In:
Cancer Research, American Association for Cancer Research (AACR), Vol. 76, No. 14_Supplement ( 2016-07-15), p. 4419-4419
Abstract:
The ability to generate interferon lambda 4 (IFNL4), the most recently discovered member of the type-III interferon family, has been associated with impaired clearance of hepatitis C virus (HCV). IFNL4 signals through an IFNL receptor complex that serves all type-III interferons and consists of receptors IFNLR1 and IL10R2. Signaling through IFNL receptor complex leads to activation of the JAK-STAT pathway and transcriptional activation of interferon-stimulated genes (ISG). Intracellular expression of IFNL4 also causes reduced proliferation and increased cell death in human hepatic cells but it remains unclear if this is a result of JAK-STAT signaling through IFNL receptor complex or through some other alternative mechanisms. To address this question, we used CRISPR-CAS9 system to knock-out IFNLR1 in a hepatoma HepG2 cells. We designed and tested six guide RNAs (gRNAs) for IFNLR1 exons. CAS9 protein and gRNAs that most efficiently induced mutations in IFNLR1 were introduced into HepG2 cells to generate stable cell lines. No obvious defects were observed in IFNLR1-KO-HepG2 cells, indicating that loss of IFNLR1 had no deleterious effects on cell viability. We selected three stable clones which had sequencing-confirmed inactivating IFNLR1 mutations and Western blot-confirmed lack of IFNLR1 protein expression. Compared to the parental HepG2 cells the IFNLR1-KO-HepG2 cells were deficient in the ability to induce the luciferase-tagged interferon-stimulated response element (ISRE-Luc) reporter after treatment with recombinant IFNL4 and IFNL3 proteins, which signal through the same receptor complex. At the same time, response to IFNa treatment was not affected because IFNa signals through its own receptor complex. These results indicate that, as other type-III interferons, IFNL4 induces ISRE signaling through IFNL receptor complex. INFLR1-KO-HepG2 cells provide a highly efficient model for studies of factors involved in signaling of type-III interferons, including IFNL4. Next, we will investigate alternative pathways involved in anti-proliferative and cell death-related phenotypes induced by IFNL4 in HepG2 cell. We will perform RNA-seq on parental and IFNLR1-KO-HepG2 cells treated with or without IFNL4, IFNL3 and IFNa. These results will contribute to a better understanding of IFNL4 function, which can be relevant for viral infections and cancer. Citation Format: Fang Wang, Olusegun O. Onabajo, Nina Rao, Ludmila Prokunina-Olsson. Exploration of alternative pathways mediated by IFNL4 and related to cell proliferation and death in a hepatoma cell line. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4419.
Type of Medium:
Online Resource
ISSN:
0008-5472
,
1538-7445
DOI:
10.1158/1538-7445.AM2016-4419
Language:
English
Publisher:
American Association for Cancer Research (AACR)
Publication Date:
2016
detail.hit.zdb_id:
2036785-5
detail.hit.zdb_id:
1432-1
detail.hit.zdb_id:
410466-3
