Temporal transcription factors determine circuit membership by permanently altering motor neuron-to-muscle synaptic partnerships
- Department of Molecular Genetics and Cell Biology, University of Chicago, United States
- Program in Cell and Molecular Biology, University of Chicago, United States
- Committee on Development, Regeneration, and Stem Cell Biology, University of Chicago, United States
- Grossman Institute for Neuroscience, University of Chicago, United States
Figures
Figure 1 with 2 supplements
Prolonged expression of Hunchback alters RP motor neuron synapses.
(A) Illustration of the late stage larvae (L3), body is organized into repeated left-right, mirror image hemisegments. Muscles (in gray) have a stereotyped pattern. (B) Illustration of muscles in a …
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Figure 1—source data 1
Test for difference in the number of synaptic branches on ventral muscles in NB3-1>Hb compared to Control.
- https://cdn.elifesciences.org/articles/56898/elife-56898-fig1-data1-v2.docx
Figure 1—figure supplement 1
NB3-1 generates RP motor neurons, with known features.
(A) Illustration of NB3-1 lineage progression. Each gray arrowhead represents cell division. Large circles are neuroblasts and smaller circles are neurons. Abbreviations: MN is motor neuron, IN is …
Figure 1—figure supplement 2
In embryos, prolonged expression of Hunchback generates more RP motor neurons with early born molecular identity.
(A–B) Illustrations of NB3-1 lineage progression. Each gray arrowhead represents cell division. Each gray arrowhead represents cell division. Large circles are neuroblasts, and smaller circles are …
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Figure 1—figure supplement 2—source data 1
Test for difference in the number of embryonic RP MN molecular identities in NB3-1>Hbcompared to Control.
- https://cdn.elifesciences.org/articles/56898/elife-56898-fig1-figsupp2-data1-v2.docx
Figure 2
Increased synaptic branches contain pre and postsynaptic markers necessary for function.
(A) Illustration of subcellular localization of neuromuscular synapse markers. Brunchpilot labels active zones, Discs large is a scaffolding protein strongly localized at post-synapse, and GluRIIA …
Figure 3
Altered synapses onto Ventral muscles are functional.
(A) Illustration of neuromuscular junction (NMJ) electrophysiology approach on third instar larvae (L3). Abbreviation NT is neurotransmitter. (B) Illustration highlighting (darkened) Muscle 30 whose …
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Figure 3—source data 1
Test for the difference in EPSP, mEPSP, and Quantal Content for muscle 14 and 6 in NB3-1>Hb compared to Control.
- https://cdn.elifesciences.org/articles/56898/elife-56898-fig3-data1-v2.docx
Figure 4 with 1 supplement
Precocious expression of Castor alters U motor neuron synapses.
(A–B) Illustrations of NB7-1 lineage progression. Each gray arrowhead represents cell division. Each gray arrowhead represents cell division. Large circles are neuroblasts, and smaller circles are …
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Figure 4—source data 1
Test for difference in the number of synaptic branches on dorsal muscles in NB7-1>Cas compared to Control.
- https://cdn.elifesciences.org/articles/56898/elife-56898-fig4-data1-v2.docx
Figure 4—figure supplement 1
In embryos, precocious expression of Castor generates fewer U motor neurons at the expense of later born neurons.
(A–B) Illustrations of NB7-1 lineage progression. Each gray arrowhead represents cell division. Each gray arrowhead represents cell division. Large circles are neuroblasts, and smaller circles are …
Figure 5 with 1 supplement
Precocious expression of Pdm alters U motor neuron synapses.
(A–C) Illustration of NB7-1 lineage progression. Each gray arrowhead represents cell division. Each gray arrowhead represents cell division. Large circles are neuroblasts, and smaller circles are …
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Figure 5—source data 1
Test for difference in the number of synaptic branches on dorsal muscles in NB7-1>Pdm compared to Control.
- https://cdn.elifesciences.org/articles/56898/elife-56898-fig5-data1-v2.docx
Figure 5—figure supplement 1
In embryos, precocious expression of Pdm generates either more or fewer U motor neurons depending on A/P positioning.
(A) Illustration of NB7-1 lineage progression. Each gray arrowhead represents cell division. Each gray arrowhead represents cell division. Large circles are neuroblasts, and smaller circles are …
Figure 6 with 1 supplement
Altered synapses onto Dorsal muscles are functional.
(A) Illustration of subcellular localization of neuromuscular synapse markers. Brunchpilot labels active zones, Discs large is a scaffolding protein strongly localized at post-synapse, GluRIIA is a …
Figure 6—figure supplement 1
Calcium imaging protocol, analysis, and examples.
(A–D) Illustration of calcium imaging protocol and analysis are shown. See Materials and methods for details. (E–H) Images of calcium signals on L3 Muscle 4. Control is NB7-1-GAL4/+; …
Figure 7
Summary of results in this study.
(A) Illustration of WT (wildtype) neuroblast lineage progression. Different patterns in the neuroblast represent temporal transcription factor expression. Outgrowth projecting from neuron represent …
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Genetic reagent (D. melanogaster) | DIP-alpha-GAL4 | Robert Carrillo (UChicago) | ||
| Genetic reagent (D. melanogaster) | ac:VP16, gsb:v8v (aka NB7-1-GAL4) | Minoree Kohwi (Columbia) | ||
| Genetic reagent (D. melanogaster) | MHC-CD8-GCaMP6f-Sh | Bloomington stock center [BL] 67739 | BDSC Cat# 67739, RRID:BDSC_67739 | |
| Genetic reagent (D. melanogaster) | UAS-Hb; UAS-Hb/TM2 | Bloomington stock center [BL] 32198 | BDSC Cat# 32198, RRID:BDSC_32198 | |
| Genetic reagent (D. melanogaster) | 5172 J-GAL4 (aka NB3-1-GAL4) | Haluk Lacin (Washington University) | ||
| Genetic reagent (D. melanogaster) | w1118 | Bloomington stock center [BL] 36005 | BDSC Cat# 36005, RRID:BDSC_36005 | |
| Genetic reagent (D. melanogaster) | UAS-Cas | Chris Doe (University of Oregon) | ||
| Genetic reagent (D. melanogaster) | UAS-HA-pdm2/CyO; UAS-HA-pdm2/TM3 | Chris Doe (University of Oregon) | ||
| Genetic reagent (D. melanogaster) | UAS-myr-GFP | Bloomington stock center [BL] 32198 | BDSC Cat# 32198, RRID:BDSC_32198 | |
| Antibody | rabbit anti-Eve (polyclonal) | Heckscher Lab | 1:1000 | |
| Antibody | rabbit anti-Castor (polyclonal) | Chris Doe (Oregon) | 1:1000 | |
| Antibody | guinea pig anti-Hb (polyclonal) | John Rientz (UChicago) | 1:1000 | |
| Antibody | guinea pig anti-Kruppel (polyclonal) | John Rientz (UChicago) | 1:1000 | |
| Antibody | Mouse anti-Eve (monoclonal) | 3C10 | DSHB Cat# 3C10 Anti-even skipped RRID:AB_528229 | 1:50 |
| Antibody | chicken anti-GFP (polyclonal) | Aves #GFP-1020 | Aves Labs Cat# GFP-1020, RRID:AB_10000240 | 1:1000 |
| Antibody | rat anti-Worniu (polyclonal) | Abcam #ab196362 | 1:250 | |
| Antibody | mouse anti-En (monoclonal) | 4D9 | DSHB Cat# 4D9 anti-engrailed/invected, RRID:AB_528224 | 1:5 |
| Antibody | guinea pig anti-Runt (polyclonal) | John Rientz (UChicago) | 1:500 | |
| Antibody | guinea pig anti-HB9 (polyclonal) | Heather Broiher (Case Western) | 1:1000 | |
| Antibody | mouse anti-Cut (monoclonal) | 2B10 | DSHB Cat# 2B10 anti-Cut homeobox RRID:AB_528186 | 1:50 |
| Antibody | rat anti-Zfh2 (polyclonal) | Chris Doe (Oregon) | 1:800 | |
| Antibody | rabbit anti-Smad3 (pMad) (polyclonal) | Abcam #52903 | Abcam Cat# ab52903, RRID:AB_882596 | 1:300 |
| Antibody | mouse anti-Brp (monoclonal) | NC82 | Creative Diagnostics Cat# DMAB9116MD, RRID:AB_2392664 | 1:50 |
| Antibody | mouse anti-DLG (monoclonal) | 4F3 | DSHB Cat# 4F3 anti-discs large, RRID:AB_528203 | 1:500 |
| Antibody | mouse anti-GluRIIA (monoclonal) | 8B4D2 | DSHB Cat# 8B4D2 (MH2B), RRID:AB_52826 | 1:25 |
| Antibody | Cy3-HRP (polyclonal) | Jackson ImmunoResearch 123-025-021 | Jackson ImmunoResearch Labs Cat# 123-025-021, RRID:AB_2338954 | 1:400 |
| Antibody | 647-Phallion (polyclonal) | Thermofisher A22287 | Thermo Fisher Scientific Cat# A22287, RRID:AB_2620155 | 1:600 |
