Structural basis for coupling of the WASH subunit FAM21 with the endosomal SNX27-Retromer complex

Guo, Qian; Chen, Kai‐En; Giménez-Andrés, Manuel; Jellett, Adam P.; Ya, Gao; Simonetti, Boris; Liu, Meihan; Danson, Chris M.; Heesom, Kate J; Cullen, Peter J.; Collins, Brett M.

doi:10.1101/2023.08.15.553351

Cited by 5 publications

(4 citation statements)

References 88 publications

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“…A second involves binding of a Leu and Phe containing sequence in Vps5 to the conserved convex surface of the Vps35 a-helical solenoid. This is present in species such as S. cerevisiae, but absent in C. thermophilum, and the binding mechanism is very similar to the recently identified LFa motifs of FAM21 interacting with the human Retromer complex 42 . These findings are completely consistent with both the cryoET structures of membrane assembled Retromer and Vps5 31,32 , as well as the cellular studies demonstrating the importance of Vps29 and Vps5 N-terminal regions in complex formation 17,33,34 .…”

Section: Discussionsupporting

confidence: 65%

“…4A, S7A and S7B). This interaction involves Leu and Phe sidechains of Vps5 binding the C-terminal end of Vps35, remarkably resembling the LFa repeat motifs (Leu, Phe and acidic sidechains) in human FAM21 protein that, as we recently showed, bind to the same region of human VPS35 42 . We speculate that Vps5 orthologues containing these three sequences together will create an even higher binding affinity for Retromer due to enhanced avidity.…”

Section: High Affinity Interaction With Vps5 Is Due To Avidity Of Bin...supporting

confidence: 71%

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Molecular basis for the assembly of the Vps5-Vps17 SNX-BAR proteins with Retromer

Chen,

Tillu,

Gopaldass

et al. 2024

Preprint

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Retromer mediates endosomal retrieval of transmembrane proteins in all eukaryotes and was first discovered in yeast in complex with the Vps5 and Vps17 sorting nexins (SNXs). Cryoelectron tomography (cryoET) studies of Retromer-Vps5 revealed a pseudo-helical coat on membrane tubules where dimers of the Vps26 subunit bind Vps5 membrane-proximal domains. However, the Vps29 subunit is also required for Vps5-Vps17 association despite being far from the membrane. Here, we show that Vps5 binds both Vps29 and Vps35 subunits through its unstructured N-terminal domain. A Pro-Leu (PL) motif in Vps5 binds Vps29 and is required for association with Retromer on membrane tubules in vitro, and for the proper recycling of the Vps10 cargo in Saccharomyces cerevisiae. CryoET of Retromer tubules with Vps5-Vps17 heterodimers show a similar architecture to the coat with Vps5-Vps5 homodimers, however, the spatial relationship between Retromer units is highly restricted, likely due to more limited orientations for docking. These results provide new mechanistic insights into how Retromer and SNX-BAR association has evolved across species.

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Section: Discussionsupporting

confidence: 65%

Section: High Affinity Interaction With Vps5 Is Due To Avidity Of Bin...supporting

confidence: 71%

Molecular basis for the assembly of the Vps5-Vps17 SNX-BAR proteins with Retromer

Chen,

Tillu,

Gopaldass

et al. 2024

Preprint

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“…The bacterial pathogen L. pneumophila interferes with retromer by binding the VPS29 pocket through its effector protein RidL (Finsel et al, 2013). Additionally, the Vrl1 homolog and RAB21 GEF VARP, the RAB7 GAP TBC1D5, the Commander complex subunit VPS35L, and the WASH complex subunit FAM21 also compete for this site (Crawley-Snowdon et al, 2020; Hesketh et al, 2014; Healy et al 2023; Guo et al 2023), suggesting that the VPS29 pocket is a multi-functional interface that could spatiotemporally control Rab dynamics and regulate interactions with sorting nexins and other accessory proteins. Further work is needed to determine the relevance of the interaction between SNX1 and the VPS29 pocket in vivo , and its role in regulating the incorporation of metazoan retromer into a variety of sorting complexes.…”

Section: Discussionmentioning

confidence: 99%

N-terminal signals in the SNX-BAR paralogs Vps5 and Vin1 guide coat complex formation

Shortill,

Frier,

Davey

et al. 2024

Preprint

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Endosomal coat complexes assemble by incorporating membrane-binding subunits such as those of the sorting nexin (SNX) family. TheS. cerevisiaeSNX-BAR paralogs Vin1 and Vps5 are respective subunits of the endosomal VINE and retromer complexes that arose from a fungal whole genome duplication. Interactions mediated by the Vin1 and Vps5 BAR domains are required for protein complex assembly and membrane association. However, a degree of promiscuity is predicted for yeast BAR-BAR pairings, suggesting that another mechanism guides the formation of specific endosomal coat complexes. Previous work by our group and others has implicated the unstructured N-terminal domains of Vin1 and Vps5 in complex assembly. Here, we map N-terminal signals in both SNX-BAR paralogs that contribute to the formation and function of two distinct endosomal coatsin vivo. Whereas Vin1 leverages a polybasic region and adjacent hydrophobic motif to bind Vrl1 and form VINE, the N-terminus of Vps5 interacts with the retromer subunit Vps29 at two separate sites. We show that one of these Vps5 motifs binds to a conserved hydrophobic pocket in Vps29 that is shared with other accessory proteins and targeted by a bacterial virulence factor in humans. Lastly, we examined the sole isoform of Vps5 from the milk yeastK. lactisand found that ancestral yeasts may have used a nested N-terminal signal to form both VINE and retromer. Our results suggest that the specific assembly of Vps5-family SNX-BAR coats depends on inputs from unique N-terminal sequence features in addition to BAR domain coupling, expanding our understanding of endosomal coat assembly mechanisms.

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“…, 2020 ; Healy et al. 2023 ; Guo et al. 2023 ), suggesting that the VPS29 pocket is a multifunctional interface that could spatiotemporally control Rab dynamics and regulate interactions with SNXs and other accessory proteins.…”

Section: Discussionmentioning

confidence: 99%

N-terminal signals in the SNX-BAR paralogs Vps5 and Vin1 guide endosomal coat complex formation

Shortill,

Frier,

Davey

et al. 2024

MBoC

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Endosomal coats incorporate membrane-binding subunits such as sorting nexin (SNX) proteins. The Saccharomyces cerevisiae SNX-BAR paralogs Vin1 and Vps5 are respective subunits of the endosomal VINE and retromer complexes whose dimerizing BAR domains are required for complex assembly and membrane association. However, a degree of promiscuity is predicted for yeast BAR-BAR pairings, and recent work has implicated the unstructured N-terminal domains of Vin1 and Vps5 in coat formation. Here, we map N-terminal signals in both SNX-BAR paralogs that contribute to the assembly and function of two distinct endosomal coats in vivo. Whereas Vin1 leverages a polybasic region and adjacent hydrophobic motif to bind Vrl1 and form VINE, the N-terminus of Vps5 interacts with the retromer subunit Vps29 at two sites, including a conserved hydrophobic pocket in Vps29 that engages other accessory proteins in humans. We also examined the sole isoform of Vps5 from the milk yeast Kluyveromyces lactis and found that ancestral yeasts may have used a nested N-terminal signal to form both VINE and retromer. Our results suggest that the specific assembly of Vps5-family SNX-BAR coats depends on inputs from unique N-terminal sequence features in addition to BAR domain coupling, expanding our understanding of endosomal coat biology.

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Structural basis for coupling of the WASH subunit FAM21 with the endosomal SNX27-Retromer complex

Cited by 5 publications

References 88 publications

Molecular basis for the assembly of the Vps5-Vps17 SNX-BAR proteins with Retromer

Molecular basis for the assembly of the Vps5-Vps17 SNX-BAR proteins with Retromer

N-terminal signals in the SNX-BAR paralogs Vps5 and Vin1 guide coat complex formation

N-terminal signals in the SNX-BAR paralogs Vps5 and Vin1 guide endosomal coat complex formation

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