The last two transmembrane helices in the APC-type FurE transporter act as an intramolecular chaperone essential for concentrative ER-exit
Authors:Yiannis Pyrris1, Georgia F. Papadaki1, Emmanuel Mikros2 and George Diallinas1,3
1 Department of Biology, National and Kapodistrian University of Athens, Panepistimioupolis, Athens, 15784, Greece.
2 Department of Pharmacy, National and Kapodistrian University of Athens, Panepistimioupolis, Athens, 15771, Greece.
3 Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology, Heraklion, 70013, Greece.
Keywords:
Aspergillus nidulans, transporter, membrane traffic, NCS1, sorting, ER exit.
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Conflict of interest statement:
The authors declare no conflict of interest.
Please cite this article as:
Yiannis Pyrris, Georgia F. Papadaki, Emmanuel Mikros and George Diallinas (2024). The last two transmembrane helices in the APC-type FurE transporter act as an intramolecular chaper-one essential for concentrative ER-exit. Microbial Cell 11: 1-15. doi: 10.15698/mic2024.01.811
© 2024 Pyrris et al. This is an open-access article released under the terms of the Creative Commons Attribution (CC BY) license, which allows the unrestricted use, distribution, and reproduc-tion in any medium, provided the original author and source are acknowledged.
Abstract:
FurE is a H+ symporter specific for the cellular uptake of uric acid, allantoin, uracil, and toxic nucleobase analogues in the fungus Aspergillus nidulans. Being member of the NCS1 protein family, FurE is structurally related to the APC-superfamily of transporters. APC-type transporters are characterised by a 5+5 inverted repeat fold made of ten transmembrane segments (TMS1-10) and function through the rocking-bundle mechanism. Most APC-type transporters possess two extra C-terminal TMS segments (TMS11-12), the function of which remains elusive. Here we present a systematic mutational analysis of TMS11-12 of FurE and show that two specific aromatic residues in TMS12, Trp473 and Tyr484, are essential for ER-exit and trafficking to the plasma membrane (PM). Molecular modeling shows that Trp473 and Tyr484 might be essential through dynamic interactions with residues in TMS2 (Leu91), TMS3 (Phe111), TMS10 (Val404, Asp406) and other aromatic residues in TMS12. Genetic analysis confirms the essential role of Phe111, Asp406 and TMS12 aromatic residues in FurE ER-exit. We further show that co-expression of FurE-Y484F or FurE-W473A with wild-type FurE leads to a dominant negative phenotype, compatible with the concept that FurE molecules oligomerize or partition in specific microdomains to achieve concentrative ER-exit and traffic to the PM. Importantly, truncated FurE versions lacking TMS11-12 are unable to reproduce a negative effect on the trafficking of co-expressed wild-type FurE. Overall, we show that TMS11-12 acts as an intramolecular chaperone for proper FurE folding, which seems to provide a structural code for FurE partitioning in ER-exit sites.
doi: 10.15698/mic2024.01.811
Volume 11, pp. 1 to 15, published 05/01/2024.