Endoplasmic reticulum localization of Sec12p is achieved by two mechanisms: Rer1p-dependent retrieval that requires the transmembrane domain and Rer1p-independent retention that involves the cytoplasmic domain

To determine which region of Sec12p is important for the ER localization, we planned to construct chimeric proteins between Sec12p and an appropriate passenger protein that is innocent for its destination. As such a passenger protein, we chose Dap2p (dipeptidyl aminopeptidase B), a vacuolar membrane protein. Dap2p is a transmembrane protein with the same topology as Sec12p (type II) but is transported to the vacuole quickly after biosynthesis (Roberts et al., 1989) . It has been shown that Dap2p has no particular localization signal in itself and thus is transported to the vacuole by default (Roberts et al., 1992) . This gives us a nice opportunity to identify the ER localization signal(s) of Sec12p by examining which chimeric constructs are localized to the ER. We dissected Sec12p and Dap2p into three parts: the NH2-terminal cytoplasmic domain, the TMD, and the COOH-terminal lumenal domain by introducing appropriate restriction sites in their corresponding genes. For convenience, we refer to the constructs with three letters composed of S and D. S stands for Sec12p and D is for Dap2p. The first letter indicates the NH2-terminal domain, the second is for the TMD, and the third is for the COOH-terminal domain. For example, SDD refers to the N (Sec12p)-TMD (Dap2p)-C (Dap2p) chimera. All the combinations of the three parts of the two proteins were made and named by this nomenclature (Fig. 1) . They were all placed under the SEC12 promoter.