Puromycin reveals a distinct conformation of neuronal ribosomes

Anadolu, MN; Sun, J; Li, JT-Y; Graber, TE; Ortega, J; Sossin, WS

Journal article

Puromycin reveals a distinct conformation of neuronal ribosomes

Abstract:: Puromycin is covalently added to the nascent chain of proteins by the peptidyl transferase activity of the ribosome and the dissociation of the puromycylated peptide typically follows this event. It was postulated that blocking the translocation of the ribosome with emetine could retain the puromycylated peptide on the ribosome, but evidence against this has recently been published [Hobson et al., Elife 9, e60048 (2020); and Enam et al., Elife 9, e60303 (2020)]. In neurons, puromycylated nascent chains remain in the ribosome even in the absence of emetine, yet direct evidence for this has been lacking. Using biochemistry and cryoelectron microscopy, we show that the puromycylated peptides remain in the ribosome exit channel in the large subunit in a subset of neuronal ribosomes stalled in the hybrid state. These results validate previous experiments to localize stalled polysomes in neurons and provide insight into how neuronal ribosomes are stalled. Moreover, in these hybrid-state neuronal ribosomes, anisomycin, which usually blocks puromycylation, competes poorly with puromycin in the puromycylation reaction, allowing a simple assay to determine the proportion of nascent chains that are stalled in this state. In early hippocampal neuronal cultures, over 50% of all nascent peptides are found in these stalled polysomes. These results provide insights into the stalling mechanisms of neuronal ribosomes and suggest that puromycylated peptides can be used to reveal subcellular sites of hybrid-state stalled ribosomes in neurons.

Publication status:: Published

Peer review status:: Peer reviewed

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APA Style

Anadolu, M. N., Sun, J., Li, J. T.-Y., Graber, T. E., Ortega, J., & Sossin, W. S. (2024). Puromycin reveals a distinct conformation of neuronal ribosomes. Proceedings of the National Academy of Sciences, 121(7), e2306993121–e2306993121.

MLA Style

Anadolu, MN, et al. “Puromycin Reveals a Distinct Conformation of Neuronal Ribosomes.” Proceedings of the National Academy of Sciences, vol. 121, no. 7, 2024, pp. e2306993121–e2306993121.

Chicago Style

Anadolu, MN, J Sun, JT-Y Li, TE Graber, J Ortega, and WS Sossin. 2024. “Puromycin Reveals a Distinct Conformation of Neuronal Ribosomes.” Proceedings of the National Academy of Sciences 121 (7): e2306993121–e2306993121.
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