Extended Data Fig. 4: Warmth increases Lim1 levels in DN1ps in a TRPA1-dependent manner.
From: Behavioral adaptation to warm conditions via Lim1-mediated acceleration of neuronal clocks
a, Levels of a pan-neuronal protein Osa (a subunit of the SWI/SNF chromatin remodeling complex) in DN1ps do not change with temperature. n = 11 and 10 at 25 °C and 30 °C, respectively, Student’s t-tests. b, Quantification of Lim1 levels in DN1p neurons of controls (w1118), TRPA11 and nocte1 mutants, and flies with aristae surgically removed. TRPA1 mutation, but not other manipulations, prevents Lim1 levels in DN1ps from increasing when temperature increases. n (25 °C, 30 °C): w1118 (32, 15); TRPA11 (32, 17); nocte1 (27, 17); flies with no aristae (33, 16), two-way ANOVA followed by Tukey’s multiple comparisons test. c, Immunostaining for Lim1 and CLK. Lim1 is expressed in DN1ps and DN2s. Lim1 signal is eliminated from DN1ps but not DN2s when lim1-RNAi is expressed with DN1-Gal4 during adulthood. Scale bar = 5 µm. d, Averaged daily locomotor activity (top) and sleep (bottom) profiles in 12-h light/12-h dark conditions (denoted by white–black bars on the bottom). Flies with Lim1 depleted from DN1 neurons during adulthood (DN1>lim1-RNAi;Tub-Gal80ts) are compared to the parental controls (DN1-Gal4 and UAS-lim1-RNAi;Tub-Gal80ts). Please note that in this experiment flies lost more sleep after shifting from 25 °C to 30 °C than in experiments shown in Figs. 1a and 7a, likely because they were raised at 18 °C; flies raised at lower temperature are more sensitive to warming (our observation). Although the combination of 18 °C rearing and 30 °C testing induced a significant baseline reduction in sleep, a statistically significant difference can be observed between Lim1 knockdown flies and controls during the predawn hours (black bars in left panels) within the 30 °C condition. n (25 °C, 30 °C): DN1-Gal4 (32, 35); UAS-lim1-RNAi;Tub-Gal80ts (13, 23); DN1>lim1-RNAi;Tub-Gal80ts (32, 28), two-way ANOVA followed by Tukey’s multiple comparisons test. Quantification represents mean ± s.e.m. ***P < 0.001.
