Proto-oncogene c-jun and c-fos messenger RNAs increase in the liver of carnitine-deficient juvenile visceral steatosis (jvs) mice

AbstractWe determined the mRNA levels of c-jun and c-fos in the liver of C3H-H-2° jvs mice. Both were higher in jvs mice than in normal mice. The level of c-jun mRNA increased gradually after birth, but in the control mice there was almost no change. In addition, α-fetoprotein and aldolase A mRNA levels were also higher than in normal littermates. These results suggest that the pattern of the gene expression in jvs mice partly resembles the one that occurs in undifferentiated hepatocytes and/or hepatocellular carcinoma

Ca2+ activation kinetics of the two aspartate-glutamate mitochondrial carriers, aralar and citrin: Role in the heart malate-aspartate NADH shuttle

Ca2+ regulation of the Ca2+ binding mitochondrial carriers for aspartate/glutamate (AGCs) is provided by their N-terminal extensions, which face the intermembrane space. The two mammalian AGCs, aralar and citrin, are members of the malate-aspartate NADH shuttle. We report that their N-terminal extensions contain up to four pairs of EF-hand motifs plus a single vestigial EF-hand, and have no known homolog. Aralar and citrin contain one fully canonical EF-hand pair and aralar two additional half-pairs, in which a single EF-hand is predicted to bind Ca2+. Shuttle activity in brain or skeletal muscle mitochondria, which contain aralar as the major AGC, is activated by Ca2+ with S0.5 values of 280-350 nM; higher than those obtained in liver mitochondria (100-150 nM) that contain citrin as the major AGC. We have used aralar- and citrin-deficient mice to study the role of the two isoforms in heart, which expresses both AGCs. The S0.5 for Ca 2+ activation of the shuttle in heart mitochondria is about 300 nM, and it remains essentially unchanged in citrin-deficient mice, although it undergoes a drastic reduction to about 100 nM in aralar-deficient mice. Therefore, aralar and citrin, when expressed as single isoforms in heart, confer differences in Ca2+ activation of shuttle activity, probably associated with their structural differences. In addition, the results reveal that the two AGCs fully account for shuttle activity in mouse heart mitochondria and that no other glutamate transporter can replace the AGCs in this pathwayThis work was supported in part by grants from the Ministerio de Educacio´ n y Ciencia (BFU2005-C02-01, GEN2003-20235-C05-03/NAC), Instituto de Salud Carlos III del Ministerio de Sanidad (PI042457), European Union (LSHM-CT-2006-518153) (to J. S.), by Grant SAF2004-06843-C03 from the Ministerio de Educacio´ n y Ciencia (to P. G.-P.), by an institutional grant from the Fundacio´ n Ramo´ n Areces to the CBMSO, and by Grants-in-Aid for Scientific Research (16390100) from the Japan Society for the Promotion of Science (to K. K.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fac

A Novel Point Mutation Affecting the Tyrosine Kinase Domain of the TRKA Gene in a Family with Congenital Insensitivity to Pain with Anhidrosis

A nerve growth factor receptor encoded by the TRKA gene plays an important part in the formation of autonomic neurons and small sensory neurons in dorsal root ganglia and in signal transduction through its intracytoplasmic tyrosine kinase domain. Recently, three mutations in the tyrosine kinase domain of TRKA have been reported in patients with congenital insensitivity to pain with anhidrosis, which is an autosomal recessive disorder characterized by recurrent fever due to absence of sweating, no reaction to noxious stimuli, self-mutilating behavior, and mental retardation. We examined the TRKA gene in five generations of a large Japanese family with many consanguineous marriages who live in a small remote island of the southern part of Japan. We found a novel point mutation at nucleotide 1825 (A→G transition) resulting in Met-581-Val in the tyrosine kinase domain. Two of the three affected patients were homozygous for this mutation; however, the third affected patient was heterozygous. Further analysis revealed that the third patient was a compound heterozygote with the Met-581-Val mutation in one allele and with a single base C deletion mutation at nucleotide 1726 in exon 14 in the other allele, resulting in a frameshift and premature termination codon

Essential role of aralar in the transduction of small Ca2+ signals to neuronal mitochondria

Aralar, the neuronal Ca2+-binding mitochondrial aspartate-glutamate carrier, has Ca2+ binding domains facing the extramitochondrial space and functions in the malate-aspartate NADH shuttle (MAS). Here we showed that MAS activity in brain mitochondria is stimulated by extramitochondrial Ca2+ with an S0.5 of 324 nM. By employing primary neuronal cultures from control and aralar-deficient mice and NAD(P)H imaging with two-photon excitation microscopy, we showed that lactate utilization involves a substantial transfer of NAD(P)H to mitochondria in control but not aralar-deficient neurons, in agreement with the lack of MAS activity associated with aralar deficiency. The increase in mitochondrial NAD(P)H was greatly potentiated by large [Ca2+]i signals both in control and aralar-deficient neurons, showing that these large signals activate the Ca2+ uniporter and mitochondrial dehydrogenases but not MAS activity. On the other hand, small [Ca2+]i signals potentiate the increase in mitochondrial NAD(P)H only in control but not in aralar-deficient neurons. We concluded that neuronal MAS activity is selectively activated by small Ca2+ signals that fall below the activation range of the Ca2+ uniporter and plays an essential role in mitochondrial Ca2+ signalingThisworkwassupportedinpartbyDireccio´nGeneraldeInvestigacio´ndelMinisterio deCienciayTecnologı´aGrantBMC2002-02072,ComunidaddeMadridGrant08.5/ 0024/2003,FondodeInvestigacionesSanitariasdelMinisteriodeSanidadyConsumo 01/0395(toJ.S.),aninstitutionalgrant fromtheFundacio´nRamo´nArecestothe CentrodeBiologı´aMolecular ‘SeveroOchoa,‘ andbyaGrant-in-aidforScientific Research16390100fromtheJapanSocietyforthePromotionofScience(toK.K.).The costsofpublicationof thisarticleweredefrayedinpartbythepaymentofpage charges.Thisarticlemustthereforebeherebymarked“advertisement”inaccordance with18U.S.C.Section1734solelytoindicatethisfac

Role of aralar, the mitochondrial transporter of aspartate-glutamate, in brain N-acetylaspartate formation and Ca2+ signaling in neuronal mitochondria

"This is the peer reviewed version of the following article: Journal of Neuroscience Research 85.15 (2007): 3359-3366, which has been published in final form at https://doi.org/10.1002/jnr.21299. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions"Aralar, the Ca2+-dependent mitochondrial aspartate-glutamate carrier expressed in brain and skeletal muscle, is a member of the malate-aspartate NADH shuttle. Disrupting the gene for aralar, SLC25a12, in mice has enabled the discovery of two new roles of this carrier. On the one hand, it is required for synthesis of brain aspartate and N-acetylaspartate, a neuron-born metabolite that supplies acetate for myelin lipid synthesis; and on the other, it is essential for the transmission of small Ca2+ signals to mitochondria via an increase in mitochondrial NADHMinisterio de Educación y Ciencia; Contract grant numbers: BFU2005-C02-01 and GEN2003-20235-C05-03/NAC; Contract grant sponsor: Instituto de Salud Carlos III del Ministerio de Sanidad; Contract grant numbers: PI042457; Contract grant sponsor: European Union; Contract grant numbers: LSHM-CT-2006-518153 (to J.S.); Contract grant sponsor: Fundación Ramón Areces (institutional grant to Centro de Biología Molecular Severo Ochoa

AGC1-malate aspartate shuttle activity is critical for dopaminehandling in the nigrostriatal pathway

This is the pre-peer reviewed version of the following article: Llorente-Folch, I. et al. "AGC1-malate aspartate shuttle activity is critical for dopaminehandling in the nigrostriatal pathway". Journal of Neurochemistry 124.3 (2013): 347-362, which has been published in final form at http://dx.doi.org/10.1111/jnc.12096This study was supported by grants from the Ministerio deEducacion y Ciencia BFU2008-04084/BMC (to JS), and Ciencia eInnovacion (SAF2010-16427 to MD), Comunidad de Madrid S-GEN-0269-2006 MITOLAB-CM (to JS), European Union GrantLSHM-CT-2006-518153 (to J.S.), and CureFXS E-Rare. EU/FISPS09102673, Spanish Ministry of Health (PI 082038 to MD),Marato TV3, Jerome Lejeune (JMLM/AC /08-044) to MD, Fundac-ion Medica Mutua Madrile~na (to BP), and by an institutional grantfrom the Fundacion Ramon Areces to the CBMSO. CIBERER is aninitiative of the ISCI

Exogenous aralar/slc25a12 can replace citrin/slc25a13 as malate aspartate shuttle component in liver

The deficiency of CITRIN, the liver mitochondrial aspartate–glutamate carrier (AGC), is the cause of four human clinical phenotypes, neonatal intrahepatic cholestasis caused by CITRIN deficiency (NICCD), silent period, failure to thrive and dyslipidemia caused by CITRIN deficiency (FTTDCD), and citrullinemia type II (CTLN2). Clinical symptoms can be traced back to disruption of the malate-aspartate shuttle due to the lack of citrin. A potential therapy for this condition is the expression of aralar, the AGC present in brain, to replace citrin. To explore this possibility we have first verified that the NADH/NAD+ ratio increases in hepatocytes from citrin(−/−) mice, and then found that exogenous aralar expression reversed the increase in NADH/NAD+ observed in these cells. Liver mitochondria from citrin (−/−) mice expressing liver specific transgenic aralar had a small (~ 4–6 nmoles x mg prot−1 x min−1) but consistent increase in malate aspartate shuttle (MAS) activity over that of citrin(−/−) mice. These results support the functional replacement between AGCs in the liver. To explore the significance of AGC replacement in human therapy we studied the relative levels of citrin and aralar in mouse and human liver through absolute quantification proteomics. We report that mouse liver has relatively high aralar levels (citrin/aralar molar ratio of 7.8), whereas human liver is virtually devoid of aralar (CITRIN/ARALAR ratio of 397). This large difference in endogenous aralar levels partly explains the high residual MAS activity in liver of citrin(−/−) mice and why they fail to recapitulate the human disease, but supports the benefit of increasing aralar expression to improve the redox balance capacity of human liver, as an effective therapy for CITRIN deficienc

Reduced N-acetylaspartate levels in mice lacking aralar, a brain- and muscle-type mitochondrial aspartate-glutamate carrier

Aralar is a mitochondrial calcium-regulated aspartate-glutamate carrier mainly distributed in brain and skeletal muscle, involved in the transport of aspartate from mitochondria to cytosol, and in the transfer of cytosolic reducing equivalents into mitochondria as a member of the malate-aspartate NADH shuttle. In the present study, we describe the characteristics of aralar-deficient (Aralar-/-) mice, generated by a gene-trap method, showing no aralar mRNA and protein, and no detectable malate-aspartate shuttle activity in skeletal muscle and brain mitochondria. Aralar-/- mice were growth-retarded, exhibited generalized tremoring, and had pronounced motor coordination defects along with an impaired myelination in the central nervous system. Analysis of lipid components showed a marked decrease in the myelin lipid galactosyl cerebroside. The content of the myelin lipid precursor, N-acetylaspartate, and that of aspartate are drastically decreased in the brain of Aralar-/- mice. The defect in N-acetylaspartate production was also observed in cell extracts from primary neuronal cultures derived from Aralar-/- mouse embryos. These results show that aralar plays an important role in myelin formation by providing aspartate for the synthesis of N-acetylaspartate in neuronal cell