Signal peptide mimicry primes Sec61 for client-selective inhibition

Rehan, Shahid; Tranter, Dale; Sharp, Phillip P.; Lowe, Eric; Anderl, Janet L.; Muchamuel, Tony; Abrishami, Vahid; Kuivanen, Suvi; Wenzell, Nicole A.; Jennings, Andy; Kalyanaraman, Chakrapani; Craven, Gregory B.; Strandin, Tomas; Javanainen, Matti; Vapalahti, Olli; Jacobson, Matt; McMinn, Dustin; Kirk, Christopher J.; Huiskonen, Juha T.; Taunton, Jack; Paavilainen, Ville O.

doi:10.1101/2022.07.03.498529

Cited by 11 publications

(25 citation statements)

References 60 publications

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“…In the cryo-EM structure, the macrocyclic cotransin inhibitor is bound to the Sec61 complex with an open lateral gate and a closed plug helix (Rehan et al 2022). This is similar as observed in the cryo-ET study of the Sec61/TRAP complex in the ER membrane, where surprisingly the lateral gate was observed to be open in majority of Sec61 molecules (Pfeffer et al 2017).…”

Section: Resultsmentioning

confidence: 99%

“…3H ). The simulations were initiated starting from an open conformation observed in our structure of Sec61 primed with a macrocyclic cotransin inhibitor (Rehan et al 2022). In simulations without TRAP, we observed the lateral gate closing rapidly, whereas simulations with TRAP included retained an open lateral gate conformation.…”

Section: Resultsmentioning

confidence: 99%

“…Our work now suggests that alterations to the ER membrane may also impact Sec61-mediated protein insertion and highlights the importance of understanding the localized membrane effects resulting from Sec61 cofactor association. Further, different natural and synthetic small molecule inhibitors of Sec61 are accommodated within Sec61 in subtly different open conformations (Rehan et al 2022); (Itskanov et al 2022) and detailed understanding of Sec61 conformational control by the surrounding lipid and protein environment will be important for targeting Sec61 with therapeutic small molecules. Further structural and mechanistic work is required to understand a possible direct role that TRAP may have in directing protein N-glycosylation with OST.…”

Section: Discussionmentioning

confidence: 99%

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Molecular view of ER membrane remodeling by the Sec61/TRAP translocon

Karki

Javanainen

Tranter

et al. 2022

Preprint

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Protein translocation across the endoplasmic reticulum (ER) membrane is an essential initial step in protein entry into the secretory pathway. The conserved Sec61 protein translocon facilitates polypeptide translocation and coordinates cotranslational polypeptide processing events. In cells, the majority of Sec61 is stably associated with a heterotetrameric membrane protein complex, the translocon associated protein complex (TRAP), yet the mechanism by which TRAP assists in polypeptide translocation or cotranslational modifications such as N-glycosylation remains unknown. Here, we demonstrate the structure of the core Sec61/TRAP complex bound to a mammalian ribosome by Cryo-EM. The interactions with the ribosome anchor the Sec61/TRAP complex in a conformation that renders the ER membrane locally thinner by significantly curving its the lumenal leaflet. We propose a model for how TRAP stabilizes the ribosome exit tunnel to assist nascent polypeptide insertion through Sec61 and provides a ratcheting mechanism into the ER lumen by direct polypeptide interactions.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Molecular view of ER membrane remodeling by the Sec61/TRAP translocon

Karki

Javanainen

Tranter

et al. 2022

Preprint

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“…Our data suggest that the density feature previously assigned as mycolactone is unlikely to be mycolactone. During the preparation of this manuscript, a medium-resolution cryo-EM structure of the mammalian Sec61 channel in association with a ribosome and a cotransin derivative has been reported 53 . While the overall structure of the channel and the location of the binding pocket seem consistent with ours, we note that the orientation of the inhibitor model is substantially different from that of cotransin from our study.…”

Section: Resultsmentioning

confidence: 99%

A common mechanism of Sec61 translocon inhibition by small molecules

Itskanov

Wang

Junne

et al. 2022

Preprint

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The Sec61 complex forms a protein-conducting channel in the endoplasmic reticulum (ER) membrane that is required for secretion of soluble proteins and production of many membrane proteins. Several natural and synthetic small molecules specifically inhibit the Sec61 channel, generating cellular effects that are potentially useful for therapeutic purposes, but their inhibitory mechanisms remain unclear. Here we present near-atomic-resolution structures of the human Sec61 channel inhibited by a comprehensive panel of structurally distinct small molecules--cotransin, decatransin, apratoxin F, ipomoeassin F, mycolactone, cyclotriazadisulfonamide (CADA) and eeyarestatin I (ESI). Remarkably, all inhibitors bind to a common lipid-exposed pocket formed by the partially open lateral gate and plug domain of the channel. Mutations conferring resistance to the inhibitors are clustered at this binding pocket. The structures indicate that Sec61 inhibitors stabilize the plug domain of Sec61 in a closed state, thereby preventing the protein-translocation pore from opening. Our study reveals molecular interactions between Sec61 and its inhibitors in atomic detail and offers the structural framework for further pharmacological studies and drug design.

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“…A recent cryo-EM study of the Sec61 translocon exposed to a novel cotransin analog, KZR-8445 containing a large cyclic heptadepsipeptide ring, reveals binding of the inhibitor to the SPbinding cleft (53). Another recent Sec61 structural study of multiple translocation inhibitors, including mycolactone and CADA, showed an overlapping binding site near the lateral gate of Sec61 for all the inhibitors (50).…”

Section: Different Inhibitor Binding Sites In Sec61mentioning

confidence: 99%

Structural insights into TRAP association with ribosome-Sec61 complex, and translocon inhibition by a CADA derivative

Pauwels

Shewakramani

Wijngaert

et al. 2022

Preprint

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During co-translational translocation, the signal peptide of a nascent chain binds Sec61 translocon to initiate protein transport through the ER membrane. Our cryo-EM structure of ribosome-Sec61 shows binding of an ordered heterotetrameric TRranslocon-Associated Protein (TRAP) complex, in which TRAP-γ is anchored at two adjacent positions of 28S rRNA and interacts with ribosomal protein L38 and Sec61α/γ. Four transmembrane helices (TMHs) of TRAP-γ cluster with one C-terminal helix of each α, α, and α subunits. The seven TMH bundle helps position a crescent-shaped trimeric TRAP-α/α/α core in the ER lumen, facing the Sec61 channel. Further, our in vitro assay establishes the CADA derivative CK147 as a translocon inhibitor. A structure of ribosome-Sec61-CK147 reveals CK147 binding the channel and interacting with the plug helix from the lumenal side. The CK147-resistance mutations surround the inhibitor. These structures help in understanding the TRAP functions and provide a new Sec61 site for designing translocon inhibitors.

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Signal peptide mimicry primes Sec61 for client-selective inhibition

Cited by 11 publications

References 60 publications

Molecular view of ER membrane remodeling by the Sec61/TRAP translocon

Molecular view of ER membrane remodeling by the Sec61/TRAP translocon

A common mechanism of Sec61 translocon inhibition by small molecules

Structural insights into TRAP association with ribosome-Sec61 complex, and translocon inhibition by a CADA derivative

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