Figures and data
Phylogenetic reconstruction of termite ORs and their transcript abundances in P. simplex workers.
A. Phylogenetic tree is based on 182 protein sequences from five species of termites and the bristletail Lepisma saccharina as a basal insect outgroup, and also includes the sequences of ORCo. The topology and branching supports were inferred using the IQ-TREE maximum likelihood algorithm with the JTT+F+R8 model and supported by 10,000 iterations of ultrafast bootstrap approximation. Protein sequences of termite ORs can be found under the same labeling in Johny et al. (2023). Lepisma saccharina sequences are listed in Thoma et al. (2019). Arrowheads highlight the four ORs from Prorhinotermes simplex selected for functional characterization. Fully annotated version of the tree is provided as Supplementary Fig. S1. B. Heatmap shows the transcript abundances of 50 ORs identified in the RNAseq data from P. simplex worker antennae available in NCBI SRA archive under accession SRX17749141.
SSR responses of transgenic D. melanogaster ab3 sensillum expressing PsimOR9, 14, 30 and 31 to the initial screening of 11 volatiles with biological relevance for termites.
A. Heatmap showing the average responses of the four ORs as Δ spikes/s from 3–6 independent replicates. B. Comparison of SSR responses of transgenic Drosophila melanogaster ab3A neurons expressing PsimOR14 (ab3A:PsimOR14) and W1118 D. melanogaster. The bars show the average Δ spikes/s values from five independent replicates ± SEM. C. Characteristic SSR traces of ab3A:PsimOR14 and W1118 flies for 1 µg dose of neocembrene and geranylgeraniol. D. Dose response curve of ab3A:PsimOR14 SSR responses to neocembrene. The graph shows average Δ spikes/s values ± SEM based on nine replicates (8 in case of 100 ng and 4 in case of 500 ng stimulations). The curve fit and ED50 value were calculated using log(agonist) vs. response non-linear algorithm with least square fit method and the constraint of minimal response > 0. The raw data for all graphs is provided in Supplementary Tables S1–S6.
SSR responses of transgenic D. melanogaster ab3 sensillum expressing PsimOR14 to the complete set of 67 compounds (Panels 1–4).
A. Heatmap showing the average responses as Δ spikes/s from 3–6 independent replicates. B. Tuning curve of PsimOR14 for the 67 compounds contained in panels 1–4. The raw data is provided in Supplementary Tables S7–S9.
Neocembrene-responding sensillum in P. simplex workers.
A. SEM photograph of the last flagellomere of P. simplex worker. Arrow shows a small multiporous grooved sensillum responding to neocembrene and geranylgeraniol. Scale bar represents 50 µm. B. HR-SEM view on the neocembrene-responding sensillum. Scale bar in the inset represents 500 nm. C. Detailed view on SSR traces recorded from the neocembrene-responding sensillum during spontaneous firing, and upon stimulation with neocembrene and geranylgeraniol.
SSR responses of the neocembrene-responding sensillum on the last flagellomere of P. simplex worker.
A. SSR responses to panel 1. The bars show the average Δ spikes/s values from 8–17 replicates ± SEM. The raw data is provided in Supplementary Table S10. B. Characteristic SSR traces of the neocembrene-detecting sensillum for neocembrene and geranylgeraniol. C. Dose response curve of the SSR responses to neocembrene by the neocembrene-responding sensillum. The graph shows average Δ spikes/s values ± SEM based on 9–11 replicates. The curve fit and ED50 value were calculated using log(agonist) vs. response non-linear algorithm with least square fit method and the constraint of minimal response > 0. The raw data is provided in Supplementary Table S11.
PsimOR14 gene, transcript and protein structures, docking and MD simulations.
A. Genomic locus containing PsimOR14 and PsimOR15. PsimOR14 gene consists of 1 non-coding and 5 protein coding exons. B. PsimOR14 transcript with 6 exons, showing the protein-coding (higher boxes) and untranslated regions (lower boxes), and ORF (arrow). C. Transmembrane architecture of PsimOR14. In red are shown seven residues interacting with neocembrene. Light blue ellipse shows the intracellular loop the most impacted by ligand binding. D. Modelled apoform of PsimOR14. Red region denotes the binding site identified via docking, light blue region represents the intracellular S4-S5 loop. E. Holoforms of PsimOR14 with three docked ligands. F. Absolute PsimOR4 dynamicity expressed as average volumes explored by atoms per simulation step in PsimOR14 apoform and upon binding the three studied ligands. G. Relative PsimOR14 dynamicity expressed as average explored atom volumes upon ligand binding relative to the volumes in PsimOR14 apoform. Nucleotide and protein sequences of PsimOR14 are provided in Supplementary Table S12 and as NCBI entry under accession OR921181.
Docking scores and energy values inferred from the docking experiment and from MM/PBSA simulations for binding interactions of neocembrene, geranylgeraniol and (+)-limonene with PsimOR14.
Caste comparison of PsimOR14 expression and EAG responses between P. simplex workers and soldiers.
A. Volcano plot representing edgeR differential gene expression analysis of all 50 P. simplex ORs in RNAseq data from soldier and workers heads (including antennae) sequenced in three independent biological replicates per caste. Colored dots mark ORs reaching absolute value of log2 fold change ≥ 1, horizontal lines represent p-value thresholds of 0.05 and 0.01. Numeric values of the edgeR and DESeq2 differential expression analysis are provided in Supplementary Table S14. Based on SRA archives accessible under SRX18952230–32 and SRX18952237–39. B. EAG responses of whole antenna preparations of workers and soldiers to neocembrene at a dose of 10 ng (mean ± SD shown on log2 scale). Inter-caste differences were compared using t-test on log2-transformed data. Raw data is shown in Supplementary Table S15.
Full version of the phylogenetic tree of termite ORs shown in Fig. 1 of the main text.
Protein sequences of termite ORs can be found under the same labeling in Johny et al. (2023). Lepisma saccharina sequences used as basal insect outgroup are listed in Thoma et al. (2019). The topology and branching supports were inferred using the IQ-TREE maximum likelihood algorithm with the JTT+F+R8 model and supported by 10,000 iterations of ultrafast bootstrap approximation.
Crossing scheme of termite ORs heterologous expression using Drosophila melanogaster empty neurons in ab3 sensilla.
SSR responses to Panel 1 for PsimOR9.
Related to Fig. 2.
SSR responses to Panel 1 for PsimOR14.
Related to Fig. 2.
SSR responses to Panel 1 for PsimOR30.
Related to Fig. 2.
SSR responses to Panel 1 for PsimOR31.
Related to Fig. 2.
SSR responses to Panel 1 for PsimOR14 vs. W1118.
Related to Fig. 2.
SSR dose response data for neocembrene and PsimOR14 fly line.
Related to Fig. 2.
SSR responses to Panel 2 for PsimOR14.
Related to Fig. 3.
SSR responses to Panel 3 for PsimOR14.
Related to Fig. 3.
SSR responses to Panel 4 for PsimOR14.
Related to Fig. 3.
SSR responses to Panel 1 by neocembrene sensillum in P. simplex workers.
Related to Fig. 5.
SSR dose response data for neocembrene and P. simplex neocembrene sensillum.
Related to Fig. 5.
Nucleotide and protein sequences of PsimOR14.
Related to Fig. 6.
MM/PBSA calculated interaction energies of ligands with PsimOR14 decomposed into per-residue contributions.
Differential expression of P. simplex ORs (DESeq2 and edge R analyses), based on heads (including antennae) of workers and soldiers (three replicates).
Related to Fig. 7.
Differential sensitivity of P. simplex workers and soldiers to neocembrene inferred from EAG responses to the dose of 10 ng.
Related to Fig. 7.
List of primers.
Characteristics and origin of D. melanogaster lines used.
Origin of chemicals.
