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. 2018 Feb 22;172(5):1063–1078.e19. doi: 10.1016/j.cell.2018.01.031

Figure 1.

Figure 1

CCP1 Promotes the Saltatory Migration of cINs

(A) Immunodetection of polyglutamate side chains (GT335 antibody) on cINs explants. cINs express Cre-GFP and nuclei are counterstained with DAPI. The white arrow points the leading process. Scale bar, 10 μm.

(B) Normalized expression levels of CCP and TTLL mRNAs in E13.5 WT cINs, n = 3 embryos per group from 3 females.

(C) ISH of Ccp1 on a coronal section of E13.5.

(D) Subcellular distribution of CCP1 (red) in WT migrating cINs. Scale bars, 5 μm (top), 2 μm (bottom).

(E) Normalized expression levels of Ccp1 exons 20 and 21 in FACS-purified cINs from 3 E13.5 CCP1 WT or cKO mouse embryos.

(F) Glutamylation levels (PolyE or GT335 antibodies) on tubulin extracted from GEs of CCP1 WT and cKO embryos at E13.5.

(G) PolyE and GT335 immunoreactivity normalized on tubulin and actin and expressed as percentage of control, n = 5–7 embryos per group, non-parametric t test, p < 0.05.

(H) Immunodetection of polyglutamate stretches (PolyE antibody) on cIN explants. INs from CCP1 WT and cKO express Cre-GFP and Dapi+ nuclei. Scale bar, 10 μm.

(I) Representative time series of time-lapse acquisition of migrating cINs from CCP1 WT and cKO embryos. cINs express GFP and secondary neurites are pointed by green arrows.

(J) Neurites life duration, n = 26–127 cells per group from at least 3 independent cultures, parametric t test, p < 0.001.

(K) Tubulin acetylation levels on protein extracts from E13.5. CCP1 WT and cKO GEs.

(L) Normalized acetylation immunoreactivity normalized, n = 4 embryos per group, p < 0.05.

(M) Nuclear displacement of representative migrating cINs from CCP1 WT and cKO E13.5 embryos. The traveled distance between two time points is plotted and every displacement above 5 μm (gray dotted line) is a nucleokinesis. Red arrows indicate sliding movement in CCP1 cKO. A nuclear pause corresponds to no displacement between two time points.

(N–P) Average speed (N), amplitude (O) of nucleokinesis, and time of nuclear pause (P) of CCP1 WT and cKO cINs measured in E13.5 organotypic slice culture (see also Figure S1; n = 45–78 cells in at least 3 embryos from at least 3 females; p < 0.01). All graphs contain bars representing the SEM.