Group Leader: Prof. Dr. Ingo Schubert
The structure of plant chromosomes (including physical gene mapping) and the mechanisms of karyotype evolution and chromosome mutagenesis are investigated by genetic, cytogenetic and molecular methods. In particular, structure and function of distinct chromatin domains and their dynamic epigenetic modification in correlation to nuclear processes such as replication and transcription are studied.
A collection of mutant karyotypes of the field bean developed at Gatersleben was used to simulate experimentally essential processes of natural karyotype evolution such as fusion/fission of chromosomes. Crossing of specifically reconstructed karyotypes revealed a hitherto unknown mechanism of evolutionary alteration of diploid chromosome numbers. Furthermore, an upper tolerance limit of chromosome arm length (~half of the spindle axis extension) has been defined. For longer arms, mitotic separation of sister chromatids becomes increasingly incomplete and results in impaired fertility and viability of carriers via genomic instability (deletions) followed by apoptotic processes.
Telomere structures and their evolution were studied in vascular plants. A new alternative telomeric structure was found on chromosome ends of Allium relatives. Genomic in situ hybridization (GISH) is used to verify interspecific hybrids or introgressions within various plant families.
DNA of microdissected or flow sorted chromosomes/chromosome regions was used for physical mapping of gene and marker sequences via PCR with sequence-specific primers.
Chromosome-specific DNA probes are applied for in situ labelling of homologous/homoeologous chromosome regions ('chromosome painting') in order to specify positions of chromosome rearrangements, to reveal homoeology/synteny between chromosomes/linkage groups of more or less related species and to study chromosome territories during different cell cycle and developmental stages. The first successful painting outside mammals and birds we did on Drosophila chromosomes. Using pools of contiguous genomic BAC clones of A. thaliana, we established chromosome painting for Arabidopsis and related species. This approached allowed us i) to elucidate the mechanism of chromosome number reduction from n=8 to n=5 in the course of evolution towards A. thaliana and ii) to study interphase arrangement of chromosomes and defined chromatin domains among Arabidopsis relatives. Recently we focussed on sister chromatid alignment and condensation during cell cycle and developmental stages. We study these phenomena in mutant backgrounds and under genotoxin exposure to better understand their mechanisms and biological meaning.
To elucidate plant centromere structures, we studied the sequence organisation of barley centromeres and found a conserved Ty3-gypsy retroposon-like sequence and a barley-specific GC-rich satellite repeat to be the main components of the seven active barley centromeres. Nevertheless, together with T.R. Endo's group from Kyoto University and with A. Houben, it could be shown by formation of novel centromeres that these sequences are neither sufficient nor necessary for functional centromeres. We trace plant kinetochore proteins in vivo by fluorescent tags in transgenic Arabidopsis plants and test their functionality applying complementation of T-DNA insertion mutants and RNAi approaches. We could show that the centromere-specific histone CENH3 is deposited during G2, in parallel with the splitting into sister kinetochores.
Chromatin modifications (acetylation, methylation, phosphorylation) in specific chromosome domains of several plant species are studied by immunostaining and FISH (fluorescence in situ hybridization) on mitotic chromosomes and nuclei of defined interphase stages in correlation with replication and transcriptional activity, to elucidate chromatin structure and heterochromatin formation in interphase nuclei, including evolutionary aspects.