CRISPR screening identifies protein methylation and ubiquitination modifications that modulate aflatoxin B1 cytotoxicity.

Abstract

Aflatoxin B₁ (AFB₁) is one of the most potent mycotoxins affecting human health and animal production. To deeply understand the host-toxin interaction, we performed CRISPR screening and identified cystathionine β-synthase (CBS) as a critical host gene affecting AFB₁ cytotoxicity. Mechanistic studies revealed that CBS affects AFB₁-induced cell death by regulating the abundance of protein post-translational modifications (PTMs) in host cells. First, AFB₁ disrupted the transfer of S-adenosylmethionine (SAM) from the cytoplasm to the mitochondria, thereby reducing the intra-mitochondrial protein methylation level. Deficient intra-mitochondrial protein methylation impaired mitochondrial function and caused cell death. CBS knockout (KO) can enhance SAM generation and mobilization to restore intra-mitochondrial SAM levels by rescuing the perturbed methionine cycle after AFB₁ exposure, thereby alleviating AFB₁-induced cell death. Second, AFB₁ decreased global protein ubiquitination levels by affecting gene expression of ubiquitin-modified enzymes. CBS-KO and pharmaceutical treatment correcting gene expression of ubiquitin-modified enzymes can rescue AFB₁-induced cell death. We also investigated two PTM-regulating small molecules, SAM and PR-619, which can increase cell viability in AFB₁-exposed cells.