Presynaptic LRP4 promotes synapse number and function of excitatory CNS neurons
- Thomas Jefferson University, United States
- Howard Hughes Medical Institute, Stanford University, United States
- Stanford University, United States
Figures
Figure 1 with 3 supplements
LRP4 is a synaptic protein expressed in excitatory neurons.
(A) Domain structure of Drosophila LRP4. Numbers indicate amino acids. EXT, extracellular side. INT, intracellular side. (B) Representative confocal image stack of a control Drosophila brain stained …
Figure 1—figure supplement 1
Sequence alignment of Drosophila, mouse, and human LRP4 homologues.
Multiple sequence alignment of Drosophila LRP4 (CG8909; accession AAF48538.1), Mus musculus LRP4 (accession NP_766256.3), and Homo sapiens LRP4 (accession NP_002325.2). Red shading = identical …
Figure 1—figure supplement 2
LRP4 reagents and patterns of LRP4 expression.
(A) Genomic region of lrp4. Top bar represents physical position on the X chromosome (in base pairs), and the blue arrow represents the lrp4 genomic region flanked by other genes (yellow). Primer …
Figure 1—figure supplement 3
Validation of expansion microscopy in Drosophila.
(A) A representative Drosophila brain stained with antibodies against endogenous Brp (green) and processed for proExM. Organization of the fly brain is maintained as are identifiable landmarks. …
Figure 2 with 4 supplements
LRP4 perturbation in excitatory neurons alters synapse number.
(A) Schematic diagram of the fly brain with major regions labeled and the olfactory regions examined in this study shaded in red (AL, antennal lobe) or yellow (LH, the lateral horn). Olfactory …
Figure 2—figure supplement 1
Representative antennal lobe images for genetic lrp4 manipulations.
(A–D) Representative high magnification confocal maximum intensity projections of VA1v ORN axon terminals in males expressing Brp-Short-mStraw and stained with antibodies against mStraw (red) and …
Figure 2—figure supplement 2
lrp4 perturbation in females affects synapse number.
(A–E) Representative high magnification confocal maximum intensity projections of VA1v ORN axon terminals in females expressing Brp-Short-mStraw and stained with antibodies against mStraw (red) and …
Figure 2—figure supplement 3
lrp4 RNAi reduces synapse number in multiple glomeruli.
(A–I) Representative high magnification confocal maximum intensity projections of ORN axon terminals in males expressing Brp-Short-mStraw (red) projecting to the DL4 and DM6 (A–C), VA1d (D–F), or …
Figure 2—figure supplement 4
lrp4 RNAi reduces Syd1 puncta in presynaptic ORN terminals.
(A–C) Representative high magnification confocal maximum intensity projections of DA1 ORN axon terminals in males expressing DSyd1-GFP and stained with antibodies to GFP (green) and N-Cadherin …
Figure 3
Loss of LRP4 causes defects in T-bar number and morphology.
(A–B) Representative transmission electron micrographs of putative ORN terminal in Control (A) and lrp4dalek (B) adult antennal lobes. Loss of lrp4 results in fewer observed T-bar profiles …
Figure 4
Effects of LRP4 perturbation on inhibitory neuron synapse formation.
(A–C) Representative high magnification confocal maximum intensity projections of GAD1-positive inhibitory neurons, which project to the DA1 glomerulus (dashed line), in males expressing …
Figure 5 with 1 supplement
LRP4 perturbations similarly affect higher order olfactory centers.
(A) Schematic diagram of the fly brain with major regions labeled and the olfactory regions examined in this study shaded in red (AL, antennal lobe) or yellow (LH, the lateral horn). Excitatory …
Figure 5—figure supplement 1
Representative lateral horn images for LRP4 genetic manipulations.
(A–C) Representative high magnification confocal maximum intensity projections of Mz19-GAL4 positive PN axon terminals in the lateral horn in males expressing Brp-Short-mStraw and mCD8-GFP and …
Figure 6
Loss of presynaptic LRP4 abolishes olfactory attraction behavior.
(A) Cartoon of the olfactory trap. (B) Quantification of preference index [(# of flies in odor vial – # of flies in control vial) / total # of flies] between apple cider vinegar (odor) and water …
Figure 7 with 1 supplement
LRP4 is required for normal synaptic SRPK79D localization in the CNS.
(A–C) Representative images of larval transverse nerves stained with antibodies to Bruchpilot (Brp, green), Synaptotagmin I (Syt I, red), and HRP (blue). Loss of lrp4 (B, lrp4dalek) and srpk79d (C, s…
Figure 7—figure supplement 1
Proximity ligation assays reveal LRP4 and SRPK79D interactions.
(A–D) Representative single confocal slices of antennal lobes expressing venus-SRPK79D alone (A), LRP4-HA alone (B) or venus-SRPK79D and LRP4-HA (C–D) in all ORNs using the pebbled-GAL4 driver and …
Figure 8 with 1 supplement
SRPK79D and LRP4 genetically interact to control synapse morphology and function.
(A–D) Representative high magnification confocal maximum intensity projections of VA1v ORN axon terminals in males expressing Brp-Short-mStraw and stained with antibodies to mStraw (red) and …
Figure 8—figure supplement 1
lrp4 and srpk79D interact genetically to control Brp-Short puncta number.
(A–D) Representative high magnification confocal maximum intensity projections of VA1v ORN axon terminals in the VA1v glomerulus of males expressing Brp-Short-mStraw and stained with antibodies …
Additional files
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Supplementary file 1
Table of genotypes for all experimental conditions.
Full genotype information for each image or analyzed condition. Notation (+; +; +; +) follows standard Drosophila genetics, corresponding to the X, 2nd, 3rd, and 4th chromosomes. Corresponding figure panels are listed as well for each genotype.
- https://doi.org/10.7554/eLife.27347.021
