The ion channel ppk301 controls freshwater egg-laying in the mosquito Aedes aegypti
- The Rockefeller University, United States
- Howard Hughes Medical Institute, United States
- Kavli Neural Systems Institute, United States
Figures
Figure 1
Ae. aegypti mosquitoes use freshwater contact to guide egg-laying, ensuring offspring survival.
(A) Still video frames of Ae. aegypti female with newly laid eggs (yellow triangles) and contact between sensory appendages and liquid water (red circles) indicated. Right panels are magnification …
Figure 2
Genomic organization and tissue-specific expression of Ae. aegypti ppk ion channels.
(A) Genomic organization of ppk ion channels on the three chromosomes of Ae. aegypti. (B) Phylogenetic tree of ppk ion channel proteins from Ae. aegypti, Anopheles gambiae, and Drosophila …
Figure 3
Mutations in the ppk301 ion channel disrupt egg-laying preference and timing.
(A, B) Females of indicated genotype who blood-fed (A) or laid >10 eggs given 18 hr to lay (B). n = 6–12 groups/genotype. (C) Embryos/ovary 72 hr post-blood-meal. n = 8–16/genotype. (D–F) Eggs laid …
Figure 4 with 1 supplement
In-frame insertion of T2A-QF2 into the ppk301 locus labels sensory neurons that project to central taste centers.
(A) Top, construct used to test the efficacy of T2A in Ae. aegypti. Bottom, confocal images of three Ae. aegypti body wall cells expressing both mCD8:GFP (green) and dsRed:NLS (red). (B) Diagram of …
Figure 4—figure supplement 1
Male expression of ppk301>mCD8:GFP and QF2/QUAS controls for ppk301-QF2 reagent.
Expression of ppk301>mCD8:GFP in males and controls for QF2/QUAS expression. (A) Male Ae. aegypti mosquito. Photo: Alex Wild. (B, C) Expression of ppk301>CD8:GFP in male brain (B) or ventral nerve …
Figure 5
ppk301 neurons respond to water and salt.
(A) Side view schematic of ventral nerve cord imaging setup. (B) Top, ventral view schematic of mosquito during imaging showing ventral nerve cord (magenta) and imaging window (gray box). Bottom, …
Figure 6
ppk301 neurons are not tuned to osmotic pressure.
(A) Top left, two-photon image of dTomato (red) and region of interest (yellow). All other panels show representative GCaMP6s fluorescence increase to indicated solutions at 200mOsm/kg compared to …
Figure 7
ppk301 gates egg laying, pointing to integration of water and salt cues that drive preference.
(A) Raster plot of eggs laid (circles) and physical contact with liquid (gray boxes) by eight individual females of the indicated genotype over 40-min observation period in the indicated NaCl …
Videos
Video 1
Ae. aegypti mosquitoes use contact with freshwater to guide egg-laying (related to Figure 1).
This video shows a single female mosquito, 4 days after a blood-meal, in a small chamber with deionized water and a vertically oriented piece of black paper partially submerged in the water. The …
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Antibody | anti-Brp (mouse monoclonal) | Developmental Studies Hybridoma Bank | DSHB:NC82 RRID:AB_2314866 | (1:50) |
| Antibody | anti-GFP (rabbit polyclonal) | Life Technologies | Molecular Probes: A11122 RRID:AB_221569 | (1:10,000) |
| Chemical compound, drug | L-serine | Sigma | Sigma:s4500 | |
| Chemical compound, drug | D-(+)-cellobiose | Sigma | Sigma:22150 | |
| Chemical compound, drug | NaCl | Sigma | Sigma:s7653 | |
| Gene (Aedes aegypti) | ppk301 | NA | NCBI:LOC5564307 | previously known as AAEL000582, ppk582 |
| Gene (Ae. aegypti) | ppk204 | NA | NCBI:LOC5573846 | previously known as AAEL010779, ppk10779 |
| Gene (Ae. aegypti) | ppk316 | NA | NCBI:LOC5567201 | previously known as AAEL000926, ppk00926 |
| Gene (Ae. aegypti) | ppk306 | NA | NCBI:LOC5563925 | previously known as AAEL014228, ppk14228 |
| Genetic reagent (Ae. aegypti) | ppk301 -/- | PMID:25818303 | NA | previously generated and referred to asAAEL00582 ECFP in Kistler et al., 2015 (PMID:25818303). Contains an insertion of a poly-ubiquitin promoter-driven ECFP into the coding sequence of ppk301 |
| Genetic reagent (Ae. aegypti) | ppk204 -/- | PMID:25818303 | NA | previously generated and referred to asAAEL010779 -/- in Kistler et al., 2015 (PMID:25818303). Contains a 150 bp deletion in the coding sequence ofppk301 with a corresponding integration of a cassette with triple-stop codons. |
| Genetic reagent (Ae. aegypti) | ppk316 -/- | PMID:25818303 | NA | previously generated and referred to asAAEL00926-/- in Kistler et al., 2015 (PMID:25818303). Contains an 86 bp deletion in the coding sequence of ppk316 with a corresponding integration of a cassette with triple-stop codons and several downstream SNPs. |
| Genetic reagent (Ae. aegypti) | ppk306 -/- | PMID:25818303 | NA | previously generated and referred to asAAEL014228-/- in Kistler et al., 2015 (PMID:25818303). Contains a 106 bp deletion in the coding sequence of ppk306 with a corresponding integration of a cassette with triple-stop codons. |
| Genetic reagent (Ae. aegypti) | ppk301-T2A-QF2 | this paper | NA | T2A-QF2 knockin into the ppk301 locus, described in Materials and methods. |
| Genetic reagent (Ae. aegypti) | Pub-mCD8:GFP-T2A-dsRed:NLS-SV40 | this paper | NA | Ae. aegypti poly-ubiquitin promoter driving expression of membrane tethered GFP and nuclear dsRed separated by a T2A peptide. Described inMaterials and methods |
| Genetic reagent (Ae. aegypti) | 15x-QUAS-mCD8-GFP | this paper | NA | QUAS reporter strain driving expression of membrane-targeted GFP. Produced by transforming wild-type Ae. aegypti with plasmid RRID:Addgene_104878 |
| Genetic reagent (Ae. aegypti) | 15x-QUAS-dTomato-T2A-GCaMP6s | this paper | NA | QUAS reporter strain driving expression of cytosolic dTomato and GCaMP6s separated by a T2A peptide. Described in Materials and methods |
Additional files
-
Supplementary file 1
- https://doi.org/10.7554/eLife.43963.012
-
Transparent reporting form
- https://doi.org/10.7554/eLife.43963.013
