Secretagogin regulates asynchronous and spontaneous glutamate release in hippocampal neurons through interaction with Doc2α

Abstract

Synaptic vesicle (SV) exocytosis is orchestrated by protein machineries consisting of the SNARE complex, Ca²⁺ sensors, and their partners. Secretagogin (SCGN) is a Ca²⁺-binding protein involved in multiple forms of vesicle secretion. Although SCGN is implicated in multiple neurological disorders, its role in SV exocytosis in neurons remains unknown. Here, using knockout and knockdown techniques, we report that SCGN could regulate the asynchronous and spontaneous forms of excitatory but not inhibitory SV exocytosis in mouse hippocampal neurons. Furthermore, SCGN functioned in glutamate release via directly interacting with Doc2α, a high-affinity Ca²⁺ sensor specific for asynchronous and spontaneous SV exocytosis. Conversely, the interaction with SCGN is also required for Doc2α to execute its Ca²⁺ sensor function in SV release. Together, our study revealed that SCGN plays an important role in asynchronous and spontaneous glutamate release through its interaction with Doc2α.

Keywords

secretagogin / Doc2α / glutamate release / synaptic vesicle / exocytosis

References

[1] Koh TW, Bellen HJ. Synaptotagmin I, a Ca²⁺ sensor for neurotransmitter release. Trends Neurosci 2003;26:413–22.

[2] Sun J, Pang ZP, Qin D, et al. A dual-Ca²⁺-sensor model for neurotransmitter release in a central synapse. Nature 2007;450:676–82.

[3] Kaeser PS, Regehr WG. Molecular mechanisms for synchronous, asynchronous, and spontaneous neurotransmitter release. Annu Rev Physiol 2014;76:333–63.

[4] Lau PM, Bi GQ. Synaptic mechanisms of persistent reverberatory activity in neuronal networks. Proc Natl Acad Sci U S A 2005;102:10333–8.

[5] Iremonger KJ, Bains JS. Integration of asynchronously released quanta prolongs the postsynaptic spike window. J Neurosci 2007;27:6684–91.

[6] Best AR, Regehr WG. Inhibitory regulation of electrically coupled neurons in the inferior olive is mediated by asynchronous release of GABA. Neuron 2009;62:555–65.

[7] Yao J, Gaffaney JD, Kwon SE, et al. Doc2 is a Ca²⁺ sensor required for asynchronous neurotransmitter release. Cell 2011;147:666–77.

[8] Deng S, Li J, He Q, et al. Regulation of recurrent inhibition by asynchronous glutamate release in neocortex. Neuron 2020;105:522–533.e4.

[9] Carter AG, Regehr WG. Quantal events shape cerebellar interneuron firing. Nat Neurosci 2002;5:1309–18.

[10] Sharma G, Vijayaraghavan S. Modulation of presynaptic store calcium induces release of glutamate and postsynaptic firing. Neuron 2003;38:929–39.

[11] Sutton MA, Ito HT, Cressy P, et al. Miniature neurotransmission stabilizes synaptic function via tonic suppression of local dendritic protein synthesis. Cell 2006;125:785–99.

[12] Sutton MA, Taylor AM, Ito HT, et al. Postsynaptic decoding of neural activity: eEF2 as a biochemical sensor coupling miniature synaptic transmission to local protein synthesis. Neuron 2007;55:648–61.

[13] Sollner T, Bennett MK, Whiteheart SW, et al. A protein assembly-disassembly pathway in-vitro that may correspond to sequential steps of synaptic vesicle docking, activation, and fusion. Cell 1993;75:409–18.

[14] Rizo J, Xu J. The synaptic vesicle release machinery. Annu Rev Biophys 2015;44:339–67.

[15] Geppert M, Goda Y, Hammer RE, et al. Synaptotagmin I: a major Ca²⁺ sensor for transmitter release at a central synapse. Cell 1994;79:717–27.

[16] Pang ZP, Melicoff E, Padgett D, et al. Synaptotagmin-2 is essential for survival and contributes to Ca²⁺ triggering of neurotransmitter release in central and neuromuscular synapses. J Neurosci 2006;26:13493–504.

[17] Xu J, Mashimo T, Sudhof TC. Synaptotagmin-1, -2, and -9: Ca(2+) sensors for fast release that specify distinct presynaptic properties in subsets of neurons. Neuron 2007;54:567–81.

[18] Bacaj T, Wu D, Yang X, et al. Synaptotagmin-1 and synaptotagmin-7 trigger synchronous and asynchronous phases of neurotransmitter release. Neuron 2013;80:947–59.

[19] Xu J, Pang ZP, Shin O-H, et al. Synaptotagmin-1 functions as a Ca²⁺ sensor for spontaneous release. Nat Neurosci 2009;12:759–66.

[20] Groffen AJ, Martens S, Díez Arazola R, et al. Doc2b is a high- affinity Ca₂₊ sensor for spontaneous neurotransmitter release. Science 2010;327:1614–8.

[21] Orita S, Sasaki T, Naito A, et al. Doc2: a novel brain protein having two repeated C2-like domains. Biochem Biophys Res Commun 1995;206:439–48.

[22] Sakaguchi G, Orita S, Maeda M, et al. Molecular cloning of an isoform of Doc2 having two C2-like domains. Biochem Biophys Res Commun 1995;217:1053–61.

[23] Sugita S, Han W, Butz S, et al. Synaptotagmin VII as a plasma membrane Ca(2+) sensor in exocytosis. Neuron 2001;30:459–73.

[24] Han W, Rhee J-S, Maximov A, et al. N-glycosylation is essential for vesicular targeting of synaptotagmin 1. Neuron 2004;41:85–99.

[25] Kojima T, Fukuda M, Aruga J, et al. Calcium-dependent phospholipid binding to the C2A domain of a ubiquitous form of double C2 protein (Doc2 beta). J Biochem 1996;120:671–6.

[26] Courtney NA, Briguglio JS, Bradberry MM, et al. Excitatory and inhibitory neurons utilize different Ca(2+) sensors and sources to regulate spontaneous release. Neuron 2018;98:977–991.e5.

[27] Pang ZP, Bacaj T, Yang X, et al. Doc2 supports spontaneous synaptic transmission by a Ca(2+)-independent mechanism. Neuron 2011;70:244–51.

[28] Wagner L, Oliyarnyk O, Gartner W, et al. Cloning and expression of secretagogin, a novel neuroendocrine- and pancreatic islet of Langerhans-specific Ca2+-binding protein. J Biol Chem 2000;275:24740–51.

[29] Rogstam A, Linse S, Lindqvist A, et al. Binding of calcium ions and SNAP-25 to the hexa EF-hand protein secretagogin. Biochem J 2007;401:353–63.

[30] Birkenkamp-Demtroder K, Wagner L, Brandt Sørensen F, et al. Secretagogin is a novel marker for neuroendocrine differentiation. Neuroendocrinology 2005;82:121–38.

[31] Yang SY, Lee J-J, Lee J-H, et al. Secretagogin affects insulin secretion in pancreatic beta-cells by regulating actin dynamics and focal adhesion. Biochem J 2016;473:1791–803.

[32] Malenczyk K, Girach F, Szodorai E, et al. A TRPV1-to-secretagogin regulatory axis controls pancreatic beta-cell survival by modulating protein turnover. EMBO J 2017;36:2107–25.

[33] Mulder J, Zilberter M, Spence L, et al. Secretagogin is a Ca²⁺-binding protein specifying subpopulations of telencephalic neurons. Proc Natl Acad Sci U S A 2009;106:22492–7.

[34] Maj M, Milenkovic I, Bauer J, et al. Novel insights into the distribution and functional aspects of the calcium binding protein secretagogin from studies on rat brain and primary neuronal cell culture. Front Mol Neurosci 2012;5:84.

[35] Krishnamurthy D, Harris LW, Levin Y, et al. Metabolic, hormonal and stress-related molecular changes in post-mortem pituitary glands from schizophrenia subjects. World J Biol Psychiatry 2013;14:478–89.

[36] Deciphering Developmental Disorders, S. Large-scale discovery of novel genetic causes of developmental disorders. Nature 2015;519:223–8.

[37] Romanov RA, Alpár A, Zhang M-D, et al. A secretagogin locus of the mammalian hypothalamus controls stress hormone release. EMBO J 2015;34:36–54.

[38] Alhowikan AM, Ayadhi LA, Halepoto DM. Secretagogin (SCGN) plasma levels and their association with cognitive and social behavior in children with Autism Spectrum Disorder (ASD). J Coll Physicians Surg Pak 2017;27:222–6.

[39] Sarkar A, Mei A, Paquola ACM, et al. Efficient generation of CA3 neurons from human pluripotent stem cells enables modeling of hippocampal connectivity in vitro. Cell Stem Cell 2018;22:684–697.e9.

[40] Zahola P, Hanics J, Pintér A, et al. Secretagogin expression in the vertebrate brainstem with focus on the noradrenergic system and implications for Alzheimer’s disease. Brain Struct Funct 2019;224:2061–78.

[41] Hanics J, Szodorai E, Tortoriello G, et al. Secretagogin-dependent matrix metalloprotease-2 release from neurons regulates neuroblast migration. Proc Natl Acad Sci U S A 2017;114:E2006–15.

[42] Qin J, Liu Q, Liu Z, et al. Structural and mechanistic insights into secretagogin-mediated exocytosis. Proc Natl Acad Sci U S A 2020;117:6559–70.

[43] Wang QW, Qin J, Chen Y-F, et al. 16p112 CNV gene Doc2alpha functions in neurodevelopment and social behaviors through interaction with Secretagogin. Cell Rep 2023;42:112691.

[44] Hagler DJ Jr, Goda Y. Properties of synchronous and asynchronous release during pulse train depression in cultured hippocampal neurons. J Neurophysiol 2001;85:2324–34.

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