Skip to main content
. Author manuscript; available in PMC: 2015 Dec 22.
Published in final edited form as: Dev Cell. 2014 Dec 22;31(6):677–689. doi: 10.1016/j.devcel.2014.11.022

Figure 4. Deletion of APC disrupts MT severing in developing interneurons.

Figure 4

(A–H) MTs in E14.5 control and APC mutant interneurons (INs) and projection neurons (PNs) are immunolabeled with anti-acetylated tubulin antibodies and imaged using a super-resolution microscope. (A–F) APC deficient INs have more breakpoints (arrow heads) than controls, indicating increased MT severing. (G–H) Such differences in MT severing is not evident between control (G) and APC deficient projection neurons (H). (I) Quantification of MT severing in control and APC mutant INs and PNs. Data shown are mean ± SEM; n=40 cells from 4 brains per group; *, significant when compared with controls at p<0.05 (Student’s t test). (J) Comparisons of MT severing protein levels in INs and PNs isolated from the GE and D. Cx of respective control, APCLox/Lox; Dlx5/6-Cre, or APCLox/Lox; Nex-Cre mice. Immunoblot analysis indicates increased p60-katanin in APC deficient INs (arrowhead), but not in PNs. No changes in other MT severing protein levels are evident. (K) Densitometric quantification of p60-katanin levels in control and APC mutants. Data shown are mean ± SEM; n=4 brains per group; *, p<0.05 (Student’s t test). (L–M) Immunolabeling of E14.5 interneurons from APCLox/+; Dlx5/6-Cre (L) and APCLox/Lox; Dlx5/6-Cre (M) GE shows increased expression of p60-katanin in APC deficient interneurons. (N) Serine phosphorylated and ubiquitylated-p60-katanin (arrowhead) are reduced in APC deficient INs. Immunoprecipitated p60-katanin are immunoblotted with anti-phosphoserine and anti-ubiquitin antibodies, respectively. (O) Densitometric quantification of phosphoserine-p60-katanin levels in control and APC mutants. Data shown are mean ± SEM; n=3 brains per group; *, p<0.05 (Student’s t test). (P) APC affects the stability of p60Katanin. Control and APC mutant interneurons were treated with 5μg/ml protein synthesis inhibitor cyclohexamide (CHX) for 3 and 6 hours. APC deletion led to increased stability of p60Katanin. C, control; M, mutant [APC deficient]; IN, interneuron-specific; PN, projection neuron-specific; GE, ganglionic eminence; D.CX, dorsal cortex. Scale bar= 3.1 μm (A–D, G–H), 2.8 μm (E and F), 15 μm (L and M).