Kendomycin Cytotoxicity against Bacterial, Fungal, and Mammalian Cells Is Due to Cation Chelation

Kendomycin is a small-molecule natural product that has gained significant attention due to reported cytotoxicity against pathogenic bacteria and fungi as well as a number of cancer cell lines. Despite significant biomedical interest and attempts to reveal its mechanism of action, the cellular target of kendomycin remains disputed. Herein it is shown that kendomycin induces cellular responses indicative of cation stress comparable to the effects of established iron chelators. Furthermore, addition of excess iron and copper attenuated kendomycin cytotoxicity in bacteria, yeast, and mammalian cells. Finally, NMR analysis demonstrated a direct interaction with cations, corroborating a close link between the observed kendomycin polypharmacology across different species and modulation of iron and/or copper levels.Peer reviewe

Decatransin, a novel natural product inhibiting protein translocation at the Sec61/SecY translocon

A new cyclic decadepsipeptide was isolated from Chaetosphaeria tulasneorum with potent bioactivity on mammalian and yeast cells. Chemogenomic profiling in S. cerevisiae indicated that the Sec61 translocon, the machinery for protein translocation and membrane insertion at the endoplasmic reticulum, is the target. The profiles were similar to those of cyclic heptadepsipeptides of a distinct chemotype (HUN-7293/cotransin) that had previously been shown to inhibit cotranslational translocation at the mammalian Sec61 translocon. Unbiased, genome-wide mutagenesis followed by full-genome sequencing in both fungal and mammalian cells identified dominant mutations in Sec61p/Sec61α1 to confer resistance. Most, but not all, of these mutations affected inhibition by both chemotypes, despite an absence of structural similarity. Biochemical analysis confirmed inhibition of protein translocation into the endoplasmic reticulum of both co- and posttranslationally translocated substrates by both chemotypes, demonstrating a mechanism independent of a translating ribosome. Most interestingly, both chemotypes were found to also inhibit SecYEG, the bacterial Sec61 homolog. We suggest "decatransin" as the name for this novel decadepsipeptide translocation inhibitor

A first update on mapping the human genetic architecture of COVID-19

peer reviewe

CRISPR-Cas9 screen identifies mechanisms of BET bromodomain inhibitor sensitivity including manganese

BET Bromodomain Inhibitors hold promise as therapeutic agents in inflammation and cancer but clinical studies show adverse side-effects at high, sustained dose. Clinical success requires further mechanistic understanding of inhibition of BET bromodomains and biomarkers to optimize efficacious dosing. To uncover the mechanisms of sensitivity and resistance to BETi, we employed a whole-genome CRISPR-Cas9 proliferation screen using colorectal cancer cells. We identify the mTOR signaling pathway as a key determinant of BETi sensitivity and that two Ca/Mn transporters mediate resistance. This later finding led to the discovery that extracellular manganese regulates sensitivity to BETi and that exposure of cells to BETi dose dependently increases intracellular manganese concentration. Our results describe new molecular pathways mediating BETi action and suggest several potential avenues for biomarker discovery

Species-specific disruption of STING-dependent antiviral cellular defenses by the Zika virus NS2B3 protease

The limited host tropism of numerous viruses causing disease in humans remains incompletely understood. One example is Zika virus (ZIKV), an RNA virus that has reemerged in recent years. Here, we demonstrate that ZIKV efficiently infects fibroblasts from humans, great apes, New and Old World monkeys, but not rodents. ZIKV infection in human—but not murine—cells impairs responses to agonists of the cGMP-AMP synthase/stimulator of IFN genes (cGAS/STING) signaling pathway, suggesting that viral mechanisms to evade antiviral defenses are less effective in rodent cells. Indeed, human, but not mouse, STING is subject to cleavage by proteases encoded by ZIKV, dengue virus, West Nile virus, and Japanese encephalitis virus, but not that of yellow fever virus. The protease cleavage site, located between positions 78/79 of human STING, is only partially conserved in nonhuman primates and rodents, rendering these orthologs resistant to degradation. Genetic disruption of STING increases the susceptibility of mouse—but not human—cells to ZIKV. Accordingly, expression of only mouse, not human, STING in murine STING knockout cells rescues the ZIKV suppression phenotype. STING-deficient mice, however, did not exhibit increased susceptibility, suggesting that other redundant antiviral pathways control ZIKV infection in vivo. Collectively, our data demonstrate that numerous RNA viruses evade cGAS/STINGdependent signaling and affirm the importance of this pathway in shaping the host range of ZIKV. Furthermore, our results explain— at least in part—the decreased permissivity of rodent cells to ZIKV, which could aid in the development of mice model with inheritable susceptibility to ZIKV and other flaviviruses

Direct Interaction of Chivosazole F with Actin Elicits Cell Responses Similar to Latrunculin A but Distinct from Chondramide

The microbial metabolite Chivosazole F has been described to affect the cytoskeleton and to inhibit actin polymerization <i>in vitro</i>. Applying orthogonal genomic and proteomics approaches, we now show for the first time that Chivosazole F exerts its effect by directly interacting with actin and demonstrate the cellular impact of Chivosazole F in an unbiased, genome-wide context in yeast and in mammalian cells. Furthermore, mutation-based resistance mapping identifies two SNPs located in the putative Chivosazole F binding site of actin. Comparing chemogenomic profiles and responses to the Chivosazole F-resistant SNPs shows a partially conserved mechanism of action for Chivosazole F and Latrunculin A, but clear divergence from Chondramide. In addition, C14orf80 is an evolutionarily highly conserved ORF, lacking any functional annotation. As editing of C14orf80 leads to Chivosazole F hyper-resistance, we propose a function for this gene product in counteracting perturbation of actin filaments

Direct Interaction of Chivosazole F with Actin Elicits Cell Responses Similar to Latrunculin A but Distinct from Chondramide

The microbial metabolite Chivosazole F has been described to affect the cytoskeleton and to inhibit actin polymerization <i>in vitro</i>. Applying orthogonal genomic and proteomics approaches, we now show for the first time that Chivosazole F exerts its effect by directly interacting with actin and demonstrate the cellular impact of Chivosazole F in an unbiased, genome-wide context in yeast and in mammalian cells. Furthermore, mutation-based resistance mapping identifies two SNPs located in the putative Chivosazole F binding site of actin. Comparing chemogenomic profiles and responses to the Chivosazole F-resistant SNPs shows a partially conserved mechanism of action for Chivosazole F and Latrunculin A, but clear divergence from Chondramide. In addition, C14orf80 is an evolutionarily highly conserved ORF, lacking any functional annotation. As editing of C14orf80 leads to Chivosazole F hyper-resistance, we propose a function for this gene product in counteracting perturbation of actin filaments

Direct Interaction of Chivosazole F with Actin Elicits Cell Responses Similar to Latrunculin A but Distinct from Chondramide

The microbial metabolite Chivosazole F has been described to affect the cytoskeleton and to inhibit actin polymerization <i>in vitro</i>. Applying orthogonal genomic and proteomics approaches, we now show for the first time that Chivosazole F exerts its effect by directly interacting with actin and demonstrate the cellular impact of Chivosazole F in an unbiased, genome-wide context in yeast and in mammalian cells. Furthermore, mutation-based resistance mapping identifies two SNPs located in the putative Chivosazole F binding site of actin. Comparing chemogenomic profiles and responses to the Chivosazole F-resistant SNPs shows a partially conserved mechanism of action for Chivosazole F and Latrunculin A, but clear divergence from Chondramide. In addition, C14orf80 is an evolutionarily highly conserved ORF, lacking any functional annotation. As editing of C14orf80 leads to Chivosazole F hyper-resistance, we propose a function for this gene product in counteracting perturbation of actin filaments

Direct Interaction of Chivosazole F with Actin Elicits Cell Responses Similar to Latrunculin A but Distinct from Chondramide

The microbial metabolite Chivosazole F has been described to affect the cytoskeleton and to inhibit actin polymerization <i>in vitro</i>. Applying orthogonal genomic and proteomics approaches, we now show for the first time that Chivosazole F exerts its effect by directly interacting with actin and demonstrate the cellular impact of Chivosazole F in an unbiased, genome-wide context in yeast and in mammalian cells. Furthermore, mutation-based resistance mapping identifies two SNPs located in the putative Chivosazole F binding site of actin. Comparing chemogenomic profiles and responses to the Chivosazole F-resistant SNPs shows a partially conserved mechanism of action for Chivosazole F and Latrunculin A, but clear divergence from Chondramide. In addition, C14orf80 is an evolutionarily highly conserved ORF, lacking any functional annotation. As editing of C14orf80 leads to Chivosazole F hyper-resistance, we propose a function for this gene product in counteracting perturbation of actin filaments

Decatransin, a novel natural product inhibiting protein translocation at the Sec61/SecY translocon

A new cyclic decadepsipeptide was isolated from Chaetosphaeria tulasneorum with potent bioactivity on mammalian and yeast cells. Chemogenomic profiling in S. cerevisiae indicated that the Sec61 translocon, the machinery for protein translocation and membrane insertion at the endoplasmic reticulum, is the target. The profiles were similar to those of cyclic heptadepsipeptides of a distinct chemotype (HUN-7293/cotransin) that had previously been shown to inhibit cotranslational translocation at the mammalian Sec61 translocon. Unbiased, genome-wide mutagenesis followed by full-genome sequencing in both fungal and mammalian cells identified dominant mutations in Sec61p/Sec61α1 to confer resistance. Most, but not all, of these mutations affected inhibition by both chemotypes, despite an absence of structural similarity. Biochemical analysis confirmed inhibition of protein translocation into the endoplasmic reticulum of both co- and posttranslationally translocated substrates by both chemotypes, demonstrating a mechanism independent of a translating ribosome. Most interestingly, both chemotypes were found to also inhibit SecYEG, the bacterial Sec61 homolog. We suggest "decatransin" as the name for this novel decadepsipeptide translocation inhibitor