Ca2+-independent insulin exocytosis induced by alpha -latrotoxin requires latrophilin, a G protein-coupled receptor

Lang, Jochen; Ushkaryov, Yuri A.; Grasso, A.; Wollheim, Claes B.

doi:10.1093/emboj/17.3.648

Cited by 78 publications

(86 citation statements)

References 67 publications

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“…NRXs) may be responsible for mediating this LTX action. Although this hypothesis contradicted our previous results (5,26,27), we decided to test it using the fact that Sr 2ϩ does not support the interaction of LTX with NRX I␣ but substitutes Ca 2ϩ in LTX-evoked exocytosis (5).The objectives of the current study were (i) to reveal the …”

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confidence: 72%

Mutant α-Latrotoxin (LTXN4C) Does Not Form Pores and Causes Secretion by Receptor Stimulation

Volynski

Capogna

Ashton

et al. 2003

Journal of Biological Chemistry

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␣-Latrotoxin (LTX) causes massive release of neurotransmitters via a complex mechanism involving (i) activation of receptor(s) and (ii) toxin insertion into the plasma membrane with (iii) subsequent pore formation. Using cryo-electron microscopy, electrophysiological and biochemical methods, we demonstrate here that the recently described toxin mutant (LTX N4C ) is unable to insert into membranes and form pores due to its inability to assemble into tetramers. However, this mutant still binds to major LTX receptors (latrophilin and neurexin) and causes strong transmitter exocytosis in synaptosomes, hippocampal slice cultures, neuromuscular junctions, and chromaffin cells. In the absence of mutant incorporation into the membrane, receptor activation must be the only mechanism by which LTX N4C triggers exocytosis. An interesting feature of this receptormediated transmitter release is its dependence on extracellular Ca 2؉ . Because Ca 2؉ is also strictly required for LTX interaction with neurexin, the latter might be the only receptor mediating the LTX N4C action. To test this hypothesis, we used conditions (substitution of Ca 2؉ in the medium with Sr 2؉ ) under which LTX N4C does not bind to any member of the neurexin family but still interacts with latrophilin. We show that, in all the systems tested, Sr 2؉ fully replaces Ca 2؉ in supporting the stimulatory effect of LTX N4C . These results indicate that LTX N4C can cause neurotransmitter release just by stimulating a receptor and that neurexins are not critical for this receptor-mediated action.␣-Latrotoxin (LTX) 1 stimulates exhaustive release of neurotransmitters in vertebrates. This toxin has been extensively used to probe molecular mechanisms that control exocytosis of both synaptic vesicles (SVs) and large dense-core vesicles (LDCVs) in such diverse models as brain, neuromuscular junctions, and endocrine cells (for reviews see Refs. 1-3).LTX acts only after binding to presynaptic receptors (4). Once receptor-bound, the toxin can trigger exocytosis by several mechanisms: (i) activation of the receptors (5-8), (ii) formation of non-selective pores in the membrane (9 -11), and (iii) hypothetical intracellular interaction with the exocytotic machinery (12).Because toxin pores damage cell membranes and produce strong cytotoxic effects (e.g. 13-15), only the receptor-transduced LTX action is likely to reveal intact, physiologically important exocytotic mechanisms. Unfortunately, this action is difficult to study using the wild-type LTX (LTX WT ) because it easily inserts into membranes and forms ionic pores (9 -11, 16). This problem could be overcome by designing LTX mutants that would lack the propensity of membrane insertion, and a promising toxin variant has been described recently (LTX N4C ) (17). This mutant had the same affinity for the receptors as LTX WT (17) but failed to form pores in synaptosomes or receptor-transfected BHK cells (8). Lacking the major (ionophore) activity, LTX N4C was originally thought to be altogether inactive (12, 17). However, we h...

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confidence: 72%

Mutant α-Latrotoxin (LTXN4C) Does Not Form Pores and Causes Secretion by Receptor Stimulation

Volynski

Capogna

Ashton

et al. 2003

Journal of Biological Chemistry

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“…However, the toxin is also active when applied in nominally Ca 2ϩ -free buffers (7,8,18) or when cation fluxes are controlled, e.g. in permeabilized cells (14,19). A possible explanation of the calcium-independent effect of ␣-latrotoxin is that it activates its membrane receptors, which results in intracellular signaling, through a G protein-linked pathway (19,20).…”

Section: Discussionmentioning

confidence: 99%

“…in permeabilized cells (14,19). A possible explanation of the calcium-independent effect of ␣-latrotoxin is that it activates its membrane receptors, which results in intracellular signaling, through a G protein-linked pathway (19,20).…”

Section: Discussionmentioning

confidence: 99%

Structural Requirements for α-Latrotoxin Binding and α-Latrotoxin-stimulated Secretion

Krasnoperov¹,

Bittner²,

Holz³

et al. 1999

Journal of Biological Chemistry

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Stimulation of neurotransmitter release by ␣-latrotoxin requires its binding to the calcium-independent receptor of ␣-latrotoxin (CIRL), an orphan neuronal G protein-coupled receptor. CIRL consists of two noncovalently bound subunits, p85, a heptahelical integral membrane protein, and p120, a large extracellular polypeptide with domains homologous to lectin, olfactomedin, mucin, the secretin receptor family, and a novel structural motif common for large orphan G proteincoupled receptors. The analysis of CIRL deletion mutants indicates that the high affinity ␣-latrotoxin-binding site is located within residues 467-891, which comprise the first transmembrane segment of p85 and the C-terminal half of p120. The N-terminal lectin, olfactomedin, and mucin domains of p120 are not required for the interaction with ␣-latrotoxin. Soluble p120 and all its fragments, which include the 467-770 residues, bind ␣-latrotoxin with low affinity suggesting the importance of membrane-embedded p85 for the stabilization of the complex of the toxin with p120. Two COOH-terminal deletion mutants of CIRL, one with the truncated cytoplasmic domain and the other with only one transmembrane segment left of seven, supported both ␣-latrotoxin-induced calcium uptake in HEK293 cells and ␣-latrotoxin-stimulated secretion when expressed in chromaffin cells, although with a different dose dependence than wild-type CIRL and its N-terminal deletion mutant. Thus the signaling domains of CIRL are not critically important for the stimulation of exocytosis in intact chromaffin cells by ␣-latrotoxin.

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“…ST6GALNAC3 is an integral Golgi membrane protein which catalyzes the transfer of sialic acids to carbohydrate groups on glycoproteins and glycolipids. LPHN2 is a G-protein-coupled receptor related to the receptor that binds black widow (Latrodectus) spider venom in synaptic membranes (21).…”

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confidence: 99%

Genome-wide association identifies candidate genes that influence the human electroencephalogram

Hodgkinson

Enoch

Srivastava

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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Complex psychiatric disorders are resistant to whole-genome analysis due to genetic and etiological heterogeneity. Variation in resting electroencephalogram (EEG) is associated with common, complex psychiatric diseases including alcoholism, schizophrenia, and anxiety disorders, although not diagnostic for any of them. EEG traits for an individual are stable, variable between individuals, and moderately to highly heritable. Such intermediate phenotypes appear to be closer to underlying molecular processes than are clinical symptoms, and represent an alternative approach for the identification of genetic variation that underlies complex psychiatric disorders. We performed a whole-genome association study on alpha (α), beta (β), and theta (θ) EEG power in a Native American cohort of 322 individuals to take advantage of the genetic and environmental homogeneity of this population isolate. We identified three genes (SGIP1, ST6GALNAC3, and UGDH) with nominal association to variability of θ or α power. SGIP1 was estimated to account for 8.8% of variance in θ power, and this association was replicated in US Caucasians, where it accounted for 3.5% of the variance. Bayesian analysis of prior probability of association based upon earlier linkage to chromosome 1 and enrichment for vesicle-related transport proteins indicates that the association of SGIP1 with θ power is genuine. We also found association of SGIP1 with alcoholism, an effect that may be mediated via the same brain mechanisms accessed by θ EEG, and which also provides validation of the use of EEG as an endophenotype for alcoholism.alcoholism | electroencephalogram | endophenotype | genetics | whole-genome association G enetic studies of behavior and psychiatric disorders are hampered by etiologic heterogeneity of these complex phenotypes. Addiction vulnerability arises from both internalizing (emotional) and externalizing (dyscontrol) behavioral dimensions (1), and both of these broad aspects of behavior are strongly influenced by early life trauma and other gene/environment interactions (2). Etiologic heterogeneity dilutes power to detect genetic effects, and is a reason for failures to detect and replicate genome-wide associations (GWAS) in complex disorders. Increasing sample size does not remove underlying heterogeneity and can introduce additional confounds. In neuropsychiatry, these considerations have led to the use of intermediate phenotypes (or endophenotypes) that are heritable, relevant to disease, and have good measurement properties and assay variation more closely related to gene function (3) as surrogates to probe the underlying biology of complex disorders.

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Ca2+-independent insulin exocytosis induced by alpha -latrotoxin requires latrophilin, a G protein-coupled receptor

Cited by 78 publications

References 67 publications

Mutant α-Latrotoxin (LTXN4C) Does Not Form Pores and Causes Secretion by Receptor Stimulation

Mutant α-Latrotoxin (LTXN4C) Does Not Form Pores and Causes Secretion by Receptor Stimulation

Structural Requirements for α-Latrotoxin Binding and α-Latrotoxin-stimulated Secretion

Genome-wide association identifies candidate genes that influence the human electroencephalogram

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