A Genome-wide CRISPR Screen Identifies ZCCHC14 as a Host Factor Required for Hepatitis B Surface Antigen Production

A hallmark of chronic hepatitis B (CHB) virus infection is the presence of high circulating levels of non-infectious small lipid HBV surface antigen (HBsAg) vesicles. Although rare, sustained HBsAg loss is the idealized endpoint of any CHB therapy. A small molecule, RG7834, has been previously reported to inhibit HBsAg expression by targeting terminal nucleotidyltransferase proteins 4A and 4B (TENT4A and TENT4B). In this study, we describe a genome-wide CRISPR screen to identify other potential host factors required for HBsAg expression and to gain further insights into the mechanism of RG7834. We report more than 60 genes involved in regulating HBsAg and identify additional factors involved in RG7834 activity, including a zinc finger CCHC-type containing 14 (ZCCHC14) protein. We show that ZCCHC14, together with TENT4A/B, stabilizes HBsAg expression through HBV RNA tailing, providing a potential new therapeutic target to achieve functional cure in CHB patients. Hyrina et al. employ a non-biased functional CRISPR screening approach to identify host factors regulating HBsAg expression as well as those targeted by RG7834, a HBsAg inhibitor. The screen highlighted over 60 genes and identified a mechanism by which ZCCHC14, together with TENT4A/B, stabilizes HBsAg expression through HBV RNA tailing

Genome-wide CRISPR screen identifies protein pathways modulating tau protein levels in neurons

Aggregates of hyperphosphorylated tau protein are a pathological hallmark of more than 20 distinct neurodegenerative diseases, including Alzheimer's disease, progressive supranuclear palsy, and frontotemporal dementia. While the exact mechanism of tau aggregation is unknown, the accumulation of aggregates correlates with disease progression. Here we report a genome-wide CRISPR screen to identify modulators of endogenous tau protein for the first time. Primary screens performed in SH-SY5Y cells, identified positive and negative regulators of tau protein levels. Hit validation of the top 43 candidate genes was performed using Ngn2-induced human cortical excitatory neurons. Using this approach, genes and pathways involved in modulation of endogenous tau levels were identified, including chromatin modifying enzymes, neddylation and ubiquitin pathway members, and components of the mTOR pathway. TSC1, a critical component of the mTOR pathway, was further validated in vivo, demonstrating the relevance of this screening strategy. These findings may have implications for treating neurodegenerative diseases in the future

Macroautophagy is dispensable for growth of KRAS mutant tumors and chloroquine efficacy

Macroautophagy is a key stress response pathway that can suppress or promote tumorigenesis depending on the cellular context. Notably, KRAS-driven tumors have been reported to rely on macroautophagy for growth and survival suggesting a potential therapeutic approach to use autophagy inhibitors based on genetic stratification. In this study, we evaluated if KRAS mutation status can predict the efficacy to inhibition of macroautophagy. By profiling 47 cell lines with pharmacological and genetic loss of function tools, we were unable to confirm that KRAS-driven tumor lines require macroautophagy for growth. Deletion of ATG7 by genome editing completely blocked macroautophagy in several tumor lines with oncogenic mutations in KRAS but did not inhibit cell proliferation in vitro or tumorigenesis in vivo. Furthermore, ATG7 knock out did not sensitize cells to irradiation or several anti-cancer agents tested. Interestingly, ATG7-deficient and -proficient cells were equally sensitive to the anti-proliferative effect of chloroquine, a lysosomotropic agent often used as a pharmacological tool to evaluate the response to macroautophagy inhibition. Moreover, both cell types manifested synergistic growth inhibition when treated with chloroquine plus the tyrosine kinase inhibitors erlotinib or sunitinib, suggesting that the antiproliferative effects of chloroquine are independent of its suppressive actions on autophagy

Genome-wide CRISPR screen identifies protein pathways modulating tau protein levels in neurons

Using an unbiased genome-wide CRISPR screen approach, Sanchez et al. identified modulators of endogenous tau protein. This study suggests that chromatin modifiers, neddylation, ubiquitination, and the mTOR pathways regulate overall levels of tau protein in neurons, which could help in future identification of therapeutics for neurodegenerative diseases