Structural insights into regulation of TRPM7 divalent cation uptake by the small GTPase ARL15

Abstract

Cystathionine-β-synthase (CBS)-pair domain divalent metal cation transport mediators (CNNMs) are an evolutionarily conserved family of magnesium transporters. They promote efflux of Mg2+ ions on their own or uptake of divalent cations when coupled to the transient receptor potential ion channel subfamily M member 7 (TRPM7). Recently, ADP-ribosylation factor-like GTPase 15 (ARL15) has been identified as CNNM binding partner and an inhibitor of divalent cation influx by TRPM7. Here, we characterize ARL15 as a GTP-binding protein and demonstrate that it binds the CNNM CBS-pair domain with low micromolar affinity. The crystal structure of the complex between ARL15 GTPase domain and CNNM2 CBS-pair domain reveals the molecular determinants of the interaction and allowed the identification of mutations in ARL15 and CNNM2 mutations that abrogate binding. Loss of CNNM binding prevented ARL15 suppression of TRPM7 channel activity in support of previous reports that the proteins function as a ternary complex. Binding experiments with phosphatase of regenerating liver 2 (PRL2 or PTP4A2) revealed that ARL15 and PRLs compete for binding CNNM, suggesting antagonistic regulation of divalent cation transport by the two proteins.

New insights into the structure and function of CNNM proteins

“…Four groups have characterized the interaction between CNNM and ADP‐ribosylation factor‐like GTPase 15 (ARL15) [97,100–102]. ARL15 locus was associated with urinary Mg 2+ excretion in a GWAS [103].…”

“…Several groups observed that ARL15 inhibits divalent ion uptake in cells [97,101]. This likely occurs through CNNM‐binding as an ARL15 mutant that is unable to bind CNNMs was unable to inhibit Zn 2+ uptake by TRPM7, a bifunctional protein with an ion channel and protein kinase domain involved in Mg 2+ homeostasis [35,101,102,105]. Knockdown and overexpression of ARL15 were also shown to affect N‐glycosylation of CNNMs [100].…”

Dimerization of the CNNM extracellular domain

“…The interaction of CNNMs with PRL phosphatases impairs the CNNM transport function, resulting in elevated intracellular magnesium levels and promoting the progression of metastatic cancer (Funato et al, 2014; Hardy et al, 2015). Additionally, recent findings have demonstrated an interaction between CNNM proteins and another Mg 2+ transporter, TRPM7 (Bai et al, 2021; Hardy et al, 2023; Kollewe et al, 2021; Mahbub et al, 2023).…”