Figure 1.
PBAN stimulates trail pheromone biosynthesis in the fire ant.
(A) Photomicrograph of sting apparatus components -Dufour's gland (DG), poison gland/sac (PG), and sting in the worker. (B) Proposed pathway of trail pheromone biosynthesis in the worker. (C) SolinPBAN induces trail pheromone biosynthesis after SolinPBAN injection into workers. DGs were dissected from adult workers 6 h after SolinPBAN or saline (control) injection. The mean production of pheromone increased more than 30 ng (>19%) per ant after SolinPBAN injection. Data were analysed by two-tailed unpaired t-test (P = 0.0054, n≥35).
Figure 2.
Phylogenetic tree of receptors for PBAN, PK2 and neuromedin U (NmU).
The tree was made by Genetyx-Tree software (ver. 10) with the neighbor-joining distance phylogram method using bootstrap 1,000 replicates. Accession number of species: Acromyrmex echinatior (EGI70561), Acyrthosiphon pisum (XP_001950091), Aedes agypti (XP_001657210), Anopheles gambiae (AAX84798), Apis melifera (NP_001091688), Bombus terrestris (XP_003395149), Bombyx mori (NP_001036977), Camponotus floridanus (EFN62034), Culex quinquefasciatus (XP_001861460), Drosophila melanogaster (CG8795), Helicoverpa armigera (AAW47417), Helicoverpa zea (AAP93921), Heliothis virescens (ABU93812), Manduca sexta (ACQ90219), Mus musculus (NP_034471), Nasonia vitripennis (XP_001600587), Pediculus humanus (XP_002424393), Plutella xylostella (AAY34744), Solenopsis invicta (JX657040), Spodoptera exigua (ABY62317), Spodoptera littoralis (ABD52277), Tribolium castaneum (EEZ97728).
Figure 3.
SolinPBAN-R binding specificity and expression.
(A) Half-maximal effective concentration (EC50) values of synthetic peptides, SolinPBAN and SolinDH, of the fire ant to bind with SolinPBAN-R. The y-axis indicates the peak peptide fluorescence value relative to ionomycin treatment (mean ± s.e.m; N≥4). (B) Photomicrograph of worker ant and SolinPBAN-R gene expression from head, Br-SG and body (without head). (C) SolinPBAN-R gene expression from the complex of PG, DG and sting (PDS), PG, DG, sting (S) and ventral nerve cord (VNC). Fire ant 18S rRNA was used for the positive control.
Table 1.
Binding activity of PBAN/pyrokinin and related peptides to SolinPBAN-R.
Figure 4.
RNAi effects on trail pheromone production and SolinPBAN and SolinPBAN-R gene expressions in workers.
(A) Suppression of trail pheromone production and (B) SolinPBAN gene expression from Br-SG of workers at 24, 48 and 72 h post-injection of nuclease-free water (−) or SolinPBAN dsRNA (+). (C) Suppression of trail pheromone production and (D) SolinPBAN-R gene expression from DG of workers at 24, 48 and 72 h post-injection of nuclease-free water (−) or SolinPBAN-R dsRNA (+). Data were analyzed by two-tailed unpaired t-test (mean ± s.e.m., asterisks indicate, SolinPBAN RNAi at 48-h, P = 0.0479, n≥10, and SolinPBAN-R RNAi at 24-h, P = 0.0302, n≥10, at 48-h, P = 0.0136, n≥10, respectively). Fire ant 18S rRNA was sued for the positive control.