Dataset Information

Differential analysis of mitochondria-associated reticulum membranes (MAMs) between standard diet-fed and high fat high sucrose diet fed mice

ABSTRACT: Type 2 diabetic cardiomyopathy (DCM) has been linked to Ca2+ signaling alterations, notably a decreased mitochondrial Ca2+ uptake. Uncovering of Ca2+ microdomains between cardiac mitochondria and reticulum launched a new investigation avenue for cardiometabolic diseases. We here aimed to study if the impairment of mitochondrial Ca2+ handling could be due to a dysregulation of the reticulum-mitochondria interactions or of the mitochondrial Ca2+ uniporter in the diabetic mice heart. Phenotypic alterations of the type 2 diabetic mouse heart, was done using an in vivo obesogenic high fat high sucrose diet fed mouse model (HFHSD: 20% proteins, 36% lipids). The composition of the cardiac MAM fractions between standard diet-fed (SD) mice and HFHSD (HF) mice at 16 weeks was analysed by MS-based quantitative proteomics.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Mus Musculus (mouse)

SUBMITTER: Yohann Couté

LAB HEAD: Virginie Brun

PROVIDER: PXD015280 | Pride | 2020-12-02

REPOSITORIES: Pride

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Reduced reticulum-mitochondria Ca2+ transfer is an early and reversible trigger of mitochondrial dysfunctions in diabetic cardiomyopathy.

Dia Maya M Gomez Ludovic L Thibault Helene H Tessier Nolwenn N Leon Christelle C Chouabe Christophe C Ducreux Sylvie S Gallo-Bona Noelle N Tubbs Emily E Bendridi Nadia N Chanon Stephanie S Leray Aymeric A Belmudes Lucid L Couté Yohann Y Kurdi Mazen M Ovize Michel M Rieusset Jennifer J Paillard Melanie M

Basic research in cardiology 20201130 6

Type 2 diabetic cardiomyopathy features Ca2+ signaling abnormalities, notably an altered mitochondrial Ca2+ handling. We here aimed to study if it might be due to a dysregulation of either the whole Ca2+ homeostasis, the reticulum-mitochondrial Ca2+ coupling, and/or the mitochondrial Ca2+ entry through the uniporter. Following a 16-week high-fat high-sucrose diet (HFHSD), mice developed cardiac insulin resistance, fibrosis, hypertrophy, lipi ...[more]

PMID: 33258101

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Project description:Mitochondrial Ca2+ (mCa2+) plays an important role in regulating cardiac metabolism and physiology. However, its contribution to metabolic and functional defects in the hearts of type 2 diabetic mice (T2D) is not well understood. Ca2+ is imported into mitochondria by the mitochondrial calcium uniporter complex (MCUC) which is a highly selective multimeric Ca2+ ion channel composed of pore-forming, scaffold, and regulatory subunits. Here, we describe new mechanisms involving the MCUC inhibitory subunit MCUb and its role in determining cardiac dysfunction by linking cardiac mitochondrial metabolism and contractile function. Hearts from T2D mice were analyzed 5 months following a single streptozotocin (STZ) injection (75 mg/kg) and high fat diet (HFD) feeding, and compared to control hearts from mice fed normal chow. Transcriptional and proteomic profiling were employed to characterize these hearts. Regulation of MCUb gene expression was investigated using a recombinant mutant inactive Cas9 (dCas9)-based gene promoter pull down approach coupled with mass spectrometry (MS). A dominant negative MCUb (MCUb W246R/V251E=dnMCUb) was engineered. dnMCUb was biochemically characterized in vitro, tested in neonatal cardiac myocytes exposed to culture media mimicking the diabetic milieu, and expressed in type 2 diabetic mice for 1 month. The impact of dnMCUb on the cardiac physiology was then investigated using omics as well as functional approaches aimed to highlight metabolic and contractile features. RNA-seq and proteomics analysis revealed that T2D hearts relied almost exclusively on fatty acid oxidative metabolism. In parallel we found a significant increase in MCUb mRNA and protein in T2D hearts, which was associated with decreased mCa2+ import rates. MS analysis of proteins co-precipitated with dCas9 revealed the presence of NCOR2 in control but not in T2D MCUb gene promoters. Downregulation of NCOR2 in isolated cardiac myocytes from control mice recapitulated the increase in MCUb gene expression. The dnMCUb mutant increased MCUC Ca2+ affinity by improving the architecture of the complex. Furthermore, when dnMCUb was introduced to cardiac myocytes and T2D mice we observed increased mitochondrial glucose oxidation, mitochondrial ATP production, and enhanced cardiac function. These were linked to the complete restoration of phospholamban (PLN) phosphorylation via Protein Kinase A (PKA) as revealed by both MS analysis of the phosphoproteome and studies dedicated on PLN only followed by PKA pharmacological inhibition. We detail a new pathological axis in which T2D induces global metabolic adaptations and inflexibility in part due to impaired mCa2+ transport into the mitochondrial matrix, resulting from MCUb overexpression. Moreover, we describe a mCa2+-dependent regulation of cytosolic Ca2+ (cCa2+) trafficking in which mCa2+ regulates the activity of SERCA2a via PKA-dependent phosphorylation of PLN.

			Action	DRS
	MAMs.mzid.gz	Mzid
	VELOS33906.mgf	Mgf
	VELOS33906.raw	Raw
	VELOS33910.mgf	Mgf
	VELOS33910.raw	Raw

Dataset Information

Differential analysis of mitochondria-associated reticulum membranes (MAMs) between standard diet-fed and high fat high sucrose diet fed mice

Dataset's files

Publications

Reduced reticulum-mitochondria Ca<sup>2+</sup> transfer is an early and reversible trigger of mitochondrial dysfunctions in diabetic cardiomyopathy.

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