Helmholtz Zentrum München Deutsches Forschungszentrum für Gesundheit und Umwelt

Ribonucleoprotein-based CRISPR approaches as novel tools for gene regulation​

Tobias was initially trained as a mechatronics engineer. Motivated by his hobbies, however, he became interested in the complex functions of "living" machines. He started his bachelor studies in chemistry and biochemistry at the LMU Munich. Fascinated by the scientific field of epigenetics, he completed his studies with a bachelor thesis in the laboratory of Dr. Stephan Hamperl at Helmholtz Zentrum München. Impressed by the complexity of the mechanisms of biological matter, a master's degree in biochemistry followed. For his master's thesis he joined the laboratory of Prof. Dr. Stefan Stricker at Helmholtz Zentrum München to investigate the influence of endogenous transcriptional regulation on cell identity.

Driven by great fascination for the research field of epigenetics, he stays in the lab of Prof. Dr. Stefan Stricker for his EpiCrossBorders PhD project. For decades, the gold standard for expressing a gene of interest was over-expression of cDNA or the open-reading-frame (ORF). Using this approach, it is difficult to control the level of exogenous gene expression or investigate the endogenous regulation of the gene. Thanks to the discovery and the experimental exploitation of the CRISPR system, it became possible to flexibly induce endogenous gene expression via CRISPR-activation (CRISPRa). However, the delivery of the CRIPRa-system still involves large DNA plasmids and/or the use of viral particles, which might lead to cytotoxicity and often requires stable genomic integration. To overcome these hurdles, we developed a RNP-based CRISPR activation system. The nuclease deficient Cas9 (dCas9) is used as recombinant protein linked to a transcriptional activator. This allows the pre-assembly of active RNP-complexes. Our approach has a high potential for the activation of multiple transcription factors either as single targets or through multiplexing. We aim to make use of the flexibility and the transient nature of the system to provide a potent gene activation system to study and manipulate gene regulation and cell identity.​