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. Author manuscript; available in PMC: 2012 Sep 16.
Published in final edited form as: Neuron. 2007 May 3;54(3):429–445. doi: 10.1016/j.neuron.2007.04.016

Figure 10. Defective localization of WAVE1 to protrusive membrane edges in Nap1 mutant cells.

Figure 10

(A) Wild type and mutant telencephalic neuroepithelial cells were labeled with phalloidin (green) and anti-WAVE1 antibodies (red). In wt cells, WAVE1 predominantly localizes to lamellipodial membrane edges (arrowhead, A), whereas in Nap1 mutant cells, WAVE1 localization to lamellipodial protrusion is mostly absent. (B) Quantification of cells with WAVE1 localization on membrane edges indicates a significant deficit in Nap1 mutant or Nap1 deficient cells. Expression of C-terminal deleted Nap1 or Nap1 shRNA in wild type neuroepithelial cells leads to disrupted WAVE1 localization. This deficit can be rescued by expression of full length Nap1. (C, D) Tracking of WAVE1 localization in Nap1 disrupted cells. Wild type, Nap1 mutant, or Nap1 shRNA expressing cells were transfected with WAVE1- Kaede (green). After localized photoconversion with a UV laser , time-lapse images of photoconverted WAVE1- Kaeda (red) were obtained. In wild type cells WAVE1, actively moved towards the protrusive edges of the cells (arrows [wt panels], C; AVI movie file #6). In contrast WAVE1 movement is significantly retarded in Nap1 mutants (arrows [Nap1 mutant panels], C; AVI movie file #7) and in Nap1 knockdown cells (arrows [Nap1 shRNA panels], C; AVI movie file #8). Time after photoconversion is indicated in minutes. (D) Measurement of relative fluorescent intensities of WAVE1-green and WAVE1-red in defined areas within the photoconverted spots indicate that in wt cells both types of WAVE1 trafficked normally, whereas in Nap1 mutants or Nap1 shRNA expressing cells, movement of WAVE1 is highly restricted. Also see supplementary movie files #6, 7, and 8. Data shown are mean ±SEM; asterisk, significant when compared with controls at p<0.01 (Student's t test).

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