Kek-6: A truncated-Trk-like receptor for Drosophila neurotrophin 2 regulates structural synaptic plasticity

Abstract

Neurotrophism, structural plasticity, learning and long-term memory in mammals critically depend on neurotrophins binding Trk receptors to activate tyrosine kinase (TyrK) signaling, but Drosophila lacks full-length Trks, raising the question of how these processes occur in the fly. Paradoxically, truncated Trk isoforms lacking the TyrK predominate in the adult human brain, but whether they have neuronal functions independently of full-length Trks is unknown. Drosophila has TyrK-less Trk-family receptors, encoded by the kekkon (kek) genes, suggesting that evolutionarily conserved functions for this receptor class may exist. Here, we asked whether Keks function together with Drosophila neurotrophins (DNTs) at the larval glutamatergic neuromuscular junction (NMJ). We tested the eleven LRR and Ig-containing (LIG) proteins encoded in the Drosophila genome for expression in the central nervous system (CNS) and potential interaction with DNTs. Kek-6 is expressed in the CNS, interacts genetically with DNTs and can bind DNT2 in signaling assays and co-immunoprecipitations. Ligand binding is promiscuous, as Kek-6 can also bind DNT1, and Kek-2 and Kek-5 can also bind DNT2. In vivo, Kek-6 is found presynaptically in motoneurons, and DNT2 is produced by the muscle to function as a retrograde factor at the NMJ. Kek-6 and DNT2 regulate NMJ growth and synaptic structure. Evidence indicates that Kek-6 does not antagonise the alternative DNT2 receptor Toll-6. Instead, Kek-6 and Toll-6 interact physically, and together regulate structural synaptic plasticity and homeostasis. Using pull-down assays, we identified and validated CaMKII and VAP33A as intracellular partners of Kek-6, and show that they regulate NMJ growth and active zone formation downstream of DNT2 and Kek-6. The synaptic functions of Kek-6 could be evolutionarily conserved. This raises the intriguing possibility that a novel mechanism of structural synaptic plasticity involving truncated Trk-family receptors independently of TyrK signaling may also operate in the human brain.

Toll and Toll-like receptor signalling in development

“…Toll/TLRS also activate the MAPK pathways upstream of ERK and JNK signalling, and the latter can be activated by Sarm (Chuang and Bargmann, 2005;Foldi et al, 2017;Gay and Gangloff, 2007;Wu et al, 2015). In addition, alternative signalling pathways are being discovered, for instance those involving FoxO or cell-to-cell communication (Chuang and Bargmann, 2005;Foldi et al, 2017;McLaughlin et al, 2016;Paré et al, 2014;Ulian-Benitez et al, 2017;Ward et al, 2015), expanding the potential mechanisms of Toll/TLR function.…”

“…Tolls also regulate structural synaptic plasticity, both under normal conditions and in response to increased neuronal activity, at the Drosophila glutamatergic neuromuscular junction (NMJ). Toll-6 and Toll-8 mutant larvae have smaller NMJs, with fewer synapses and, consequently, larvae exhibit a slow crawling behaviour (Ballard et al, 2014;McIlroy et al, 2013;Ulian-Benitez et al, 2017). Genetic evidence indicates that Toll-8 promotes NMJ growth independently of NF-κB signalling and via JNK instead (Ballard et al, 2014).…”

“…In Drosophila, NMJ growth also depends on the retrograde factor DNT2 produced from the muscle, which binds a receptor complex formed of Toll-6 and Kek6, whereby Kek6 causes the activation of CaMKII (Ulian-Benitez et al, 2017). However, the mechanism might differ from that of C. elegans, as Toll-6 and Kek6 function presynaptically.…”

Synaptic Properties and Plasticity Mechanisms of Invertebrate Tonic and Phasic Neurons

“…These include pathways that result in functional increases in presynaptic output due to increased SV pools, increased presynaptic Ca 2+ influx, or enhanced membrane excitability. Numerous pathways that control structural plasticity and regulation of AZ number at Drosophila NMJs have also been identified, including Neurexin/Neuroligin, Teneurins, neurotrophins, Synaptotagmin 4-mediated retrograde signaling, BMPs, Wingless, and regulated proteolysis ( Wan et al, 2000 ; Aberle et al, 2002 ; Marqués et al, 2002 ; DiAntonio and Hicke, 2004 ; Yoshihara et al, 2005 ; Ataman et al, 2006 , 2008 ; Collins et al, 2006 ; Collins and DiAntonio, 2007 ; Li et al, 2007 ; Johnson et al, 2009 ; Wairkar et al, 2009 ; Banovic et al, 2010 ; Sun et al, 2011 ; Miller et al, 2012 ; Mosca et al, 2012 ; Owald et al, 2012 ; Berke et al, 2013 ; Korkut et al, 2013 ; Piccioli and Littleton, 2014 ; Harris and Littleton, 2015 ; Muhammad et al, 2015 ; Harris et al, 2016 ; Banerjee et al, 2017 ; Ulian-Benitez et al, 2017 ). Defining if and how these well-known molecular pathways for synaptic growth and function are uniquely employed in tonic vs. phasic motoneurons should complement RNA profiling approaches and help decipher how differences in synaptic structure and function arise.…”

The Toll Route to Structural Brain Plasticity

“…Drosophila neurotrophins are encoded by the spätzle ( spz ) paralog gene family, which includes Drosophila neurotrophin-1 (DNT1, also called spz-2), DNT2/spz-5, and Spz-1 ( Zhu et al, 2008 ; Sutcliffe et al, 2013 ; Foldi et al, 2017 ). During nervous system development, DNTs promote neuronal survival, cell death, connectivity, structural synaptic plasticity, and compensatory plasticity ( Zhu et al, 2008 ; Sutcliffe et al, 2013 ; Foldi et al, 2017 ; Ulian-Benitez et al, 2017 ). Mammalian neurotrophins bind Trk and p75 receptors, but DNTs bind kinase-free Trk homolog encoded by the kekkon genes and Toll receptors ( Mandai et al, 2009 ; Ulian-Benitez et al, 2017 ).…”