Neurexin directs partner-specific synaptic connectivity in C. elegans
- University of Massachusetts Medical School, United States
- Assumption College, United States
Figures
Figure 1 with 2 supplements
Characterization of post-synaptic specializations in the DD neurons.
(A) Diagrams of C. elegans showing the six DD GABAergic neurons in the ventral nerve cord and expanded view of DD1. After the L1/L2 transition, the DD neurons exclusively make dorsal synaptic …
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Figure 1—source data 1
Raw values for synaptic vs asynaptic ACR-12::GFP receptor clusters.
- https://doi.org/10.7554/eLife.35692.006
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Figure 1—source data 2
Raw values for wild type spine length, numbers of spines and ACR-12 receptor clusters throughout development.
Relates to Figure 1—figure supplements 1 and 2.
- https://doi.org/10.7554/eLife.35692.007
Figure 1—figure supplement 1
Spine-like protrusions are located on DD dendrites.
(A) Distribution of dendritic protrusion lengths. 65 protrusions from 11 L4 animals expressing flp-13::mCherry are included in the analysis. (B) DD1 and surrounding ventral cord region in an animal …
Figure 1—figure supplement 2
Spine-like protrusions increase developmentally in a correlated manner with ACR-12 receptor clusters.
Quantification of the number of DD1 spine-like protrusions (red) or receptor clusters (green) at the indicated time points after hatch. Protrusions and receptor clusters from ≥10 animals were …
Figure 2 with 1 supplement
Heteromeric ACR-12 receptors are localized at the cell surface opposite sites of release.
(A) Confocal images of presynaptic (acr-2::SNB-1::GFP) and postsynaptic (flp-13::mCherry) domains in the DD1 synaptic region. Note violet/green coloring to more clearly depict presynaptic structures …
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Figure 2—source data 1
Raw values for UNC-29::GFP and UNC-63::GFP receptor cluster number.
- https://doi.org/10.7554/eLife.35692.010
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Figure 2—source data 2
Raw values for ACR-12::GFP receptor cluster number and normalized UNC-29::GFP and UNC-63::GFP fluorescence intensity.
Relates to Figure 2—figure supplement 1.
- https://doi.org/10.7554/eLife.35692.011
Figure 2—figure supplement 1
Mutations in specific AChR subunits and accessory proteins disrupt ACR-12 synaptic delivery and clustering.
(A) Confocal images of the DD1 cell body and synaptic region for a wild type animal expressing flp-13::ACR-12::GFP::3xHA. Animals were injected with anti-GFP antibodies conjugated to Alexa594 …
Figure 3 with 4 supplements
nrx-1/neurexin is required for AChR localization in GABAergic motor neurons, but not muscles.
(A) Quantification of ACR-12::GFP clusters in the synaptic region of the DD1 dendrite for the genotypes indicated, normalized to wild type. Red line indicates 50% reduction in puncta number. Colored …
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Figure 3—source data 1
Raw values for normalized ACR-12::GFP receptor clusters, muscle UNC-29::GFP receptor clusters, and ventral/dorsal ACR-12::GFP receptor clusters in nrx-1mutants.
- https://doi.org/10.7554/eLife.35692.017
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Figure 3—source data 2
Raw values for UNC-29::GFP and UNC-63::GFP receptor cluster number, normalized wild type and nrx-1 dendrite fluorescence, and SNB-1::GFP clusters.
Relates to Figure 3—figure supplements 1, 2 and 3.
- https://doi.org/10.7554/eLife.35692.018
Figure 3—figure supplement 1
Description of genes surveyed in candidate-based genetic screen.
Description of genes surveyed in Figure 3A and their relation to mammalian genes. Candidates predominantly encode scaffold and cell-cell interaction proteins previously implicated in synapse …
Figure 3—figure supplement 2
LEV-10 and MADD-4 are required for muscle L-AChR clustering, and nrx-1 deletion does not disrupt transgene expression.
(A) Representative confocal images of the adult dorsal nerve cord region in wild type (WT), lev-10(ok2111), and madd-4(ok2854) mutants expressing myo-3::UNC-29::GFP. Scale bar, 5 μm. (B) …
Figure 3—figure supplement 3
Loss of functional nrx-1 disrupts ACR-12 AChR localization, but nrx-1 is not required for AChR membrane insertion or synaptic remodeling.
(A) Top, schematic depicting pre-synaptic (red) and post-synaptic (green) remodeling of DD motor neurons during development. Bottom, confocal images of wildtype and nrx-1(ok1649) mutants imaged …
Figure 3—figure supplement 4
Neurexin is not essential for AMPAR localization or synaptic vesicle clustering in cholinergic motor neurons.
(A) Representative confocal images of GLR-1::GFP in the anterior ventral nerve cord of adult animals. Scale bar, 5 μm. (B) Left, representative confocal images of the dorsal nerve cord from five …
Figure 4 with 1 supplement
Synaptic architecture is dependent on nrx-1/neurexin.
(A) Fluorescent confocal images of spine-like protrusions in the synaptic region of the DD1 dendrite (flp-13::mCherry) for the genotypes indicated. Inverted images show expanded views of the …
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Figure 4—source data 1
Raw values for spine-like protrusion number.
- https://doi.org/10.7554/eLife.35692.021
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Figure 4—source data 2
Raw values for adult ACR-12::GFP receptor cluster number in DD3 and DD1 spine-like protrusion number at L2.
Relates to Figure 4—figure supplement 1.
- https://doi.org/10.7554/eLife.35692.022
Figure 4—figure supplement 1
NRX-1 is required for ACR-12 receptor clustering and spine outgrowth in DD motor neurons.
(A) Left, representative confocal images of ACR-12::GFP clusters in the dendrite of the DD3 neuron (flp-13 promoter) for wild type and nrx-1(ok1649) mutants. Scale bar, 5 μm. Right, quantification …
Figure 5 with 3 supplements
Cell-specific expression of neurexin in cholinergic motor neurons restores ACR-12 localization to synapses.
(A) Confocal images of the ventral nerve cord in a transgenic strain expressing nrx-1L::GFP together with the cholinergic motor neuron marker acr-2::mCherry. Asterisks indicate coexpression. Scale …
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Figure 5—source data 1
Raw values for ACR-12::GFP receptor cluster number with expression of neurexin rescuing constructs and for colocalization of mCherry::Rab-3 and NRX-1::GFP fluorescence.
- https://doi.org/10.7554/eLife.35692.027
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Figure 5—source data 2
Raw values for spine-like protrusions and ACR-12::GFP receptor clustering with rescue constructs, and for NRX-1::GFP fluorescence in indicated mutants.
Relates to Figure 5—figure supplements 1, 2 and 3.
- https://doi.org/10.7554/eLife.35692.028
Figure 5—figure supplement 1
NRX-1 acts presynaptically to regulate spine outgrowth.
(A) Inverted images of spine-like protrusions in the synaptic region of the DD1 dendrite (flp-13::mCherry) for the genotypes indicated. Rescue refers to cholinergic expression (ufEx1114) of NRX-1L …
Figure 5—figure supplement 2
NRX-1 protein lacking the PDZ binding motif rescues ACR-12 receptor clustering defects in nrx-1 mutants and localizes to cholinergic axon terminals.
(A) Representative confocal images showing ACR-12::GFP clusters in the synaptic region of the DD1 dendrite for the genotypes indicated. In A and B, +ΔPDZ rescue refers to cholinergic-specific (unc-17…
Figure 5—figure supplement 3
Loss of nlg-1 or acr-12 does not affect NRX-1 localization to cholinergic axons.
(A) Confocal images of NRX-1L::GFP in the dorsal nerve cord of wild type or nlg-1(ok259) mutant. Pharynx is outlined by white dashed line. Scale bar, 5 μm. (B) Confocal images of the dorsal nerve …
Figure 6
Transcriptional control of neurexin expression in ACh motor neurons by UNC-3.
(A) Confocal images of the DD1 synaptic region and cell soma in nrx-1(ok1649) and unc-3(e151) mutants expressing flp-13::mCherry (upper) or flp-13::ACR-12::GFP (lower). Note the absence of …
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Figure 6—source data 1
Raw values for spine-like protrusion and ACR-12::GFP receptor cluster number, and for nrx-1 mRNA FISH signal.
- https://doi.org/10.7554/eLife.35692.030
Figure 7 with 1 supplement
nrx-1 mutants show functional defects in synaptic connectivity.
(A) Confocal images of pre-synaptic (acr-2::mCherry::RAB-3) and post-synaptic (flp-13::ACR-12::GFP) specializations in the DD1 synaptic region for the genotypes indicated. Arrowheads indicate …
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Figure 7—source data 1
Raw values for calcium imaging peak fluorescence and normalized peak fluorescence.
- https://doi.org/10.7554/eLife.35692.033
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Figure 7—source data 2
Raw values for body bend amplitude ratio during movement.
Relates to Figure 7—figure supplement 1.
- https://doi.org/10.7554/eLife.35692.034
Figure 7—figure supplement 1
nrx-1 mutants have defects in transmission onto GABAergic neurons and abnormalities in dorsoventral bending.
(A) Simplified schematic of experimental setup for calcium imaging studies. GCaMP6s fluorescence was recorded in GABAergic neurons or body wall muscle cells in a single focal plane over a 20 s …
Figure 8
Distinct molecular scaffolds direct partner-specific connectivity.
Distinct molecular scaffolds coordinate post-synaptic development in GABAergic neurons vs muscle. NRX-1/neurexin located at sites of presynaptic cholinergic release acts to coordinate ACR-12 …
Additional files
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Supplementary file 1
Strain list.
- https://doi.org/10.7554/eLife.35692.036
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Supplementary file 2
Plasmid list.
- https://doi.org/10.7554/eLife.35692.037
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Supplementary file 3
Primer list.
- https://doi.org/10.7554/eLife.35692.038
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Transparent reporting form
- https://doi.org/10.7554/eLife.35692.039
