Extended Data Figure 7: Sleep characteristics of wild-type and genetically manipulated flies under ramping temperature cycles and constant conditions after entrainment to ramping temperature cycles.

a–l, Sleep data analysis for Canton S and genetically modified flies under 18 °C to 25 °C temperature ramping cycles (see Methods). Daily total sleep time (a, e, i); total sleep time during heating (red) and cooling (blue) phases (b, f, j); sleep bout number during heating (red) and cooling (blue) phases (c, g, k); and sleep bout duration during heating (red) and cooling (blue) phases (d, h, l) of the genotypes indicated. Number of flies used in the analysis: a–d, Canton S (n = 214), per⁰¹ (n = 96); e–h, nocte⁺ (n = 40), nocte⁺ AL (n = 28), nocte¹ (n = 49) and nocte¹ AL (n = 39); i–l, Clk4.1m/+ (n = 17), UAS-TNT/+ (n = 23) and Clk4.1m>TNT (n = 100). Data are shown as population average ± s.e.m. n = number of flies. m, Statistical analysis. For determining statistical significance, ANOVA was performed across each individual group. Tukey’s honest significant difference tests were conducted in pairwise fashion with P values indicated on the graphs: **P < 0.005, *P < 0.05. Unpaired two-tailed Student’s t-tests were conducted for two-group comparison of the heating phase and the cooling phase within each individual genotype. The number of flies is shown in panels a–l. n, Averaged population sleep plots on day 1 of constant conditions after entrainment to ramping temperature cycles for wild-type Canton S (black, n = 198) and mutant per⁰¹ (red, n = 87) flies. Circadian time (CT) 0 is the start of the subjective heating phase and CT12 is the start of the subjective cooling phase, the times when the temperature transitions would have occurred had the temperature cycle continued. Dark lines indicate mean and shaded regions in the plots indicate s.e.m. s, Averaged population sleep plots on day 4 of constant temperature after entrainment for Canton S (black, n = 198) and per⁰¹ (red, n = 83) flies. Dark lines indicate mean and shaded regions in the plots indicate s.e.m. o–r, t–w, Daily total sleep time, sleep times during subjective heating (red) and cooling (blue) phases, sleep bout number during subjective heating (red) and cooling (blue) phases, and sleep bout duration during subjective heating (red) and cooling (blue) phases of Canton S and per⁰¹ on day 1 (o–r) and day 4 (t–w) of constant conditions. The number of flies in panels o–r is shown in n, and the number of flies in panels t–w is shown in s. Data are presented as mean ± s.e.m. Canton S flies cycles under constant conditions display similar sleep patterns as observed during temperature cycles. By contrast, per⁰¹ flies do not display any differences in their sleep time or bout number or bout duration during the subjective heating phase and cooling phase, consistent with a lack of circadian timekeeping. n = number of flies. Statistical analysis was performed using unpaired two-tailed Student’s t-test. **P < 0.005, *P < 0.05. Individual P values are reported in the Source Data for this figure. Analysis of the sleep data revealed the following: 1. Wild-type flies sleep significantly more during the cooling phase than during the heating phase. 2. Wild-type flies have significantly more sleep bouts during the heating phase than during the cooling phase, indicating that sleep is more fragmented during the heating phase. 3. Wild-type flies have significantly longer sleep bout durations during the cooling phase compared to during the heating phase, indicating that sleep is more consolidated during the cooling phase. 4. Manipulation of chordotonal organ or aristae function does not produce changes in total sleep or sleep quality, only in the timing of sleep. 5. nocte¹ mutant flies that lack aristae (nocte¹ AL) fail to show significant differences in the amount of sleep during the heating and cooling phases. 6. Inhibition of a subset of DN1_ps results in decreased total daily sleep time and specifically reduces sleep time during the heating phase.

Source data