Goran Josipović | Senior Researcher
Genos
Dr. Goran Josipović is a Senior Researcher in the Epigenetic Laboratory at Genos Ltd. in Zagreb, Croatia. He obtained his PhD in Biology from the Faculty of Science, University of Zagreb, where his research focused on epigenome editing and the design of a modular CRISPR/dCas9 platform based on orthogonal Cas9 proteins. His current research uses CRISPR/dCas9-based epigenome editing to investigate how the interplay between DNA methylation and post-translational histone modifications regulates gene expression and influences the establishment and maintenance of DNA methylation patterns. His research also addresses the molecular mechanisms regulating IgA1 glycosylation. He continues to develop the modular Epi-Toolbox by incorporating additional dCas9 orthologues, epigenetic effector domains and new vector backbones, thereby expanding its capacity for targeted, multiplexed and combinatorial manipulation of epigenetic marks.
Appearances:
Festival of Biologics Day 2 @ 17:30
CRISPR-Based Epigenome Editing: From Targeted DNA Methylation to the Development of a Modular Epi-Toolbox
Can gene activity be precisely and durably controlled without changing the underlying DNA sequence? CRISPR-based epigenome editing offers this possibility by targeting regulatory regions and modifying the epigenetic signals that control transcription. We were the first to develop a CRISPR/dCas9–DNMT3A system for targeted DNA methylation. Recruitment of the DNMT3A catalytic domain induced site-specific CpG methylation, while the use of multiple guide RNAs enabled the modification of broader promoter regions. Targeted methylation of the BACH2 and IL6ST promoters was maintained through cell divisions and reduced their expression, demonstrating that the precise introduction of an DNA methylation can influence transcriptional activity. Building on this work, we developed a modular and extensible Epi-Toolbox based on Golden Gate assembly. The Epi-Toolbox combines interchangeable promoters, orthogonal dCas9 proteins, epigenetic effector domains, selection markers and multiplexed guide RNA modules. The parallel use of dCas9 orthologues from Streptococcus pyogenes and Staphylococcus aureus enabled distinct epigenetic effectors to be recruited either to separate genomic regions for simultaneous epigenetic editing or to the same regulatory region for synergistic modulation of gene expression. Targeted methylation and demethylation using DNMT3A and TET1 produced gene-specific transcriptional responses. Simultaneous editing of HNF1A and MGAT3 also altered the cellular N-glycan profile, directly linking engineered epigenetic states to a complex molecular phenotype. Furthermore, combined recruitment of TET1 and the transcriptional activator VPR to the same regulatory region produced strong synergistic gene activation that persisted for up to 30 days. Our current work focuses on expanding the Epi-Toolbox with additional dCas orthologues, backbones and epigenetic effector domains for the targeted modification of histone marks. This adaptable platform enables precise, combinatorial and potentially durable control of gene expression, offering new opportunities for functional epigenetics, disease modelling and the future development of epigenetic therapies.
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