SEC14L2, a lipid-binding protein, regulate HCV replication in culture with inter- and intra- genotype variations
2018
Authors: Costa R, Todt D, Zapatero-Belinchón F, Schenk C, Anastasiou OE, Walker A, Hertel B, Timmer L, Bojkova D, Ruckert M, Sarrazin C, Timm J, Lohmann V, Manns MP, Steinman E, Hahn TV, Ciesek S
CellNetworks People: Lohmann Volker
Journal: J Hepatol. 2018 Nov 22. pii: S0168-8278(18)32544-3. doi: 10.1016/j.jhep.2018.11.012

BACKGROUND & AIMS:
Since the discovery of HCV, efficient viral replication of clinical isolates in cell culture was a challenge. This status quo changed with SEC14L2 mediated viral replication of clinical isolates in cell culture. Given the role of SEC14L2 in HCV replication, we aimed to study a large number of HCV positive sera carrying genotypes 1-4, to identify viral factors associated with efficient replication in cell culture. Additionally, we investigated whether 13 single nucleotide polymorphisms (SNPs) of SEC14L2 have an impact on RNA replication of natural occurring HCV isolates.

METHODS:
We generated Huh-7.5 cell lines overexpressing SEC14L2 or 13 coding SNPs and tested 73 different HCV positive sera for in vitro replication. Furthermore, we genotyped a cohort of 262 patients with chronic HCV for the common SNP (rs757660) and investigated its effect on the clinical phenotype.

RESULTS:
HCV isolates from genotype 1, 2, 3 and 4 replicate in Huh-7.5 cells overexpressing SEC14L2. Interestingly, only subgenomic replicons from genotypes 1 and 3 showed enhanced replication whereas genotypes 2 and 4 remained unaffected. Furthermore, replication was independent from viral load. Importantly, all tested SNPs supported HCV RNA replication in vitro, while one SNP was associated with decreased SEC1L2 expression and viral RNA. All SNPs exhibited comparable cellular cholesterol and vitamin E abundance in naïve Huh-7.5 cells.

CONCLUSIONS:
This large screen of natural HCV isolates of 4 genotypes underscores the relevance of SEC14L2 as an in vitro HCV host factor. Additionally, SEC14L2 variants appear to recapitulate the wild type enhancement of HCV replication. Variant rs191341134 showed a decreased effect due to lowered stability, whereas variant rs757660, a high prevalence mutant, showed similar phenotype to the wild type.

LAY SUMMARY:
Until the year 2015, consistent replication of patient derived isolates of hepatitis C virus (HCV) in an in vitro model remained a limitation in HCV research. In 2015 a group of authors identified a protein named SEC14L2 that enabled the replication of isolates in a human hepatoma cell line named Huh-7.5. We implemented this model in our laboratory and performed a large screen encompassing 73 isolates of 4 different HCV genotypes. Additionally, we replaced the natural SEC14L2 with 13 different mutants to test if the protein variation could significantly alter its HCV replication enhancing functions. We showed that different genotypes of HCV react differently to the presence of this protein and the variants of the protein mimic the behaviour of the wild type.