Retention of p63 in an ER-Golgi intermediate compartment depends on the presence of all three of its domains and on its ability to form oligomers

All further chimeras were generated by PCR using the overlap extension technique (Horton et al., 1989) . All chimeric constructs were precisely joined at the transitions between two domains. Final PCR products were treated as described above for the p63 cytoplasmic tail mutants, resulting in constructs pECE-DDP (cytoplasmic tail of DPPIV, transmembrane domain of DPPIV, and lumenal domain of p63), pECE-DPP (cytoplasmic tail of DPPIV, transmembrane domain of p63, and lumenal domain of p63), and pECE A24-101PDP (A24-101 cytoplasmic tail of p63, transmembrane domain of DPPIV, and lumenal domain of p63), respectively. The following templates were used for the PCR reactions: plasmids pBSK-p63 and pBSK-DPPIV (DPPIV cDNA subcloned as EcoRI fragment into the Bluescript KS-vector) to generate constructs DPP and DDP, and plasmids pBKS I)63 A2.4-101 and pECE-DDP to construct the A94-101PDP chimera. Upstream and downstream flanking primers were the same as above. Appropriate pairs of partially complementary oligonucleotides which encoded the desired fusion were chosen as internal primers.