The crystal structure of MICU 2 provides insight into Ca ²⁺ binding and MICU 1‐ MICU 2 heterodimer formation

Abstract

The mitochondrial calcium uniporter (MCU) complex mediates the uptake of Ca2+ into mitochondria. Its activity is regulated by a heterodimer of MICU1 and MICU2, two EF‐hand‐containing proteins that act as the main gatekeeper of the uniporter. Herein we report the crystal structure of human MICU2 at 1.96 Å resolution. Our structure reveals a dimeric architecture of MICU2, in which each monomer adopts the canonical two‐lobe structure with a pair of EF‐hands in each lobe. Both Ca2+‐bound and Ca2+‐free EF‐hands are observed in our structure. Moreover, we characterize the interaction sites within the MICU2 homodimer, as well as the MICU1–MICU2 heterodimer in both Ca2+‐free and Ca2+‐bound conditions. Glu242 in MICU1 and Arg352 in MICU2 are crucial for apo heterodimer formation, while Phe383 in MICU1 and Glu196 in MICU2 significantly contribute to the interaction in the Ca2+‐bound state. Based on our structural and biochemical analyses, we propose a model for MICU1–MICU2 heterodimer formation and its conformational transition from apo to a more compact Ca2+‐bound state, which expands our understanding of this co‐regulatory mechanism critical for MCU's mitochondrial calcium uptake function.

Structure of the MICU1–MICU2 heterodimer provides insights into the gatekeeping threshold shift

“…Similarly, Arg221(MICU1)-Asp330(MICU2), in the heterodimer, would also be important for the heterodimerization and function, as consistent with previous results (Li et al, 2016). In the case of the other salt bridge, Asp231(MICU1)-Arg352(MICU2), Wu et al already reported that this salt bridge is essential for the heterodimerization of MICU1 and MICU2 in the apo state (Wu et al, 2019). Thus, we assumed that the F-F interface of the MICU1-MICU2 heterodimer is biologically relevant.…”

“…The tight interaction in the apo state of MICU1-MICU2 might hinder the conformational changes required for Ca 2+ binding, resulting in a lower Ca 2+ binding affinity in the MICU1-MICU2 heterodimer compared with that of the MICU1 homodimer. tissues (Plovanich et al, 2013;Paillard et al, 2017;Patron et al, 2019), MICUs in cells generally form MICU1-MICU2 heterodimers, which is consistent with prior biochemical studies showing that the MICU1-MICU2 heterodimer is more stable (K d = 224 nM in the apo state) compared with homodimers of MICU1 and MICU2 (Wagner et al, 2015;Patron et al, 2014;Matesanz-Isabel et al, 2016;Mallilankaraman et al, 2012b;Csordá s et al, 2013;Ahuja & Muallem, 2014;Wu et al, 2019;Kamer et al, 2017). In particular, it has been reported that MICU2 functions as a gatekeeper of the MCU for suppressing the cellular damage caused by excessive mitochondrial Ca 2+ influx Patron et al, 2014).…”

“…The heterodimers were formed by a face-to-face (F-F) interface similar to other MICU homodimer structures [Figs. 1(a) and 1(b)] (Wang et al, 2014;Kamer et al, 2019;Xing et al, 2019;Wu et al, 2019). Unlike MICU homodimer structures, the MICU1-MICU2 heterodimer comprised two interfaces, interfaces 1 and 2, as a result of the asymmetry of the heterodimer.…”

“…Notably, the Asp231(MICU1) was located on the interloop (L6) of EF-hand 1 participating in the Ca 2+ coordination, indicating that Ca 2+ binding induces structural changes directly on interface 1. Arg352(MICU2) was identified as one of two important residues, Glu242(MICU1) and Arg352(MICU2), whose alanine mutation disturbs the heterodimerization of MICU1 and MICU2 in the apo state (Wu et al, 2019). However, Glu242(MICU1) did not directly participate in interface 1 of the structure but stabilized the conformation of the interloop (L6) by forming a hydrogen bond through the backbone amide nitrogen of Leu232 [ Fig.…”

“…This excessive Ca 2+ uptake induces several diseases such as proximal myopathy, learning difficulties and progressive extrapyramidal movement disorders (Logan et al, 2014). However, the structure of the heterodimers is not known and the mechanism underlying the regulation of Ca 2+ uptake by the heterodimer through the MCU is also unclear, although the structures of the homodimers in MICU1, MICU2 and MICU3 have been reported previously (Wang et al, 2014;Kamer et al, 2019;Xing et al, 2019;Wu et al, 2019).…”