α-latrotoxin action probed with recombinant toxin:: receptors recruit α-latrotoxin but do not transduce an exocytotic signal

被引:68
作者
Ichtchenko, K
Khvotchev, M
Kiyatkin, N
Simpson, L
Sugita, S
Südhof, TC
机构
[1] Univ Texas, SW Med Ctr, Ctr Basic Neurosci, Dallas, TX 75235 USA
[2] Univ Texas, SW Med Ctr, Howard Hughes Med Inst, Dept Mol Genet, Dallas, TX 75235 USA
[3] Thomas Jefferson Univ, Dept Med, Jefferson Clin Ctr Occupat & Environm Med, Philadelphia, PA 19107 USA
关键词
alpha-latrotoxin; CIRL; latrophilin; neurexins; neurotransmitter release;
D O I
10.1093/emboj/17.21.6188
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
alpha-Latrotoxin stimulates neurotransmitter release probably by binding to two receptors, CIRL/latrophilin 1 (CL1) and neurexin I alpha, We have now produced recombinant alpha-latrotoxin (Ltx(WT)) that is as active as native alpha-latrotoxin in triggering synaptic release of glutamate, GABA and norepinephrine, We have also generated three alpha-latrotoxin mutants with substitutions in conserved cysteine residues, and a fourth mutant with a four-residue insertion. All four alpha-latrotoxin mutants were found to be unable to trigger release. interestingly, the insertion mutant Ltx(N4C) exhibited receptor-binding affinities identical to wild-type Ltx(WT), bound to CL1 and neurexin I alpha as well as Ltx(WT), and similarly stimulated synaptic hydrolysis of phosphatidylinositolphosphates. Therefore, receptor binding by alpha-latrotoxin and stimulation of phospholipase C are insufficient to trigger exocytosis, This conclusion was confirmed in experiments with La3+ and Cd2+. La3+ blocked release triggered by Ltx(WT), whereas Cd2+ enhanced it, Both cations, however, had no effect on the stimulation by Ltx(WT) of phosphatidylinositolphosphate hydrolysis, Our data show that receptor binding by alpha-latrotoxin and activation of phospholipase C do not by themselves trigger exocytosis, Thus receptors recruit alpha-latrotoxin to its point of action without activating exocytosis, Exocytosis probably requires an additional receptor-independent activity of alpha-latrotoxin that is selectively inhibited by the Ltx(N4C) mutation and by La3+.
引用
收藏
页码:6188 / 6199
页数:12
相关论文
共 34 条
  • [1] CAPOGNA M, 1996, J NEUROPHYSIOL, V76, P149
  • [2] CA2+-DEPENDENT RECYCLING OF SYNAPTIC VESICLES AT THE FROG NEUROMUSCULAR-JUNCTION
    CECCARELLI, B
    HURLBUT, WP
    [J]. JOURNAL OF CELL BIOLOGY, 1980, 87 (01) : 297 - 303
  • [3] Vesicle exocytosis stimulated by α-latrotoxin is mediated by latrophilin and requires both external and stored Ca2+
    Davletov, BA
    Meunier, FA
    Ashton, AC
    Matsushita, H
    Hirst, WD
    Lelianova, VG
    Wilkin, GP
    Dolly, JO
    Ushkaryov, YA
    [J]. EMBO JOURNAL, 1998, 17 (14) : 3909 - 3920
  • [4] HIGH-AFFINITY BINDING OF ALPHA-LATROTOXIN TO RECOMBINANT NEUREXIN I-ALPHA
    DAVLETOV, BA
    KRASNOPEROV, V
    HATA, Y
    PETRENKO, AG
    SUDHOF, TC
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 1995, 270 (41) : 23903 - 23905
  • [5] Isolation and biochemical characterization of a Ca2+-independent alpha-latrotoxin-binding protein
    Davletov, BA
    Shamotienko, OG
    Lelianova, VG
    Grishin, EV
    Ushkaryov, YA
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 1996, 271 (38) : 23239 - 23245
  • [6] Phosphoinositides as regulators in membrane traffic
    DeCamilli, P
    Emr, SD
    McPherson, PS
    Novick, P
    [J]. SCIENCE, 1996, 271 (5255) : 1533 - 1539
  • [7] Cloning and structure of delta-latroinsectotoxin, a novel insect-specific member of the latrotoxin family - Functional expression requires C-terminal truncation
    Dulubova, IE
    Krasnoperov, VG
    Khvotchev, MV
    Pluzhnikov, KA
    Volkova, TM
    Grishin, EV
    Vais, H
    Bell, DR
    Usherwood, PNR
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 1996, 271 (13) : 7535 - 7543
  • [8] BLACK-WIDOW SPIDER VENOM - EFFECT OF PURIFIED TOXIN ON LIPID BILAYER MEMBRANES
    FINKELSTEIN, A
    RUBIN, LL
    TZENG, MC
    [J]. SCIENCE, 1976, 193 (4257) : 1009 - 1011
  • [9] PURIFICATION FROM BLACK-WIDOW SPIDER VENOM OF A PROTEIN FACTOR CAUSING DEPLETION OF SYNAPTIC VESICLES AT NEUROMUSCULAR-JUNCTIONS
    FRONTALI, N
    CECCARELLI, B
    GORIO, A
    MAURO, A
    SIEKEVITZ, P
    TZENG, MC
    HURLBUT, WP
    [J]. JOURNAL OF CELL BIOLOGY, 1976, 68 (03) : 462 - 479
  • [10] SEPARATION OF 3 TOXICOLOGICALLY DIFFERENT PROTEIN COMPONENTS FROM VENOM OF SPIDER LATRODECTUS TREDECIMGUTTATUS
    FRONTALI, N
    GRASSO, A
    [J]. ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS, 1964, 106 (1-3) : 213 - &