CRISPR/Cas9-Mediated Whole Genomic Wide Knockout Screening Identifies Specific Genes Associated With PM2.5-Induced Mineral Absorption in Liver Toxicity.

Bin Yi,Jieqiong Tan,Mengyao Wang,Zhijun Huang,Jinfu Peng

doi:10.3389/fbioe.2021.669434

Abstract

PM2.5, also known as fine particles, refers to particulate matter with a dynamic diameter of ≦2.5 μm in air pollutants, that carries metals (Zn, Co, Cd) which can pass through the alveolar epithelium and enter the circulatory system and tissues. PM2.5 can cause serious health problems, such as non-alcoholic fatty liver and hepatocellular carcinoma, although the underlying mechanisms of its toxic effect are poorly understood. Here, we exposed L02 cells to PM2.5 and performed a pooled genome−wide clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) to assess loss of function and identify new potential PM2.5targets. Enrichr and KEGG pathway analyses were performed to identify candidate genes associated with PM2.5 toxicity. Results revealed that four key genes, namely ATPase Na+/K+ transporting subunit alpha 2 (ATP1A2), metallothionein 1M (MT1M), solute carrier family 6 members 19 (SLC6A19) and transient receptor potential cation channel subfamily V member 6 (TRPV6) were associated with PM2.5 toxicity, mainly in regulating the mineral absorption pathway. Downregulating these genes increased cell viability and attenuated apoptosis in cells exposed to PM2.5. Conversely, overexpressing TRPV6 exacerbated cell apoptosis caused by PM2.5, while a reactive oxygen species (ROS) inhibitor N-acetyl-l-cysteine (NAC) alleviated PM2.5-induced apoptosis. In conclusion, ATP1A2, MT1M, SLC6A19 and TRPV6 may be contributing to absorption of metals in PM2.5 thereby inducing apoptosis mediated by ROS. Therefore, they hold potential as therapeutic targets for PM2.5-related diseases.

Highlights

Particulate Matter (PM) refers to a suspended mixture of solid and liquid particles in the air (Oh et al, 2021), whereas inhalable fine particles are described by PM2.5, usually 2.5 microns or less in diameter (Xing et al, 2016)
We identified some interesting gene defects that were potentially associated with protection from PM2.5 toxicity (Supplementary Table 2)
Overexpressing TRPV6 positively promoted reactive oxygen species (ROS) production (Figure 5B). These findings suggest that TPRV6 promotes the toxic effects of PM2.5, thereby aggravating ROS production in the cell, leading to apoptosis

Summary

Introduction

Particulate Matter (PM) refers to a suspended mixture of solid and liquid particles in the air (Oh et al, 2021), whereas inhalable fine particles are described by PM2.5, usually 2.5 microns or less in diameter (Xing et al, 2016) These potentially harbors various toxic substances, including heavy metals or minerals such as copper (Cu), zinc (Zn), calcium (Ca), potassium (K), and cadmium (Cd). Previous studies have described the mechanism of PM2.5induced liver disease, with ROS and lysosome implicated in PM2.5-induced cell apoptosis (Dornhof et al, 2017; Jing Piao et al, 2018; Zhu et al, 2018). The underlying mechanism of PM2.5 -mediated hepatotoxicity remains unclear, necessitating further exploration

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Discussion

Conclusion