Figure 5: Nuclear expression of YAP is regulated by nerve elongation

(a) Schematic of the technique we used to mechanically distract the adult (11 to 13 weeks old) mouse femur and to artificially extend the sciatic nerve (blue) that is attached to the spinal cord (device not to scale, see Online Methods for more details). (b) An entire extended sciatic nerve (E) and the non-extended contralateral nerve (NE) of the same mouse are shown after femoral distraction using reflected white light. The extended sciatic nerve is 30% longer than the contralateral nerve. (c) Representative mSCs infected with control (Cont sh) or Crb3 (Crb3 sh) shRNA virus in an extended (E) or a non-extended nerve (NE). (d) Quantification of internodal lengths of cells infected with control, YAP shRNA or Crb3 shRNA viruses in extended (E) or non-extended nerves (NE). Between 106 and 171 cells (from three animals) were measured for each condition. (e) Nuclear YAP (red) immunostaining in mSC nuclei (arrows) is higher in artificially extended nerves (E, right panel) than in non-extended nerves (NE, left panel). (f) Quantification of YAP immunostaining in nuclei of mSCs infected with control or Crb3 shRNA viruses in extended (E) or non-extended nerves (NE). Nuclei quantified: 52 (NE Cont sh), 131 (E Cont sh), 19 (E Crb3 sh). Three animals for each condition. (g) In an extended nerve, the nucleus (arrows) of a cell expressing Crb3 shRNA (red) has more YAP (green) than the nucleus (DAPI, blue) of a nearby non-infected cell (NI). a.u., arbitrary unit. Statistical two-tailed Student’s t-tests: *P<0.05; **P<0.01; ***P<0.001. Scale bars, 1 mm (b), 100 μm (c), 20 μm (e,g). Error bars show s.e.m.