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. Author manuscript; available in PMC: 2009 Jul 16.
Published in final edited form as: Nat Neurosci. 2004 Sep 19;7(10):1059–1069. doi: 10.1038/nn1317

Figure 6.

Figure 6

Several protein-binding sites on FAK are required to control axonal branching as assessed by coexpression experiments with EGFP-Cre. (a–c) FAK re-expression reverses Cre-mediated fak deletion phenotype into a wild-type pattern in mice hippocampal neurons. Fluorescence images and drawings of neurons transfected with pCDNA3, EGFP-Cre and EGFP-Cre plus wild-type FAK. (d–f) Fluorescence images and representative drawings taken from coexpression experiments of FAK-defective variants and Cre plasmids. (d) Myc-FAKY397F (e) Myc-FAK878A (f) Myc-FAK1034S. All neurons were transfected at 3 DIV and recorded at 6 DIV. High-magnification images in color show colocalization (yellow or white) of DsRed, EGFP-Cre and Myc. (g) Quantification of the total number of branches per neuron. One asterisk denotes significant difference from control; two asterisks denote significant difference from Cre experiments. P < 0.01–0.0001 (control, 6.21 ± 1.07; Cre, 30.53 ± 2.07; FAKwt, 8.07 ± 0.68; FAKY397F, 18.88 ± 2.39; FAKP878A, 23.50 ± 2.80; FAKL1034S, 19.66 ± 1.76; n = 93 neurons from three independent experiments in g,h,j,k). (h) Quantification of the number of increasing branch order per neuron. P < 0.05 to P < 0.0001 versus control. (i) Position of point mutations and protein domains in FAK. FAT, focal adhesion targeting region, PR1 and PR2, proline-rich regions. (j) Quantification of the total neurite length per neuron expressed in um. P < 0.05–0.0001 versus control (control, 1,705.71 ± 265.34; Cre, 4,748.76 ± 402.85; FAKwt (3,061.96 ± 335.56; FAKY397F, 4,120.26 ± 543.10; FAKP878A, 6279.38 ± 922.08; FAKL1034S, 4,018.83 ± 347.92). (k) Quantification of the total number of branches per µm of neurite. P < 0.05–0.0001 versus control (control, 0.0034 ± 0.0004; Cre, 0.0068 ± 0.0006; FAKwt, 0.0032 ± 0.0003; FAKY397F, 0.005 ± 0.0005; FAKP878A, 0.0045 ± 0.0005; FAKL1034S, 0.0052 ± 0.0004) Note that ectopic expression of FAK rescues both the branching and the overall increase in axon length phenotypes in FAK-deficient neurons. This result is consistent with our observations that loss of FAK resulted in reduced axonal growth speed, and supports the idea that FAK controls axonal dynamics by participating both in axonal growth and branch formation independently. Note that, as shown in Figure 4, most branching neurites at these stages are developing axons (∼95%), although a minor percentage of them could be dendrites. Data are the mean ± s.e.m. Scale bars, 200 µm (a–g).