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. 2023 Nov 22;44(13):e0374232023. doi: 10.1523/JNEUROSCI.0374-23.2023

Figure 1.

Figure 1.

Katanin inhibition affects MT growth in dendrites of premature but not mature neurons. A, Immunostaining of endogenous p60 katanin in dendrites of cultured hippocampal neurons at DIV 13–17. n = 3 experiments. B, Anti-p60 katanin immunoelectron microscopy with diaminobenzidine (DAB) showing MTs from hippocampal neurons. C,D, Anti-GFP immunoelectron microscopy with diaminobenzidine (DAB) showing MTs from hippocampal neurons transfected with either GFP-p60 katanin (C) or GFP-p60DEID (D). E, Overview image of GFP-p60 katanin (green) and endogenous p60 katanin (red). F, Magnifications of boxed region in E. G, Quantification of GFP-p60 katanin/p60 (endogenous) katanin colocalization levels. Mean ± S.E.M. = 91.16 ± 1.006%. Three independent experiments. H,I, EB3-Tomato time-lapse imaging in dendrites of DIV4 hippocampal neurons. Right: diagrams of EB3 tracks for GFP in H and for GFP-p60DEID in I. Neurons express EB3-Tomato (red) with either GFP or GFP-p60DEID (green). The green channel is not shown. J, Quantification of EB3 growth velocity (µm/s). GFP: 0.24 ± 0.01, GFP-p60DEID: 0.21 ± 0.01. Mann–Whitney U test p = 0.0152. GFP: n = 48 comets; GFP-p60DEID: n = 24 comets, three independent experiments. K,L, EB3-Tomato time-lapse imaging in dendrites of DIV14 hippocampal neurons. Right: diagrams of EB3 tracks for GFP in K and for GFP-p60DEID in L. Neurons express EB3-Tomato (red) with either GFP or GFP-p60DEID (green). The green channel is not shown. M, Quantification of K and L shows comparable growth velocity among conditions. GFP: 0.2054 + 0.01584 μm/s, GFP-p60DEID: 0.2387 + 0.01736 μm/s, Unpaired t test, two-tailed, p = 0.1712, (GFP: n = 12 ROIs, 503 comets); (GFP-p60DEID: n = 12 ROIs, 340 comets), four independent experiments.