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Figure 3. The beta-cat-binding site and the TM are essential for Xcad-11 function. (A) Xcad-11 constructs for reconstitution. (S) Signal peptide (green); (EC) extracellular domains (orange); TM (red); (p120/beta-cat) p120-binding site (blue) or beta-cat-binding site (dark red); intracellular domain (yellow). (B) Whole-mount ISH. Lateral view of CNC at stage 26, analyzed by expression of twist. (Left column)Noninjected side (nis). (Right column) Injected side (is). Embryos injected with Xcad-11MO (2 pmol) together with 50 pg of DELTA-e or DELTA-e/DELTA-p120 showed twist signal in pharyngeal pouches, while coinjection of 75 pg of DELTA-c, 50 pg DELTA-e/DELTA-beta-cat, DELTA-e/DELTA-TM, or DELTA-e/DELTA-IMD did not. (C) Statistics for reconstitution. In the case of Xcad-11MO injection, only 28% of embryos showed normal migration. Coinjection of 75
pg of FLXcad-11 resulted in 82%, 75 pg of DELTA-e in 88%, and 75 pg of DELTA-e/DELTA-p120 in 82% of embryos with CNC present in pharyngeal pouches; in the case of DELTA-c, DELTA-e/DELTA-beta-cat, DELTA-e/DELTA-TM, and DELTA-e/DELTA-IMD coinjection, only 30%, 27%, 28%, and 52%, respectively. (D) CNC explants. Coinjection of FLXcad-11, DELTA-e, or DELTA-e/DELTA-p120 with Xcad-11MO restored formation of lamellipodia and filopodia in CNC; DELTA-e/DELTA-beta-cat coinjection showed blebbing. Bar, 10 mm. |
