genome. Indeed, at elevated temperatures that lead to growth arrest, many genes lose nucleosomes at their regulatory elements, and many of these genes become active. These genes include transposable elements that are kept silent under growth-promoting temperatures. Heat stress can therefore open a window of opportunity for their propagation within the genome. Heat stress does not only change the pattern of gene expression but alters the architecture of the whole nucleus. With adapted microscopy techniques that allow to follow individual nuclei in living root cells over long periods, it could be demonstrated that previously spindleshaped nuclei turn spherical under heat stress, and dense chromosomal regions become loose. The microscopic setups for life imaging of plant cells will be applicable to many other biological questions. Remarkably, if the heat stress is over, the plants can restore the original arrangement. This includes return to the regular gene expression, the nucleosome association and the nuclear structure, which all, after the recovery period, resemble that of non-stressed control plants. The results indicate that the observed processes reflect active processes of adaptation to extreme stress conditions, which in plants are probably especially important due to their sessile life style. Whether, under certain circumstances, externally triggered changes in a plants nucleus can become permanent and be inherited by progeny is an open and much discussed question. If so, the changes must affect the stem cells that form the seeds for the next generation. Attempts to isolate these rare stem cells and to investigate their epigenetic information are ongoing.