Influence of nuclear architecture on gene vector persistence and expression

• Antragsteller: Privatdozent Dr. Steffen Dietzel
• Fachliche Zuordnung: Hämatologie, Onkologie
• Förderung: Förderung von 2006 bis 2010
• Projektkennung: Deutsche Forschungsgemeinschaft (DFG) - Projektnummer 22792375

This grant application deals with aspects of gene vector persistence and integration. For some vectors a number of integration sites have been mapped to exact nucleotide positions. The virusspecific integration characteristics delineated in these studies suggest vector-specific risk potentials for gene therapy. However, the nuclear positions and possible dynamics of integration loci have not been investigated so far. Here we wish to investigate the three-dimensional nuclear organization of gene vectors, their integration loci and changes in the large-scale chromatin structure caused by transcriptional activation. A major goal is to determine topological requirements or preferences for integration of viral gene vectors, whether persistence of integrated vectors is coupled to specific nuclear locations and whether these locations are changed by vector transcription. We will determine transcriptional activity and then visualize mapped integration sites together with other structures in the nucleus by fluorescence in situ hybridization and three-dimensional microscopy. Quantitative image analysis will be applied to measure positions of integration sites relative to the nuclear space, their harboring chromosome territories, and specific nuclear structures such as heterochromatin, splicing speckles and early and late replicating chromatin. By comparing integration loci of HIV and Foamy virus systems in different cell types, we will analyze the virus and cell type specificity of targeting of nuclear regions and the topological changes caused by viral vectors. In addition we will analyze the effects of transcriptional activation by viral transactivators on the nuclear architecture.

• DFG-Verfahren: Schwerpunktprogramme
• Teilprojekt zu: SPP 1230: Mechanisms of gene vector entry and persistence
• Beteiligte Personen: Privatdozent Dr. Jochen Bodem; Professorin Dr. Ruth Brack-Werner