Genetic analysis of alpha-latrotoxin receptors reveals functional interdependence of CIRL/latrophilin 1 and neurexin 1 alpha.
alpha-Latrotoxin triggers massive neurotransmitter release from nerve terminals by binding to at least two distinct presynaptic receptors, neurexin 1 alpha and CIRL1/latrophilin1 (CL1). We have now generated knockout (KO) mice that lack CL1 and analyzed them alone or in combination with neurexin 1 alpha KO mice. Mice lacking only CL1, or both CL1 and neurexin 1 alpha, were viable and fertile. Ca(2+)-independent binding of alpha-latrotoxin to brain membranes was impaired similarly in CL1 single and in CL1/neurexin 1 alpha double KO mice (approximately 75% decrease) but not in neurexin 1 alpha single KO mice. In contrast, Ca(2+)-dependent binding (approximately 2 times above Ca(2+)-independent binding) was altered in both CL1 (approximately 50% decrease) and neurexin 1 alpha single KO mice (approximately 25% decrease) and was decreased further in double KO mice (approximately 75% decrease). Synaptosomes lacking CL1 exhibited the same decrease in alpha-latrotoxin-stimulated glutamate release in the presence and absence of Ca(2+) (approximately 75%). In contrast, synaptosomes lacking neurexin 1 alpha exhibited only a small decrease in alpha-latrotoxin-triggered release in the absence of Ca(2+) (approximately 20%) but a major decrease in the presence of Ca(2+) (approximately 75%). Surprisingly, synaptosomes lacking both CL1 and neurexin 1 alpha displayed a relatively smaller decrease in alpha-latrotoxin-stimulated glutamate release than synaptosomes lacking only CL1 in the absence of Ca(2+) (approximately 50 versus approximately 75%), but the same decrease in the presence of Ca(2+) (approximately 75%). Our data suggest the following two major conclusions. 1) CL1 and neurexin 1 alpha together account for the majority (75%) of alpha-latrotoxin receptors in brain, with the remaining receptor activity possibly due to other CL and neurexin isoforms, and 2) the two receptors act additively in binding alpha-latrotoxin but not in triggering release. Together these data suggest that the two receptors act autonomously in binding of alpha-latrotoxin but cooperatively in transducing the stimulation of neurotransmitter release by alpha-latrotoxin.
Tobaben
Sönke
S
Max-Planck-Institute for Experimental Medicine, 37075 Göttingen, Germany.
Südhof
Thomas C
TC
Stahl
Bernd
B
D000818:Animals; D001665:Binding Sites; D004533:Egtazic Acid / Q000494:pharmacology; D015698:Genomic Library; D018698:Glutamic Acid / Q000378:metabolism; D006023:Glycoproteins; D007700:Kinetics; D051379:Mice; D018345:Mice, Knockout; D009419:Nerve Tissue Proteins / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D009479:Neuropeptides; D016133:Polymerase Chain Reaction; D011188:Potassium / Q000494:pharmacology; D018000:Receptors, Peptide / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D015183:Restriction Mapping; D013111:Spider Venoms / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D013574:Synaptosomes / Q000187:drug effects / Q000378:metabolism
D006023:Glycoproteins; D009419:Nerve Tissue Proteins; D009479:Neuropeptides; D018000:Receptors, Peptide; D013111:Spider Venoms; C034800:alpha-latrotoxin receptor; C112162:neurexophilin; D018698:Glutamic Acid; D004533:Egtazic Acid; C020763:alpha-latrotoxin; D011188:Potassium