Specific residues in a purine transporter are critical for dimerization, ER-Exit and function.
File(s)
Accepted version
Author(s)
Type
Journal Article
Abstract
Transporters are transmembrane proteins that mediate the selective translocation of solutes across biological membranes. Recently, we have shown that specific interactions with plasma membrane phospholipids are essential for formation and/or stability of functional dimers of the purine transporter, UapA, a prototypic eukaryotic member of the ubiquitous NAT family. Here, we provide strong evidence that distinct interactions of UapA with membrane lipids are essential for ab initio formation of functional dimers in the ER or ER-exit and further subcellular trafficking. Through genetic screens we identify mutations that restore defects in dimer formation and/or trafficking. Suppressors of defective dimerization restore ab initio formation of UapA dimers in the ER. Most of these suppressors are located in the movable core domain, but also in the core-dimerization interface and in residues of the dimerization domain exposed to lipids. Molecular Dynamics suggest the majority of suppressors stabilize interhelical interactions in the core domain and thus assist the formation of functional UapA dimers. Among suppressors restoring dimerization, a specific mutation, T401P, was also isolated independently as a suppressor restoring trafficking, suggesting that stabilization of the core domain restores function by sustaining structural defects caused by abolishment of essential interactions with specific lipids. Importantly, introduction of mutations topologically equivalent to T401P into a rat homologue of UapA, namely rSNBT1, permitted the functional expression of a mammalian NAT in A. nidulans Thus, our results provide a potential route for the functional expression and manipulation of mammalian transporters in the model Aspergillus system.
Date Issued
2019-12-01
Date Acceptance
2019-10-02
Citation
Genetics, 2019, 213 (4), pp.1357-1372
ISSN
1943-2631
Publisher
Oxford University Press
Start Page
1357
End Page
1372
Journal / Book Title
Genetics
Volume
213
Issue
4
Copyright Statement
© Genetics 2019
This article is published and distributed under the terms of the Oxford University Press, Standard Journals Publication Model (https://academic.oup.com/journals/pages/open_access/funder_policies/chorus/standard_publication_model). This is a pre-copy-editing, author-produced version of an article accepted for publication in Genetics following peer review. The definitive publisher-authenticated version Anezia Kourkoulou, Pothos Grevias, George Lambrinidis, Euan Pyle, Mariangela Dionysopoulou, Argyris Politis, Emmanuel Mikros, Bernadette Byrne, George Diallinas, Specific Residues in a Purine Transporter Are Critical for Dimerization, ER Exit, and Function, Genetics, Volume 213, Issue 4, 1 December 2019, Pages 1357–1372, https://doi.org/10.1534/genetics.119.302566
This article is published and distributed under the terms of the Oxford University Press, Standard Journals Publication Model (https://academic.oup.com/journals/pages/open_access/funder_policies/chorus/standard_publication_model). This is a pre-copy-editing, author-produced version of an article accepted for publication in Genetics following peer review. The definitive publisher-authenticated version Anezia Kourkoulou, Pothos Grevias, George Lambrinidis, Euan Pyle, Mariangela Dionysopoulou, Argyris Politis, Emmanuel Mikros, Bernadette Byrne, George Diallinas, Specific Residues in a Purine Transporter Are Critical for Dimerization, ER Exit, and Function, Genetics, Volume 213, Issue 4, 1 December 2019, Pages 1357–1372, https://doi.org/10.1534/genetics.119.302566
Sponsor
Biotechnology and Biological Sciences Research Council (BBSRC)
Identifier
https://www.ncbi.nlm.nih.gov/pubmed/31611232
PII: genetics.119.302566
Grant Number
BB/K017292/1
Subjects
Aspergillus nidulans
NAT
UapA
nucleobase
rSNBT1
Publication Status
Published
Coverage Spatial
United States
Date Publish Online
2019-10-14