Extended Data Figure 5. Leg mechanosensory sensilla neurons but not chordotonal organs are necessary and sufficient for Encounter Responses.

We identified five lines expressing Gal4 in different subsets of mechanosensory neurons (R55B01-Gal4, piezo-Gal4, piezo-Gal4:cha3-Gal80, R86D09-Gal4, and R46H11-Gal4) and one line expressing Gal4 in the fan-shaped body (R65C03-Gal4) as a control for fan-shaped body expression in R55B01-Gal4. a, Brain and thoracic ganglion expression for Gal4 lines driving UAS-CD4:tdGFP. Immunostaining is shown for the neuropil marker nc82 (magenta) and CD4:tdGFP (green). Sensory neuron projections from the wings (‘W’) and legs (R1-R3 and L1-L3) are labelled for R55B01-Gal4. Importantly, neurons expressing GFP in the brains of R55B01-Gal4 and piezo-Gal4, cha3-Gal80 flies are different, implying that they are not responsible for the production of Encounter Responses. The scale bars are 40 μm. b, Transmitted light images, inverted GFP fluorescence images (GFP indicated in black), and summed fluorescence of Gal4 driver legs expressing CD4:tdGFP. Autofluorescent cuticle and pretarsus debris are indicated in black. GFP expression is shown in green. When present, the femoral chordotonal organ (‘fChO’), tibial chordotonal organ (‘tChO’) and mechanosensory sensilla neurons (‘MS’) are labelled. The scale bar is 100 μm. c, The frequency of Encounter Responses for a parental control (‘Gal4’), Gal4 line neurons expressing an inactive tetanus toxin control (‘Gal4 & ‘Inactive Tnt’), or Gal4 line neurons expressing tetanus toxin (‘Gal4’ & ‘Tnt’). N = 10-15 experiments for each condition. d, Blue laser pulse stimulation responses of Gal4 line flies expressing UAS-ChR2 in the absence (left) or presence (right) of the essential cofactor all trans-Retinal (n = 6-12 flies for each condition). Each box indicates the response for a single fly (‘walk’, ‘groom’, ‘leg shift’, ‘none’, or ‘jump’).