Selective transfer of calcium from an acidic compartment to the mitochondrion of Trypanosoma brucei -: Measurements with targeted aequorins

被引:36
作者
Xiong, ZH
Ridgley, EL
Enis, D
Olness, F
Ruben, L [1 ]
机构
[1] So Methodist Univ, Dept Biol Sci, Dallas, TX 75275 USA
[2] So Methodist Univ, Dept Phys, Dallas, TX 75275 USA
关键词
D O I
10.1074/jbc.272.49.31022
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Organelle compartments are used by cells as reservoirs of exchangeable Ca2+ and as Ca2+ buffers, The following study uses recombinant aequorins (CYT-AEQ and MT-AEQ) to measure the dynamics of Ca2+ flux between organelles in procyclic forms of the pathogenic protozoan, Trypanosoma brucei. Emphasis is placed on the exchange between an acidic Ca2+ reservoir and the mitochondrion, The mammalian mitochondrial targeting sequence was functional in trypanosomes as determined by immunoblots, immunolocalizations, and the observation that MT-AEQ was in a compartment whose Ca2+ uptake was inhibited 82% with carbonyl cyanide p-trifluoromethoxyphenylhydrazone and KCN, The resting level of free calcium ion concentration in the mitochondrion ([Ca2+](mit),,,) was slightly higher than that in the cytoplasm ([Ca2+],(cyt)) (400 +/- 50 nM and 290 +/- 40 nM?I, respectively), Melittin (125 nM) disrupted Ca2+ homeostasis by inducing Ca2+ influx across the plasma membrane, [Ca2+](cyt) became slightly elevated to 410 +/- 100 nM, whereas [Ca2+],(mit), was selectively increased approximately la fold, with a broad peak at 4.8 +/- 1.9 mu M. At the peak, the mitochondrion contained approximately three times more free Ca2+ than the cytosol. However, mitochondrial retention of the Ca2+ was transient, Similar selective transport into the mitochondrion was observed when Ca2+ efflux from an acidic compartment was induced with monensin (2 mu g/ml) in the presence of 5 mM EGTA, [Ca2+](cyt) was transiently elevated to 400 +/- 50 nM, whereas [Ca2+],(mit), was elevated to 3.3+/-1.3 mu M. When cells were treated sequentially with monensin (2 mu g/ml) and then melittin (200 nM), mitochondrial Ca2+ transport was normal, However, [Ca2+](cyt),, became elevated to a level that was 1.4-fold higher than with melittin alone, Overall, these data demonstrate that the trypanosome mitochondrion is not a reservoir of exchangeable Ca2+ in the resting cell, However, Ca2+ is selectively channeled to the mitochondrion from the plasma membrane or acidic Ca2+ storage compartment, Additionally, the acidic compartment contributes to maintenance of Ca2+ homeostasis in response to melittin.
引用
收藏
页码:31022 / 31028
页数:7
相关论文
共 48 条
  • [1] Differential regulation of nuclear and cytosolic Ca2+ in HeLa cells
    Badminton, MN
    Campbell, AK
    Rembold, CM
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 1996, 271 (49) : 31210 - 31214
  • [2] NUCLEOPLASMIN-TARGETED AEQUORIN PROVIDES EVIDENCE FOR A NUCLEAR CALCIUM BARRIER
    BADMINTON, MN
    KENDALL, JM
    SALANEWBY, G
    CAMPBELL, AK
    [J]. EXPERIMENTAL CELL RESEARCH, 1995, 216 (01) : 236 - 243
  • [4] A CALMODULIN-STIMULATED CA2+ PUMP IN PLASMA-MEMBRANE VESICLES FROM TRYPANOSOMA-BRUCEI - SELECTIVE-INHIBITION BY PENTAMIDINE
    BENAIM, G
    LOPEZESTRANO, C
    DOCAMPO, R
    MORENO, SNJ
    [J]. BIOCHEMICAL JOURNAL, 1993, 296 : 759 - 763
  • [5] MITOCHONDRIAL DEVELOPMENT IN TRYPANOSOMA-BRUCEI-BRUCEI TRANSITIONAL BLOOD-STREAM FORMS
    BIENEN, EJ
    SARIC, M
    POLLAKIS, G
    GRADY, RW
    CLARKSON, AB
    [J]. MOLECULAR AND BIOCHEMICAL PARASITOLOGY, 1991, 45 (02) : 185 - 192
  • [6] TRANSFECTED AEQUORIN IN THE MEASUREMENT OF CYTOSOLIC CA2+ CONCENTRATION ([CA2+](C)) - A CRITICAL-EVALUATION
    BRINI, M
    MARSAULT, R
    BASTIANUTTO, C
    ALVAREZ, J
    POZZAN, T
    RIZZUTO, R
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 1995, 270 (17) : 9896 - 9903
  • [7] NUCLEAR CA2+ CONCENTRATION MEASURED WITH SPECIFICALLY TARGETED RECOMBINANT AEQUORIN
    BRINI, M
    MURGIA, M
    PASTI, L
    PICARD, D
    POZZAN, T
    RIZZUTO, R
    [J]. EMBO JOURNAL, 1993, 12 (12) : 4813 - 4819
  • [8] BRUN R, 1979, ACTA TROP, V36, P289
  • [9] CLARKSON AB, 1989, J BIOL CHEM, V264, P17770
  • [10] CA-2+ AS A 2ND MESSENGER WITHIN MITOCHONDRIA OF THE HEART AND OTHER TISSUES
    DENTON, RM
    MCCORMACK, JG
    [J]. ANNUAL REVIEW OF PHYSIOLOGY, 1990, 52 : 451 - 466