Transsynaptic interactions between IgSF proteins DIP-α and Dpr10 are required for motor neuron targeting specificity
- University of Chicago, United States
- Baylor College of Medicine, United States
- University of California, Los Angeles, United States
- California Institute of Technology, United States
Figures
Figure 1 with 1 supplement
DIP-α is expressed in a subset of neurons that includes MNISN-1s/RP2.
(A–B) DIP-α-T2A-GAL4 driving EGFP expression in (A) embryonic and (B) third instar larval ventral nerve cords labeled with anti-GFP (green) and anti-Eve (red). Arrowheads denote segmentally …
Figure 1—figure supplement 1
DIP-α is expressed in a subset of VNC neurons and DIP-α protein is enriched in the neuropil.
(A) Ventral confocal projections of VNC in st16 embryos depicting DIP-α-GAL4 expression of EGFP. Arrow denotes MNISNb/d-1s. (B) Dorsal confocal projections of VNC in larval animals depicting DIP-α-GA…
Figure 2 with 1 supplement
DIP-α is required for MNISN-1s branch formation on m4.
(A–C) Larval neuromuscular junctions labeled with anti-GFP (green) and anti-HRP (red) from (A) heterozygous DIP-α-GAL4 animals, (B) homozygous DIP-α-GAL4 mutants and (C) rescue of homozygous DIP-α-GA…
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Figure 2—source data 1
Data for graphs in Figure 2.
- https://doi.org/10.7554/eLife.42690.006
Figure 2—figure supplement 1
DIP-α protein from a transgene localizes normally in rescue animals.
(A,B) The DIP-α rescue construct, tagged with a c-terminal Myc tag, labeled with (A) anti-Myc (green), anti-HRP (red) and anti-Dlg (blue) or (B) anti-DIP-α (green), anti-HRP (red) or phalloidin …
Figure 3 with 1 supplement
MNISN-1s axons in DIP-α mutants produce protrusions on m4 but are unable to form stable 1s branches.
(A–D) Embryonic NMJs labeled with anti-GFP (green), anti-HRP (red) and phalloidin (blue) from (A,C) heterozygous and (B,D) hemizygous DIP-α-GAL4 mutants. (A–B) are st15 embryos; (C–D) are st17 …
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Figure 3—source data 1
Data for graphs in Figure 3—figure supplement 1.
- https://doi.org/10.7554/eLife.42690.009
Figure 3—figure supplement 1
DIP-α is expressed in MNISN-1s motor neurons in st14 embryos, and 1s arbors form over distal muscles before proximal.
(A) A ventral nerve cord labeled with anti-GFP (green) and anti-Fas2 (red) from a DIP-α-GAL4 driving EGFP st14 embryo. (B) Frequency of MNISN-1s terminal swellings (varicosity precursors) from …
Figure 4 with 1 supplement
Loss of DIP-α causes selective loss of MNISN-1s branches on proximal muscles and ectopic innervation of distal muscles.
(A,B) Dorsal body wall hemisegments labeled with anti-GFP (green), anti-HRP (red) and phalloidin (blue) from (A) heterozygous DIP-α-GAL4 animals, and (B) homozygous DIP-α-GAL4 mutants. Arrow denotes …
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Figure 4—source data 1
Data for graphs in Figure 4 and Figure 4—figure supplement 1.
- https://doi.org/10.7554/eLife.42690.012
Figure 4—figure supplement 1
Quantification of MNISN-1s arbor branches on m2.
(A) Quantification of the number of MNISN-1s branches on m2 in controls (DIP-α-T2A-GAL4/+) and hemizygous nulls (DIP-α-T2A-GAL4/Y). n (animals/hemisegments) = (13/54), (15/59) (respectively). Error …
Figure 5 with 1 supplement
Dpr10 is expressed in muscles and can localize to the SSR and bind to DIP-α.
(A,C) Embryonic ventral nerve cord from (A) dpr6-GAL4 and (C) dpr10-GAL4 driving EGFP expression, labeled with anti-GFP (green), anti-Eve (red) and anti-Elav (blue). dpr6 is expressed in aCC and RP2 …
Figure 5—figure supplement 1
Dpr10 is expressed in a subset of neurons in the embryonic ventral nerve cord, and anti-V5 staining in a control sample.
(A) An embryonic ventral nerve cord from dpr10-GAL4 > EGFP labeled with anti-GFP (green) and anti-Elav (red) from sections below the MNISN-1s neuron cell bodies seen in Figure 4. (B) A larval m4 NMJ …
Figure 6 with 1 supplement
Dpr10 is required for MNISN-1s innervation of muscle 4.
(A–C) Larval m4 NMJs labeled with anti-DLG (green) and anti-HRP (red) from (A) control w1118 animals, (B) dpr1014-5/Df null mutants and (C) dpr6-GAL4/Df. The difference in anti-Dlg staining …
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Figure 6—source data 1
Data for graphs in Figure 6 and Figure 6—figure supplement 1.
- https://doi.org/10.7554/eLife.42690.017
Figure 6—figure supplement 1
MNISN-1s innervation of dorsal muscles and branching on m2.
(A) Frequency of MNISN-1s innervation of dorsal muscles from control (w1118) and dpr10 mutants. (B) Quantification of MNISN-1s branches on m2 in control (UAS-Dpr10/+) and muscle …
Figure 7
Temporal and spatial expression of dpr10 in embryonic muscles.
(A,C,E) Embryonic dpr10-GAL4>EGFP abdominal body wall hemisegments labeled with anti-GFP (green), anti-Fas2 (red) and phalloidin (blue) from (A) late st14/early st15, (C) late st15/early st16, and (E…
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Figure 7—source data 1
Data for graphs in Figure 7.
- https://doi.org/10.7554/eLife.42690.019
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Genetic reagent (D. melanogaster) | w1118 | Bloomington Drosophila Stock center (BDSC) | ||
| Genetic reagent (D. melanogaster) | Mef2-GAL4 | PMID: 9671578 | Gift of Hugo Bellen | |
| Genetic reagent (D. melanogaster) | DIP-α-T2A-GAL4 | PMID: 21985007 | Gift of Hugo Bellen | |
| Genetic reagent (D. melanogaster) | DIP-α-EGFP-DIP-α | PMID: 26687361 | Gift of Hugo Bellen | |
| Genetic reagent (D. melanogaster) | DIP-α1-178 | PMID: 30467079 | Gift of Lawrence Zipursky | |
| Genetic reagent (D. melanogaster) | dpr1014-5 | PMID: 30467079 | Gift of Lawrence Zipursky | |
| Genetic reagent (D. melanogaster) | UAS-DIP-α-Myc | PMID: 30467079 | Gift of Lawrence Zipursky | |
| Genetic reagent (D. melanogaster) | UAS-Dpr10-V5 | PMID: 30467079 | Gift of Lawrence Zipursky | |
| Genetic reagent (D. melanogaster) | dpr10-T2A-GAL4 | PMID: 21985007 | Gift of Hugo Bellen | |
| Genetic reagent (D. melanogaster) | dpr6-T2A-GAL4 | PMID: 21985007 | Gift of Hugo Bellen | |
| Genetic reagent (D. melanogaster) | UAS-2XEGFP | PMID: 12324968 | BDSC #6874 | |
| Genetic reagent (D. melanogaster) | Elav-GAL4 | PMID: 22319582 | BDSC #8765 | |
| Genetic reagent (D. melanogaster) | 24B-GAL4 | PMID: 8223268 | BDSC #1767 | |
| Genetic reagent (D. melanogaster) | MHC-GeneSwitch | PMID: 11675495 | Gift of Haig Keshishian | |
| Genetic reagent (D. melanogaster) | EveRN2-GAL4 | PMID: 14624243 | BDSC #7470 | |
| Genetic reagent (D. melanogaster) | UAS-dpr10-RNAi | PMID: 17625558 | Vienna Drosophila Resource Center #103511 | |
| Genetic reagent (D. melanogaster) | UAS-DIP-α-RNAi | PMID: 26320097 | BDSC #38965 | |
| Genetic reagent (D. melanogaster) | Df(3L)BSC673 | BDSC #26525 | Deficiency covering dpr6 and dpr10 | |
| Antibody | Goat anti-HRP-TRITC | Jackson Immunological Research | #123-025-021 | 1:50 |
| Antibody | Goat anti-HRP-Alexa405 | Jackson Immunological Research | #123-475-021 | 1:50 |
| Antibody | Mouse anti-Dlg | Developmental Studies Hybridoma Bank | #4F3 | 1:100 |
| Antibody | Mouse anti-Fas2 | Developmental Studies Hybridoma Bank | #1D4 | 1:100 |
| Antibody | Mouse anti-DIP-α | PMID: 30467079 | Gift of Lawrence Zipursky | 1:20 |
| Antibody | Mouse anti-V5 | ThermoFisher | #R960-25 | 1:400 |
| Antibody | Rabbit anti-GFP | ThermoFisher | #A11122 | 1:1000 |
| Antibody | Rabbit anti-Dlg | PMID: 9354326 | Gift of Vivian Budnik | 1:40,000 |
| Antibody | Rabbit anti-Myc | Cell Signaling Technology | #71D10 | 1:200 |
| Antibody | Goat anti-Mouse-Alexa488 | ThermoFisher | #A11029 | 1:500 |
| Antibody | Goat anti-Mouse-Alexa568 | ThermoFisher | #A11031 | 1:500 |
| Antibody | Goat anti-Rabbit-Alexa488 | ThermoFisher | #A11008 | 1:500 |
| Antibody | Goat anti-Rabbit-Alexa568 | ThermoFisher | #A11036 | 1:500 |
| Chemical compound, drug | Phalloidin-Alexa647 | ThermoFisher | #A22287 | 1:100 |
| Antibody | Rabbit anti-alkaline phosphatase | Abcam | #ab16695 | 1:100 |
Additional files
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Transparent reporting form
- https://doi.org/10.7554/eLife.42690.020
