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Figure 4


Fig. 4. Expression of Dapper1 abolishes the ability of WNT3a, as well as the ability of expression of Q205L G{alpha}o to stimulate JNK activation. F9 cells expressing rFZ1 were either transfected (1 µg/well in a 12-well plate) with an expression vector harboring Myc-tagged mouse Dapper1 (A) or co-transfected with Myc-Dapper1 and Q205L G{alpha}o (B) for 24 hours, and JNK activity was determined, as described in the Materials and Methods. The expression of Dapper1 was monitored by immunoblotting with antibodies against Myc. (C) F9 cells were either transfected (1 µg/well in a 6-well plate) with individual HA-DVL-GFP2 or co-transfected with HA-DVL-GFP2 and MycDapper1 (1 µg/well of each plasmid) for 24 hours followed by cell lysis and affinity pull-downs with anti-HA-sepharose beads. The interaction of Dapper1 with individual isoforms of DVL was made visible by probing the blots with the antibody against Myc. The expression of the exogenous DVL-GFP2 was established by stripping the Myc blot and probing with an antibody against HA. The upper panel displays mean values±s.e.m. obtained from three independent experiments; the lower panel displays the corresponding representative blots. The blot of the DVL-Dapper1 interaction is representative of two independent experiments that proved highly reproducible. *P<0.05; **P<0.01 versus –WNT3a control; #, P<0.05; ##, P<0.01 versus +WNT3a control. The expression of DVL3 appears low in the absence of Dapper 1, but this was not a consistent observation.





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