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izvor podataka: crosbi

Antagonistic and synergistic epigenetic modulation using orthologous CRISPR/dCas9-based modular system (CROSBI ID 268369)

Prilog u časopisu | izvorni znanstveni rad | međunarodna recenzija

Josipović, Goran ; Tadić, Vanja ; Klasić, Marija ; Zanki, Vladimir ; Bečeheli, Ivona ; Chung, Felicia ; Ghantous, Akram ; Keser, Toma ; Madunić, Josip ; Bošković, Maria et al. Antagonistic and synergistic epigenetic modulation using orthologous CRISPR/dCas9-based modular system // Nucleic acids research, 47 (2019), 18; 9637-9657. doi: 10.1093/nar/gkz709

Podaci o odgovornosti

Josipović, Goran ; Tadić, Vanja ; Klasić, Marija ; Zanki, Vladimir ; Bečeheli, Ivona ; Chung, Felicia ; Ghantous, Akram ; Keser, Toma ; Madunić, Josip ; Bošković, Maria ; Lauc, Gordan ; Herceg, Zdenko ; Vojta, Aleksandar ; Zoldoš, Vlatka

engleski

Antagonistic and synergistic epigenetic modulation using orthologous CRISPR/dCas9-based modular system

Establishing causal relationship between epigenetic marks and gene transcription requires molecular tools, which can precisely modify specific genomic regions. Here, we present a modular and extensible CRISPR/dCas9- based toolbox for epigenetic editing and direct gene regulation. It features a system for expression of orthogonal dCas9 proteins fused to various effector domains and includes a multi-gRNA system for simultaneous targeting dCas9 orthologs to up to six loci. The C- and N- terminal dCas9 fusions with DNMT3A and TET1 catalytic domains were thoroughly characterized. We demonstrated simultaneous use of the DNMT3A- dSpCas9 and TET1-dSaCas9 fusions within the same cells and showed that imposed cytosine hyper- and hypo-methylation altered level of gene transcription if targeted CpG sites were functionally relevant. Dual epigenetic manipulation of the HNF1A and MGAT3 genes, involved in protein N-glycosylation, resulted in change of the glycan phenotype in BG1 cells. Furthermore, simultaneous targeting of the TET1- dSaCas9 and VPR-dSpCas9 fusions to the HNF1A regulatory region revealed strong and persistent synergistic effect on gene transcription, up to 30 days following cell transfection, suggesting involvement of epigenetic mechanisms in maintenance of the reactivated state. Also, modulation of dCas9 expression effectively reduced off-target effects while maintaining the desired effects on target regions.

Epigenetics ; Methylation ; CRISPR/dCas9 ; HNF1A ; MGAT3 ; N-glycosylation

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Podaci o izdanju

47 (18)

2019.

9637-9657

objavljeno

0305-1048

1362-4962

10.1093/nar/gkz709

Povezanost rada

Biologija, Biotehnologija u biomedicini (prirodno područje, biomedicina i zdravstvo, biotehničko područje), Farmacija

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