Tetracenomycin X inhibits translation by binding within the ribosomal exit tunnel

Osterman, Ilya А.; Wieland, Maximiliane; Maviza, Tinashe P.; Lashkevich, Kseniya A; Lukianov, Dmitrii A.; Komarova, Ekaterina S.; Zakalyukina, Yu. V.; Buschauer, Robert; Shiriaev, Dmitrii I.; Leyn, Semen A.; Zlamal, Jaime E.; Biryukov, Mikhail V.; Skvortsov, Dmitry A.; Tashlitsky, Vadim N.; Polshakov, Vladimir I.; Cheng, Jingdong; Polikanov, Y.S.; Bøgdanov, A.; Osterman, Andrei L.; Dmitriev, Sergey E.; Beckmann, Roland; Dontsova, Olga А.; Wilson, Daniel N.; Сергиев, Петр В.

doi:10.1038/s41589-020-0578-x

Cited by 60 publications

(83 citation statements)

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“…Thus, we hypothesized that RPL3 methylation may impact protein synthesis. To understand the potential role of methylation in RPL3, we reanalyzed published cryo-electron microscopy data (Osterman et al, 2020) and assessed the τ- N -methyl moiety on His245 of RPL3 (Figure 3A), which indicated that τ- N -methylated RPL3 is a stoichiometric component of the ribosome. A similar density of methylation on the histidine of RPL3 has also been found in rabbit ribosomes (Bhatt et al, 2021).…”

Section: Resultsmentioning

confidence: 99%

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METTL18-mediated histidine methylation on RPL3 modulates translation elongation for proteostasis maintenance

Matsuura-Suzuki

Shimazu

Takahashi

et al. 2021

Preprint

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Protein methylation occurs predominantly on lysine and arginine residues, but histidine also serves as a substrate for the modification. However, a limited number of enzymes responsible for this modification have been reported. Moreover, the biological role of histidine methylation has remained poorly understood. Here, we report that human METTL18 is a histidine methyltransferase for the ribosomal protein RPL3 and that the modification specifically slows ribosome traverse on tyrosine codons, allowing the proper folding of synthesized proteins. By performing an in vitro methylation assay with a methyl donor analog and quantitative mass spectrometry, we found that His245 of RPL3 is methylated at the τ-N position by METTL18. Structural comparison of the modified and unmodified ribosomes showed stoichiometric modification and suggested a role in translation tuning. Indeed, genome-wide ribosome profiling revealed suppressed ribosomal translocation at tyrosine codons by RPL3 methylation. Because the slower elongation provides enough time for nascent protein folding, RPL3 methylation protects cells from the cellular aggregation of Tyr-rich proteins. Our results reveal histidine methylation as an example of a “ribosome code” that ensures proteome integrity in cells.

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Section: Resultsmentioning

confidence: 99%

“…(A) Stick models of 244 GHR 246 of RPL3 and G1595 of the 28S rRNA of the human ribosome are shown with the cryo-EM density map around His245. The τ- N -methyl group was manually added to the original model (PDB ID: 6Y6X) (Osterman et al, 2020) based on the cryo-EM density map.…”

Section: Resultsmentioning

confidence: 99%

METTL18-mediated histidine methylation on RPL3 modulates translation elongation for proteostasis maintenance

Matsuura-Suzuki

Shimazu

Takahashi

et al. 2021

Preprint

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“…It is remarkable that in spite of these differences, PF846 and TEL achieve context-specific inhibition of translation by binding to overlapping sites in the NPET. Interestingly, it has been recently shown that the compound tetracenomycin X binds to bacterial and eukaryotic ribosomes in a cavity of the NPET located on the wall opposite to the macrolide binding site and appears to act, at least during bacterial translation, in a sequence-specific manner 64 . Therefore, the PTC-proximal NPET segment emerges as the best target for the inhibitors, whose action depends on the sequence context of the growing polypeptide.…”

Section: Discussionmentioning

confidence: 99%

“…b-e Representative gels of two independent experiments. and eukaryotic ribosomes in a cavity of the NPET located on the wall opposite to the macrolide binding site and appears to act, at least during bacterial translation, in a sequence-specific manner 64 . Therefore, the PTC-proximal NPET segment emerges as the best target for the inhibitors, whose action depends on the sequence context of the growing polypeptide.…”

Section: Discussionmentioning

confidence: 99%

Context-specific action of macrolide antibiotics on the eukaryotic ribosome

et al. 2021

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Macrolide antibiotics bind in the nascent peptide exit tunnel of the bacterial ribosome and prevent polymerization of specific amino acid sequences, selectively inhibiting translation of a subset of proteins. Because preventing translation of individual proteins could be beneficial for the treatment of human diseases, we asked whether macrolides, if bound to the eukaryotic ribosome, would retain their context- and protein-specific action. By introducing a single mutation in rRNA, we rendered yeast Saccharomyces cerevisiae cells sensitive to macrolides. Cryo-EM structural analysis showed that the macrolide telithromycin binds in the tunnel of the engineered eukaryotic ribosome. Genome-wide analysis of cellular translation and biochemical studies demonstrated that the drug inhibits eukaryotic translation by preferentially stalling ribosomes at distinct sequence motifs. Context-specific action markedly depends on the macrolide structure. Eliminating macrolide-arrest motifs from a protein renders its translation macrolide-tolerant. Our data illuminate the prospects of adapting macrolides for protein-selective translation inhibition in eukaryotic cells.

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“…The tetracenomycins were previously thought to exhibit a mechanism of action like the anthracyclines, namely, binding to DNA topoisomerase II and induction of DNA damage [3] . Recently, Osterman and coworkers demonstrated that 3 does not induce DNA damage, but rather it inhibits peptide translation via binding to the large ribosomal subunit polypeptide exit channel [5] . Impressively, Osterman et al demonstrated that 3 binds to the same binding site within the exit channel of the E. coli 50S ribosomal subunit and the H. sapiens 60S ribosomal subunit, a stunning display of evolutionarily conserved molecular recognition that accounts for the cytotoxic activities of the TCMs [5] .…”

Section: Introductionmentioning

confidence: 99%

A BioBricks^® toolbox for multiplexed metabolic engineering of central carbon metabolism in the tetracenomycin pathway

Nguyen

Riebschleger

Brown

et al. 2021

Preprint

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The tetracenomycins are aromatic anticancer polyketides that inhibit peptide translation via binding to the large ribosomal subunit. Here, we expressed the elloramycin biosynthetic gene cluster in the heterologous host Streptomyces coelicolor M1146 to facilitate the downstream production of tetracenomycin analogs. We developed a BioBricks genetic toolbox of genetic parts for substrate precursor engineering in S. coelicolor M1146::cos16F4iE. We cloned a series of integrating vectors based on the VWB, TG1, and SV1 integrase systems to interrogate gene expression in the chromosome. We genetically engineered three separate genetic constructs to modulate tetracenomycin biosynthesis: 1) the vhb hemoglobin from obligate aerobe Vitreoscilla stercoraria to improve oxygen utilization; (2) the accA2BE acetyl-CoA carboxylase to enhance condensation of malonyl-CoA; (3) lastly, the sco6196 acyltransferase, which is a metabolic regulatory switch responsible for mobilizing triacylglycerols to beta-oxidation machinery for acetyl-CoA. In addition, we engineered the tcmO 8-O-methyltransferase and newly identified tcmD 12-O-methyltransferase from Amycolatopsis sp. A23 to generate tetracenomycins C and X. We also co-expressed the tcmO methyltransferase with oxygenase urdE to generate the analog 6-hydroxy-tetracenomycin C. Altogether, this system is compatible with the BioBricks [RFC 10] cloning standard for the co-expression of multiple gene sets for metabolic engineering of Streptomyces coelicolor M1146::cos16F4iE.

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Tetracenomycin X inhibits translation by binding within the ribosomal exit tunnel

Cited by 60 publications

References 60 publications

METTL18-mediated histidine methylation on RPL3 modulates translation elongation for proteostasis maintenance

METTL18-mediated histidine methylation on RPL3 modulates translation elongation for proteostasis maintenance

Context-specific action of macrolide antibiotics on the eukaryotic ribosome

A BioBricks^® toolbox for multiplexed metabolic engineering of central carbon metabolism in the tetracenomycin pathway

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Tetracenomycin X inhibits translation by binding within the ribosomal exit tunnel

Cited by 60 publications

References 60 publications

METTL18-mediated histidine methylation on RPL3 modulates translation elongation for proteostasis maintenance

METTL18-mediated histidine methylation on RPL3 modulates translation elongation for proteostasis maintenance

Context-specific action of macrolide antibiotics on the eukaryotic ribosome

A BioBricks® toolbox for multiplexed metabolic engineering of central carbon metabolism in the tetracenomycin pathway

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Product

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A BioBricks^® toolbox for multiplexed metabolic engineering of central carbon metabolism in the tetracenomycin pathway