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Figure 5


Fig. 5. Bcl-2 interacts with EGL-1. (A) The two-hybrid yeast strain HF7c was transformed with the indicated plasmids or empty vector controls. Transformants were spotted (as described in the legend to Fig. 1) onto minimal medium either containing histidine (growth confirms the presence of the plasmids) or lacking histidine (growth indicates interaction between the prey and bait proteins). (B) In vitro translated [35S]methionine-labelled FLAG-Tab-1, [35S]methionine-labelled CED-9 or [35S]methionine-labelled Bcl-2 were incubated with bacterial lysates containing recombinant HIS-tagged p35, HIS-tagged EGL-1 or HIS-tagged CED-4. The protein complexes were then bound to Ni-NTA beads and subjected to SDS-PAGE. [35S]-labelled proteins, bound through the recombinant proteins to the beads, were detected (upper panel). The complexes were also immunoblotted to detect the HIS-tagged proteins (lower panel). The arrow denotes full-length CED-4-6HIS. The asterisk indicates a smaller protein that might be a breakdown product. (C) Yeast were transformed with the indicated plasmids and spotted onto either repressing or inducing media as described in Fig. 1. Immunoblotting was used to detect Bax, Bcl-2, Bim and myc-EGL-1. Coomassie staining illustrates protein loading.





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