Fig. 8: BRAWNIN is necessary and sufficient downstream of NUAK1 for cortical axons branching. | Nature Communications

Fig. 8: BRAWNIN is necessary and sufficient downstream of NUAK1 for cortical axons branching.

From: The AMPK-related kinase NUAK1 controls cortical axons branching by locally modulating mitochondrial metabolic functions

Fig. 8

A, B Representative images of mVenus expressing cortical neurons electroporated with an shRNA against Brawnin. Red stars indicate collateral branches. C, D Quantification of axon length and collateral branches in the indicated conditions. Statistical test: two-tailed Mann-Whitney. N(control)= 295, N(shRNA)= 277 neurons out of 3 independent experiments. Data: box-and-whisker plots. The horizontal line inside the box is the median value. The endpoints of the box represent the 25th−75th percentiles, with whiskers at minimum and maximum values. E Effect of Brawnin shRNA on the mitochondrial membrane potential (ΔΨm). Each point represents average values for a given neuron. N(control)= 51, N(shRNA)= 53 out of 3 distinct cell cultures. Data: average ± SEM. Statistical test: two-tailed Mann-Whitney. F–H Measurement of oxygen consumption rate in 5DIV cortical neurons transduced with shRNA against Brawnin. OCR values are normalized to cell number. Basal and maximal respiration values are represented in (G) and (H) respectively. Data: average ± SEM. N(control) = 9, N(shRNA) = 9 different embryos out of 3 independent litters. Statistical test: two-tailed Mann-Whitney. I–L Representative images of cortical neurons electroporated with an shRNA against Nuak1 and overexpressing BRAWNIN. Red stars indicate collateral branches. M, N Axon length and collateral branches of 5DIV neurons in the indicated conditions. Statistical test: Kruskal-Wallis test with Dunn’s post-test. N(Control)= 295, N(NUAK1sh)= 302, N(Control+BRAWNIN) = 277, N(NUAK1sh+BRAWNIN) = 316 neurons out of 3 independent experiments. Data: box-and-whisker plots. The horizontal line inside the box is the median value. The endpoints of the box represent the 25th−75th percentiles, with whiskers at minimum and maximum values. O Measurements of the inner mitochondrial membrane potential (IMM) in the indicated conditions. Data: average ± 95% CI. Statistical test: Kruskal-Wallis test with Dunn’s post-test (each condition compared to the control condition). Data: average ± 95% CI. N(control)= 97, N(NUAK1 KO) = 26, N(control+BRAWNIN) = 45, N(NUAK1 KO+BRAWNIN) = 36 neurons out of 4 independent experiments. ns not significant, *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001. Source data are provided as a Source Data file.

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