Extended Data Fig. 6: Related to Fig. 3. Control experiments for the effect of Letm1 KD on fly olfactory memory.
From: Mitochondrial Ca2+ efflux controls neuronal metabolism and long-term memory across species
(A-B) Letm1 RNAi #2 flies were used to test memory. (A) MTM tested 3 h after 1x conditioning in control and Letm1 RNAi #2 flies. Group differences were assessed with a one-way ANOVA (n = 12 per group). ANOVA: F(2,24) = 0.5172, P = 0.6027 (B) LTM tested 24 h after 1x conditioning in control and Letm1 RNAi #2 flies, is increased LTM in Letm1 KD flies. (one-way ANOVA F(2,33) = 6.565, P = 0.0040, n = 12,12,12, Tukey’s post-test Control 1 vs Letm1 KD, **P = 0.0045; Control 2 vs Letm1 KD, *P = 0.0271. Control 1 vs Control 2, n.s., P = 0.7579). (C) No RNAi expression was induced and LTM was tested 24 h after 1x conditioning in control and Letm1 RNAi #1 or #2 lines and no significant differences were detected. P values were determined using one-way ANOVA (left: Letm1 RNAi #1: F(2,51) = 0.2267, P = 0.7670, n = 18,18,18; right Letm1 RNAi #2: F(2,33) = 0.1892, P = 0.8285, n = 12,12,12) (D) Avoidance of control and Letm1 RNAi #1 or #2 lines to the odors used for olfactory conditioning shows no difference in odor avoidance. P values were determined using one-way ANOVA (top panel: Letm1 RNAi #1: Octanol: F(2,33) = 0.8653, P = 0.4303, n = 12,12,12; Methyl-cyclohexanol: F(2,33) = 2.749, P = 0.0787, n = 12,12,12; bottom panel: Letm1 RNAi #2: Octanol: F(2,33) = 2.859, P = 0.0716, n = 12,12,12; Methyl-cyclohexanol: F(2,33) = 0.2520, P = 0.7787, n = 12,12,12) (E) Avoidance of control and Letm1 RNAi #1 or #2 lines to the shock stimulus shows no difference in shock avoidance. P values were determined using one-way ANOVA (left: Letm1 RNAi #1: F(2,33) = 0.5629, P = 0.5749, n = 12,12,12; right Letm1 RNAi #2: F(2,33) = 0.7137, P = 0.4972, n = 12,12,12) (F) Testing for LTM formation 24 hours after a 5x spaced training in control and Letm1 RNAi #1 or #2 flies show normal LTM formation in all conditions. P values were determined using one-way ANOVA (left: Letm1 RNAi #1: F(2,51) = 0.2275, P = 0.7973, n = 18,18,18; right Letm1 RNAi #2: F(2,33) = 0.9020, P = 0.4155, n = 12,12,12) For graph, A-F: Data are represented as mean ± SEM and genotypes are shown in the top legend. (G) Letm1 RNA levels measured by qPCR from fly brains, normalized to α-Tub84B as reference gene. Note that knock down is underestimated as RNAi is only expressed in MB neurons and not in the entire brain. P values were determined using a one-way ANOVA followed by Tukey’s multiple comparison test versus control with n = 4 for control, n = 4 for Letm1 RNAi #1 and n = 5 for Letm1 RNAi #2, one-way ANOVA: F(2,10) = 8.838, P = 0.0.0062, Tukey’s multiple comparison P values: Letm1 RNAi #1 vs control, *P = 0.0084; Letm1 RNAi #2 vs control, *P = 0.0147. Data are represented as mean ± SEM.
