A Transient Expression System with Stably Integrated CRISPR-dCas9 Fusions for Regulation of Genes Involved in Immunoglobulin G Glycosylation

Mijakovac, Anika; Miškec, Karlo; Krištić, Jasminka; Vičić Bočkor, Vedrana; Tadić, Vanja; Bošković, Maria; Lauc, Gordan; Zoldoš, Vlatka; Vojta, Aleksandar (2022) A Transient Expression System with Stably Integrated CRISPR-dCas9 Fusions for Regulation of Genes Involved in Immunoglobulin G Glycosylation. The CRISPR Journal, 5 (2). pp. 237-253. ISSN 2573-1599

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Abstract

Alternative glycosylation of immunoglobulin G (IgG) is functionally important in multiple human physiological and pathological states. Our understanding of molecular mechanisms that regulate IgG glycosylation is vague because of the complexity of this process, which involves hundreds of genes. Several genome-wide association (GWA) studies have revealed a network of genes associated with IgG glycosylation that are pleiotropic for a number of diseases. Here, we report a design of a versatile system for IgG production and gene manipulations that can be used for in vitro functional follow-up of GWA hits or any gene of interest. The system is based on CRISPR-dCas9, extended by a piggyBac integrase compatible vector, and drives IgG production in HEK-293F cells. We validated our systems that stably express VPR-dCas9 and KRAB-dCas9 by manipulation of four glyco-genes with a known role in IgG glycosylation, and then functionally validated three GWAS hits for IgG glycosylation with an as-yet-unknown role in this process.

Item Type:	Article
Uncontrolled Keywords:	CRISPR ; regulation ; epigenetics
Subjects:	NATURAL SCIENCES > Biology
Divisions:	Division of Molecular Biology
Projects:	Project title Project leader Project code Project type Centar kompetencija u molekularnoj dijagnostici-CEKOM Damir Knjaz; Gordan Lauc; Tomislav Krističević; Branka Matković; Saša Janković; Tomislav Rupčić; Ivan Bonić; Josip Zekić; Marin Vidović; Mateja Očić; Nina Briški; Robert Zekić; Vedran Dukarić; Oliver Vugrek; Ozren Polašek KK.01.2.2.03.0006 EK Znanstveni centar izvrsnosti za personaliziranu brigu o zdravlju-ZCIPersonHealth Aleksandar Secenji KK.01.1.1.01.0010 EK Genomsko inženjerstvo i genska regulacija u staničnim linijama i modelnim organizacijama tehnologija CRISPR / Cas9 CasMouse-CRISPR/Cas9-CasMouse Vlatka Zoldoš; Srećko Gajović KK.01.1.1.04.0085 EK
Depositing User:	Martina Žugaj
Date Deposited:	28 Apr 2026 13:10
URI:	https://fulir.irb.hr:/id/eprint/11814
DOI:	10.1089/crispr.2021.0089

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