Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2019 Feb 7;176(4):687-701.e5.
doi: 10.1016/j.cell.2018.12.004.

Small-Molecule Agonists of Ae. aegypti Neuropeptide Y Receptor Block Mosquito Biting

Laura B Duvall  1 , Lavoisier Ramos-Espiritu  2 , Kyrollos E Barsoum  1 , J Fraser Glickman  2 , Leslie B Vosshall  3
Affiliations

Small-Molecule Agonists of Ae. aegypti Neuropeptide Y Receptor Block Mosquito Biting

Laura B Duvall et al. Cell. .

Abstract

Female Aedes aegypti mosquitoes bite humans to obtain blood to develop their eggs. Remarkably, their strong attraction to humans is suppressed for days after the blood meal by an unknown mechanism. We investigated a role for neuropeptide Y (NPY)-related signaling in long-term behavioral suppression and discovered that drugs targeting human NPY receptors modulate mosquito host-seeking. In a screen of all 49 predicted Ae. aegypti peptide receptors, we identified NPY-like receptor 7 (NPYLR7) as the sole target of these drugs. To obtain small-molecule agonists selective for NPYLR7, we performed a high-throughput cell-based assay of 265,211 compounds and isolated six highly selective NPYLR7 agonists that inhibit mosquito attraction to humans. NPYLR7 CRISPR-Cas9 null mutants are defective in behavioral suppression and resistant to these drugs. Finally, we show that these drugs can inhibit biting and blood-feeding on a live host, suggesting a novel approach to control infectious disease transmission by controlling mosquito behavior. VIDEO ABSTRACT.

Keywords: Aedes aegypti; CRISPR-Cas9; Zika; blood-feeding; dengue; feeding; high-throughput small-molecule screen; host-seeking behavior; mosquito; neuropeptide Y.

PubMed Disclaimer

Figures

Figure 1.
Figure 1.. Protein-rich blood-meals induce sustained host-seeking suppression
(A) Ae. aegypti females feeding on human skin or Glytube membrane feeders (photos by Alex Wild). (B) Female engorgement on the indicated meal delivered via Glytube (median with range, n=12, 10 - 15 females/trial, Kruskal-Wallis test with Dunn’s multiple comparison, n.s.: not significant p>0.05). (C) Weight per female of the indicated meal (median with range, n=12, 10 - 15 females/trial, one-way AnOVA followed by Tukey’s multiple comparison test). (D) Eggs produced per female after feeding on the indicated meal (median with range, n=37 - 48 females, Kruskal-Wallis test with Dunn’s multiple comparison). (E) Time course of human host-seeking in the uniport olfactometer after feeding with the indicated meal (median with range, n= 6 - 48, 15 - 25 females/trial, Kruskal-Wallis test with Dunn’s multiple comparison). Data labeled with different letters in C-E are significantly different from each other (p<0.05). Experimental groups denoted “AB” are not significantly different from either A or B groups.
Figure 2.
Figure 2.. Human NPY receptor drugs modulate Ae. aegypti host-seeking behavior
(A) Schematic of miniport olfactometer. Mosquitoes not drawn to scale. (B) Flowchart of human NPY receptor compound behavioral screen. (C) Effect of human NPY Y2/Y4 agonist TM30335 on host-seeking in a miniport olfactometer (n= 5 - 14, 15 - 25 females/trial). (D) Time course of human host-seeking in the uniport olfactometer after feeding with the indicated meal (n= 5 - 28, 15 - 25 females/trial. (E) Effect of human NPY Y2 antagonist BIIE0246 on host-seeking in a miniport olfactometer (n= 5 – 15, 15 – 25 females/trial). Data in C - E are plotted as median with range and data in D are plotted as median with interquartile range Data labeled with different letters in C and E are significantly different from each other (Kruskal-Wallis test with Dunn’s multiple comparison, p<0.05). Experimental groups denoted “AB” are not significantly different from either A or B groups.
Figure 3.
Figure 3.. In vitro screen identifies NPY-like receptor 7 as the target of behaviorally active compounds
(A-B) Response to TM30335 of all Ae. aegypti peptide receptors annotated in the L3 genome assembly and annotation (A) and additional receptors annotated in the L5 genome assembly and annotation (B) (mean ± SD, n=3 trials, 4 replicates/trial). Non-gray data points are statistically different from non-transfected control (black open circle), one-way aNoVA with Bonferroni correction, p<0.001. (C-D) In vitro response of 61 predicted Ae. aegypti peptides against all predicted peptide receptors in the L3 genome annotation (C) and the L5 genome annotation (D). See Data S1 for raw data. (E) Dose-response curve of TM30335 against the indicated receptors. NPYLR7 EC50 = 448 nM, NPYLR5 EC50 = 7.7 μM calculated using log(agonist) versus response nonlinear fit (mean ± SD, n=3 trials,4 replicates/trial). (F) Dose-inhibition curve of TM30335 activation NPYLR7 and NPYLR5 responses by BIIE0246. NPYLR7 IC50 =5.1 μM calculated using log(inhibitor) versus response nonlinear fit (mean ± SD, n=3 trials, 4 replicates/trial). (G) Summary of in vitro findings.
Figure 4.
Figure 4.. NPYLR7 mutants blood-feed normally but do not maintain sustained host-seeking suppression
(A) Snake plot of wild-type NPYLR7 and predicted truncated amino acid sequence of NPYLR7Δ4 mutant. Stop codon is indicated by the purple asterisk. (B) Engorgement of females of the indicated genotype on sheep blood delivered via Glytube (median with range, n=4 – 5, 35 - 50 females/trial, Kruskal-Wallis test with Dunn’s multiple comparison test, n.s., not significant, p>0.05). (C) Weight per female of non-blood-fed and sheep-blood-fed females (median with range, n= 8-10, 10 females/trial, one-way ANOVA with Sidak’s multiple comparisons test, n.s., not significant, p>0.05). (D) Percentage of females attracted to human host in the uniport olfactometer. Females were allowed to lay eggs between days 4 and 5. (median with range, n=5-27, 15-25 females/trial). Data labeled with different letters are significantly different from each other (Kruskal-Wallis test with Dunn’s multiple comparison, p<0.05). Experimental groups denoted “AB” are not significantly different from either A or B groups.
Figure 5.
Figure 5.. Small molecule screen identifies NPYLR7 agonists with in vivo activity
(A) Schematic of high-throughput small molecule screen for NPYLR7 agonists. (B) 24 confirmed in vitro hits tested for in vivo activity using the miniport olfactometer (median with interquartile range, n = 4 – 116, 15 - 25 females/trial). Compounds are indicated at the top of the figure with identifier number in a circle. Groups in green are significantly different compared to saline meal control (Kruskal-Wallis test with Dunn’s multiple comparison p<0.05). (c) In vivo dose-response tests of 6 primary hits in the miniport olfactometer (median with interquartile range, n=4 – 6, 15 - 25 females/trial). Data from inactive compound 8 are replotted in all 6 panels. (D-E) In vitro response profile of NPYLR7-activating compounds against all predicted peptide receptors in the L3 (D) and L5 (E) annotation of the Ae. aegypti genome. (F) In vitro response profile of NPYLR7-activating compounds against human NPY receptors.
Figure 6.
Figure 6.. Small molecule screen compounds target Ae. aegypti NPYLR7 with high specificity in vitro and in vivo
(A) Chemical structures of compound 18 and six structural analogues. (B) In vitro dose-response curve of compounds in (A) against NPYLR7 (mean ± SEM, n=1, 3 replicates/trial) (C) Host-seeking in a miniport olfactometer 2 days after feeding the indicated meals (n = 4 – 82, 15 - 25 females/trial) (D) Host-seeking in a miniport olfactometer 2 days after feeding the indicated genotypes with the indicated meals (n = 4-26, 15 - 25 females/trial). Data in C-D are plotted as median with range. Data labeled with different letters are significantly different from each other (Kruskal-Wallis test with Dunn’s multiple comparison, p<0.05). Experimental groups denoted “AB” are not significantly different from either A or B groups.
Figure 7.
Figure 7.. NPYLR7 agonist compound 18 inhibits mosquito blood-feeding on a live host
(A) Schematic of live host assay experiment. (B) Percentage of females fed the indicated meal 2 days prior to the experiment, which freshly blood-fed on an anesthetized mouse after a 15 min exposure (median with range, n = 5 – 24, 58 – 62 females/trial. Data labeled with different letters are significantly different from each other (Kruskal-Wallis test with Dunn’s multiple comparison, p<0.05).
See this image and copyright information in PMC

Comment in

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Alfonso-Parra C, Avila FW, Deewatthanawong P, Sirot LK, Wolfner MF, and Harrington LC (2014). Synthesis, depletion and cell-type expression of a protein from the male accessory glands of the dengue vector mosquito Aedes aegypti. Journal of insect physiology 70, 117–124. - PMC - PubMed
    1. Anderson JF, and Magnarelli LA (2008). Biology of ticks. Infectious disease clinics of North America 22, 195–215. - PubMed
    1. Babicki S, Arndt D, Marcu A, Liang Y, Grant JR, Maciejewski A, and Wishart DS (2016). Heatmapper: web-enabled heat mapping for all. Nucleic acids research 44, W147–153. - PMC - PubMed
    1. Batterham RL, Cowley MA, Small CJ, Herzog H, Cohen MA, Dakin CL, Wren AM, Brynes AE, Low MJ, Ghatei MA, et al. (2002). Gut hormone PYY(3-36) physiologically inhibits food intake. Nature 418, 650–654. - PubMed
    1. Bhatt S, Gething PW, Brady OJ, Messina JP, Farlow AW, Moyes CL, Drake JM, Brownstein JS, Hoen AG, Sankoh O, et al. (2013). The global distribution and burden of dengue. Nature 496, 504–507. - PMC - PubMed

Publication types

Substances

LinkOut - more resources