Crimpy Sorts a BMP into the Regulated Secretory Pathway for Activity-Dependent Release in Drosophila Motorneurons

Neural circuits integrate experience and store information by the formation and remodeling of synapses. Defining the signaling pathways that underlie such plasticity has been a major goal of modern neuroscience. BMP signaling critically regulates both morphological plasticity and neurotransmission in motorneurons at the Drosophila neuromuscular junction (NMJ). However the mechanisms that establish BMP pathway directionality and diversify pathway action in motorneurons remain elusive. In this thesis, I describe a novel regulator of the BMP signaling pathway in Drosophila motorneurons that I name Crimpy (Cmpy). Cmpy acts at the interface between morphological plasticity and synaptic transmission to specify a role for motorneuronal BMP ligand in promoting synaptic transmission, as opposed to morphological growth, at the NMJ.Using the Drosophila NMJ as a model system, I examined the role of Cmpy in motorneuron development. BMP signaling scales growth of the presynaptic axon arbor to growth of the postsynaptic muscle during larval development in order to preserve synaptic input onto the muscle cell. Genetic analyses suggest that the relevant BMP ligand at the NMJ, Glass bottom boat (Gbb), acts in a retrograde fashion, secreted from the postsynaptic muscle to activate presynaptic BMP receptors and promote motoraxon growth. However establishment of this directionality is unclear since Gbb is also produced by and active within motorneurons, albeit to regulate synaptic transmission as opposed to morphological growth. I found that loss of cmpy results in excessive presynaptic growth due to ectopic motorneuron-derived, autocrine pro-growth BMP signaling at the NMJ. Biochemical analyses demonstrated that Cmpy binds to precursor BMP. Expressing a novel Gbb-HA transgene within larval motorneurons that preserves protein processing and secretion, and functions like a wild-type Gbb transgene, I detected presynaptic Gbb-HA localization at the NMJ. Cmpy is necessary and sufficient for Gbb-HA localization at the NMJ. Gbb-HA and Cmpy-Venus transgenes colocalize at presynaptic terminals and also within motor nerves, where they are observed in discrete puncta reminiscent of trafficking vesicles. Furthermore, Gbb-HA and Cmpy-Venus both colocalize with a marker for dense core vesicles (DCVs) of the regulated secretory pathway (RSP) at the NMJ and in individual puncta within motor nerves, which indicated that Gbb is secreted from DCVs in response to synaptic activity. Indeed, by stimulating synaptic activity in motorneurons using two independent measures, high K+ depolarization and by activating the blue light-gated cation channel, channelrhodopsin, in motorneurons, we detect activity-dependent changes in Gbb-HA levels at the larval NMJ. Growth factors of the neurotrophin and TGFß superfamilies are secreted from vertebrate neurons in response to synaptic activity in addition to constitutive secretion. Sorting receptors for the neurotrophin BDNF have been identified that interact with precursor BDNF and direct its delivery into the RSP. This reminded us of interactions between Cmpy and precursor Gbb. Additionally, activity-dependent growth factor secretion promotes synaptic transmission at vertebrate synapses. Similarly, neuronal Gbb is critical for proper neurotransmission at the Drosophila NMJ. We hypothesized that Cmpy is a sorting receptor that delivers Gbb into the RSP for activity-dependent secretion in larval motorneurons, and that activity-dependent Gbb signaling strengthens synaptic transmission. In line with this hypothesis, I found that Cmpy is necessary for activity-dependent changes in Gbb-HA levels at the NMJ. Also, evoked synaptic transmission is impaired at cmpy mutant NMJs, despite overgrowth of cmpy mutant terminals. Taken together, my thesis work identifies a novel pathway that diversifies BMP pathway action in larval motorneurons by sorting the BMP ligand into DCVs of the RSP for activity-dependent secretion. We propose that sorting of Gbb into the RSP in Drosophila motorneurons defines its role in synaptic transmission, as opposed to growth, at the NMJ. To date, Cmpy is the first BMP sorting receptor identified in any species. Given the high degree of conservation of key developmental signaling pathways between vertebrates and invertebrates, it is likely that vertebrate homologs of Cmpy similarly sort TGFß ligands into the RSP for activity-dependent secretion from vertebrate neurons.

Name thesis:degree_name

Doctor of Philosophy

Level thesis:degree_level

doctoral

Discipline thesis:degree_discipline

Neurosciences

Grantor dc:publisher

Case Western Reserve University School of Graduate Studies

Year dc:date

2013

Author dc:creator

• James, Rebecca E.

Contributors dc:contributor

• Strowbridge, Ben
• Broihier, Heather

• Neurosciences
• Developmental Biology
• Drosophila melanogaster
• motorneuron
• neuromuscular junction
• Glass bottom boat
• Crimpy
• activity-dependent BMP secretion
• BMP sorting receptor

Statement dc:rights

• unrestricted
• This thesis or dissertation is protected by copyright: all rights reserved. It may not be copied or redistributed beyond the terms of applicable copyright laws.

Language dc:language

English

Repository record dc:identifier

http://rave.ohiolink.edu/etdc/view?acc_num=case1364994680

OAI identifier oai:identifier

oai:etd.ohiolink.edu:case1364994680