Transcriptional control of ribosomal genes is a key regulatory element in the reaction of cells to environmental cues. We study the spatial arrangement of the different epigenetically defined populations of ribosomal genes, transcriptionally active, silent and primed ribosomal genes in relation to the structural correlate of ribosomal gene expression, the nucleolus. These studies are conducted under conditions favorable for ribosome production and cell growth as well as under nutrition deprivation and treatment with inhibitors of nucleolar activity. In the latter respect, we conduct a detailed study about the influence of a novel anti-cancer drug, CX-5461 on alterations of ribosomal gene arrangement and nucleolar structure.
We employ a predominately microscopic approach including time-lapse microscopy of dCas-tagged ribosomal genes and correlated light- and electron microscopy (CLEM).
We have the exciting and novel opportunity to study the entire life span of an organism by exploiting the model system Nothobranchius furzeri that is maintained in our in-house Austrian Notho Project facility (see Group Pusch). In more detail, we study chromatin modifications that are specific for transcriptionally active versus modifications that are typical for inactive chromatin during defined stages of the entire life cycle representing embryonic development, childhood, adolescence, adulthood and age. One part addresses the changes of expression of histone deacetylases (HDACs), especially class I and class III (Sirt) enzymes that play crucial roles in histone modifications regulating the transcriptional activity of chromatin. Overlapping with project 1 we will also investigate changes in the ribosome production and thus nucleolar function across the life span. An important part represent our studies of age-related alterations of structural tissue homeostasis. Importantly, we investigate the tissue dynamics of intestinal stem cells in the gastrointestinal tract during the lifetime of N. furzeri.
Here we employ a wide variety of biochemical, molecular and microscopic techniques and can produce genetically modified animals to study loss-of-function effects.
Several proteins of the inner nuclear membrane (INM) belong to the LEM-domain (Lap2-Emerin-MAN1) protein family. The LEM-domain binds BAF, which itself interacts with DNA. LEM-domain proteins have developmental functions, as exemplified by mutations in emerin causing Emery-Dreifuss Muscular Dystrophy. They also interact with transcriptional repressors. In order to examine the collective influence of the LEM-domain on chromatin architecture and post-mitotic nuclear envelope formation we are creating cell lines deficient for one or several LEM-domain proteins of the INM using the CRISPR/Cas methodology and time-lapse microscopy.