Structural basis of autoinhibition of the human NHE3-CHP1 complex

Dong, Yanli; Li, Hang; Ilie, Alina; Gao, Yongfei; Boucher, Annie; Zhang, Xuejun C.; Orlowski, John; Zhao, Yan

doi:10.1126/sciadv.abn3925

Cited by 16 publications

(19 citation statements)

References 59 publications

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“…However, in the presence of PA, His73 interacts with the lipid, and the TM2′–TM3′ extracellular region has rearranged to coordinate the PA headgroup through water-bridged His132 and Asp129. The lipid-induced rearrangement observed in SLC9C1 is centred around an extracellular helical motif (ECH1) that is not present in the NHE1 19 , NHE3 35 and NHE9 18 structures (Fig. 2a and Supplementary Fig.…”

Section: The Nhe Transporter Module Of Slc9c1mentioning

confidence: 99%

Structure and electromechanical coupling of a voltage-gated Na+/H+ exchanger

Yeo,

Mehta,

Gulati

et al. 2023

Nature

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Voltage-sensing domains control the activation of voltage-gated ion channels, with a few exceptions1. One such exception is the sperm-specific Na+/H+ exchanger SLC9C1, which is the only known transporter to be regulated by voltage-sensing domains2–5. After hyperpolarization of sperm flagella, SLC9C1 becomes active, causing pH alkalinization and CatSper Ca2+ channel activation, which drives chemotaxis2,6. SLC9C1 activation is further regulated by cAMP2,7, which is produced by soluble adenyl cyclase (sAC). SLC9C1 is therefore an essential component of the pH–sAC–cAMP signalling pathway in metazoa8,9, required for sperm motility and fertilization4. Despite its importance, the molecular basis of SLC9C1 voltage activation is unclear. Here we report cryo-electron microscopy (cryo-EM) structures of sea urchin SLC9C1 in detergent and nanodiscs. We show that the voltage-sensing domains are positioned in an unusual configuration, sandwiching each side of the SLC9C1 homodimer. The S4 segment is very long, 90 Å in length, and connects the voltage-sensing domains to the cytoplasmic cyclic-nucleotide-binding domains. The S4 segment is in the up configuration—the inactive state of SLC9C1. Consistently, although a negatively charged cavity is accessible for Na+ to bind to the ion-transporting domains of SLC9C1, an intracellular helix connected to S4 restricts their movement. On the basis of the differences in the cryo-EM structure of SLC9C1 in the presence of cAMP, we propose that, upon hyperpolarization, the S4 segment moves down, removing this constriction and enabling Na+/H+ exchange.

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Section: The Nhe Transporter Module Of Slc9c1mentioning

confidence: 99%

Structure and electromechanical coupling of a voltage-gated Na+/H+ exchanger

Yeo,

Mehta,

Gulati

et al. 2023

Nature

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“…Here, we have compared the lipid-mediated oligomerization in the Na + /H + exchanger E. coli NhaA and the endosomal exchanger horse NHE9, which both harbour β-hairpins located between topologically equivalent helices in the scaffold domain. The scaffold β-hairpin loop domain is absent is bacterial Na + /H + NapA (Lee et al ., 2013) and NhaP (Paulino et al ., 2014) members, and in mammalian NHEs localised to the plasma membrane (Dong et al ., 2021; Dong et al , 2022). Moreover, the high structural-similarity between plasma membrane localised NHE1 and endosomal NHE9, implies that the additional TM2-TM3 β-hairpin loop domain has an additional, regulatory role.…”

Section: Discussionmentioning

confidence: 99%

PI-(3,5)P2-mediated oligomerization of the endosomal sodium/proton exchanger NHE9

Kokane,

Meier,

Gulati

et al. 2023

Preprint

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Na+/H+exchangers (NHE) are found in all cells to regulate intracellular pH, sodium levels and cell volume. The NHE isoform 9 (SLC9A9) fine-tunes endosomal pH, and its activity is linked to glioblastoma, epilepsy, autism spectrum and attention-deficit-hyperactivity disorders. Here, we report cryo-EM structures ofhorseNHE9 and a cysteine-variant at 3.6 and 3.1 Å resolution, respectively. We show how lysine residues, from a previously unresolved TM2-TM3 β-hairpin loop domain, are positioned above the dimerization interface and interact with the endosomal-specific PI-(3,5)P2lipid, together with residues located on dimer domain helices. Thermal-shift assays, solid-state membrane (SSM) electrophysiology and MD simulations, corroborates that NHE9 can specifically bind PI-(3,5)P2, and that its addition stabilizes the homodimer and enhances NHE9 activity. We have further determined the cryo-EM structure ofE. coliNhaA, confirming the expected coordination of cardiolipin at the dimerization interface, solidifying the concept that Na+/H+exchanger dimerization and transporter activity can be regulated by specific lipids. Taken together, we propose that the activity of NHE9 is regulated by the PI-(3,5)P2lipid upon reaching endosomes, which we refer to as an lipid-activation-upon-arrival model.

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“…1d and Supplementary Video 1 ). The overall arrangement of the cytoplasmic domains and ARD of sp9C1 is similar to the SOS1 transporter 17 , which is a plant ortholog of SLC9s, although the latter does not feature a VSD and is not regulated by cAMP.…”

Section: Introductionmentioning

confidence: 91%

“…SLC9C exhibit characteristic differences from other SLC9s in terms of their structural and regulatory attributes. In SLC9As and 9Bs, and their prokaryotic orthologs of known structure, the membrane-delimited ion-exchange domains (EDs) are tethered to relatively short or largely unstructured C-terminal soluble domains [13][14][15][16][17][18] . In contrast, SLC9Cs, feature a homologous ED, a canonical voltagesensing domain (VSD) and a cyclic nucleotide binding domain (CNBD), interconnected via long, structured linkers, all within a single polypeptide.…”

Section: Introductionmentioning

confidence: 99%

Architecture and rearrangements of a sperm-specific Na⁺/H⁺exchanger

Pal,

Chowdhury

2023

Preprint

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The sperm-specific sodium hydrogen exchanger, SLC9C1, underlies hyperpolarization and cyclic nucleotide stimulated proton fluxes across sperm membranes and regulates their hyperactivated motility. SLC9C1 is the first known instance of an ion transporter that uses a canonical voltage-sensing domain (VSD) and an evolutionarily conserved cyclic nucleotide binding domain (CNBD) to influence the dynamics of its ion-exchange domain (ED). The structural organization of this ‘tripartite transporter’ and the mechanisms whereby it integrates physical (membrane voltage) and chemical (cyclic nucleotide) cues are unknown. In this study, we use single particle cryo-electron microscopy to determine structures of a metazoan SLC9C1 in different conformational states. We find that the three structural domains are uniquely organized around a distinct ring-shaped scaffold that we call the ‘allosteric ring domain’ or ARD. The ARD undergoes coupled proton-dependent rearrangements with the ED and acts as a ‘signaling hub’ enabling allosteric communication between the key functional modules of sp9C1. We demonstrate that binding of cAMP causes large conformational changes in the cytoplasmic domains and disrupts key ARD-linked interfaces. We propose that these structural changes rescue the transmembrane domains from an auto-inhibited state and facilitate their functional dynamics. Our study provides a structural framework to understand and further probe electrochemical linkage in SLC9C1.

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Structural basis of autoinhibition of the human NHE3-CHP1 complex

Cited by 16 publications

References 59 publications

Structure and electromechanical coupling of a voltage-gated Na+/H+ exchanger

Structure and electromechanical coupling of a voltage-gated Na+/H+ exchanger

PI-(3,5)P2-mediated oligomerization of the endosomal sodium/proton exchanger NHE9

Architecture and rearrangements of a sperm-specific Na⁺/H⁺exchanger

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Structural basis of autoinhibition of the human NHE3-CHP1 complex

Cited by 16 publications

References 59 publications

Structure and electromechanical coupling of a voltage-gated Na+/H+ exchanger

Structure and electromechanical coupling of a voltage-gated Na+/H+ exchanger

PI-(3,5)P2-mediated oligomerization of the endosomal sodium/proton exchanger NHE9

Architecture and rearrangements of a sperm-specific Na+/H+exchanger

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Architecture and rearrangements of a sperm-specific Na⁺/H⁺exchanger