Dataset Information

MICU1 Serves as a Molecular Gatekeeper to Prevent In Vivo Mitochondrial Calcium Overload.

ABSTRACT: MICU1 is a component of the mitochondrial calcium uniporter, a multiprotein complex that also includes MICU2, MCU, and EMRE. Here, we describe a mouse model of MICU1 deficiency. MICU1(-/-) mitochondria demonstrate altered calcium uptake, and deletion of MICU1 results in significant, but not complete, perinatal mortality. Similar to afflicted patients, viable MICU1(-/-) mice manifest marked ataxia and muscle weakness. Early in life, these animals display a range of biochemical abnormalities, including increased resting mitochondrial calcium levels, altered mitochondrial morphology, and reduced ATP. Older MICU1(-/-) mice show marked, spontaneous improvement coincident with improved mitochondrial calcium handling and an age-dependent reduction in EMRE expression. Remarkably, deleting one allele of EMRE helps normalize calcium uptake while simultaneously rescuing the high perinatal mortality observed in young MICU1(-/-) mice. Together, these results demonstrate that MICU1 serves as a molecular gatekeeper preventing calcium overload and suggests that modulating the calcium uniporter could have widespread therapeutic benefits.

SUBMITTER: Liu JC

PROVIDER: S-EPMC5316484 | biostudies-literature | 2016 Aug

REPOSITORIES: biostudies-literature

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MICU1 Serves as a Molecular Gatekeeper to Prevent In Vivo Mitochondrial Calcium Overload.

Liu Julia C JC Liu Jie J Holmström Kira M KM Menazza Sara S Parks Randi J RJ Fergusson Maria M MM Yu Zu-Xi ZX Springer Danielle A DA Halsey Charles C Liu Chengyu C Murphy Elizabeth E Finkel Toren T

Cell reports 20160728 6

MICU1 is a component of the mitochondrial calcium uniporter, a multiprotein complex that also includes MICU2, MCU, and EMRE. Here, we describe a mouse model of MICU1 deficiency. MICU1(-/-) mitochondria demonstrate altered calcium uptake, and deletion of MICU1 results in significant, but not complete, perinatal mortality. Similar to afflicted patients, viable MICU1(-/-) mice manifest marked ataxia and muscle weakness. Early in life, these animals display a range of biochemical abnormalities, incl ...[more]

PMID: 27477272

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Project description:BACKGROUND:The mitochondrial calcium uniporter (mtCU) is an ?700-kD multisubunit channel residing in the inner mitochondrial membrane required for mitochondrial Ca2+ (mCa2+) uptake. Here, we detail the contribution of MCUB, a paralog of the pore-forming subunit MCU, in mtCU regulation and function and for the first time investigate the relevance of MCUB to cardiac physiology. METHODS:We created a stable MCUB knockout cell line (MCUB-/-) using CRISPR-Cas9n technology and generated a cardiac-specific, tamoxifen-inducible MCUB mutant mouse (CAG-CAT-MCUB x MCM; MCUB-Tg) for in vivo assessment of cardiac physiology and response to ischemia/reperfusion injury. Live-cell imaging and high-resolution spectrofluorometery were used to determine intracellular Ca2+ exchange and size-exclusion chromatography; blue native page and immunoprecipitation studies were used to determine the molecular function and impact of MCUB on the high-molecular-weight mtCU complex. RESULTS:Using genetic gain- and loss-of-function approaches, we show that MCUB expression displaces MCU from the functional mtCU complex and thereby decreases the association of mitochondrial calcium uptake 1 and 2 (MICU1/2) to alter channel gating. These molecular changes decrease MICU1/2-dependent cooperative activation of the mtCU, thereby decreasing mCa2+ uptake. Furthermore, we show that MCUB incorporation into the mtCU is a stress-responsive mechanism to limit mCa2+ overload during cardiac injury. Indeed, overexpression of MCUB is sufficient to decrease infarct size after ischemia/reperfusion injury. However, MCUB incorporation into the mtCU does come at a cost; acute decreases in mCa2+ uptake impair mitochondrial energetics and contractile function. CONCLUSIONS:We detail a new regulatory mechanism to modulate mtCU function and mCa2+ uptake. Our results suggest that MCUB-dependent changes in mtCU stoichiometry are a prominent regulatory mechanism to modulate mCa2+ uptake and cellular physiology.

Project description:Background Translocation of miR-181c into cardiac mitochondria downregulates the mitochondrial gene, mt-COX1. miR-181c/d-/- hearts experience less oxidative stress during ischemia/reperfusion (I/R) and are protected against I/R injury. Additionally, miR-181c overexpression can increase mitochondrial matrix Ca2+ ([Ca2+]m), but the mechanism by which miR-181c regulates [Ca2+]m is unknown. Methods and Results By RNA sequencing and analysis, here we show that hearts from miR-181c/d-/- mice overexpress nuclear-encoded Ca2+ regulatory and metabolic pathway genes, suggesting that alterations in miR-181c and mt-COX1 perturb mitochondria-to-nucleus retrograde signaling and [Ca2+]m regulation. Quantitative polymerase chain reaction validation of transcription factors that are known to initiate retrograde signaling revealed significantly higher Sp1 (specificity protein) expression in the miR-181c/d-/- hearts. Furthermore, an association of Sp1 with the promoter region of MICU1 was confirmed by chromatin immunoprecipitation-quantitative polymerase chain reaction and higher expression of MICU1 was found in the miR-181c/d-/- hearts. Conversely, downregulation of Sp1 by small interfering RNA decreased MICU1 expression in neonatal mouse ventricular myocytes. Changes in PDH activity provided evidence for a change in [Ca2+]m via the miR-181c/MICU1 axis. Moreover, this mechanism was implicated in the pathology of I/R injury. When MICU1 was knocked down in the miR-181c/d-/- heart by lentiviral expression of a short-hairpin RNA against MICU1, cardioprotective effects against I/R injury were abrogated. Furthermore, using an in vitro I/R model in miR-181c/d-/- neonatal mouse ventricular myocytes, we confirmed the contribution of both Sp1 and MICU1 in ischemic injury. Conclusions miR-181c regulates mt-COX1, which in turn regulates MICU1 expression through the Sp1-mediated mitochondria-to-nucleus retrograde pathway. Loss of miR-181c can protect the heart from I/R injury by modulating [Ca2+]m through the upregulation of MICU1.

Dataset Information

MICU1 Serves as a Molecular Gatekeeper to Prevent In Vivo Mitochondrial Calcium Overload.

Publications

MICU1 Serves as a Molecular Gatekeeper to Prevent In Vivo Mitochondrial Calcium Overload.

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