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 examine whether SDDm is in fact retained in the ER, we performed immunofluorescence microscopy. As shown in Fig. 5 C, the anti-Dap2p antibody again stained the ER in the Adap2 cells expressing SDDm. This led us to conclude that the cytoplasmic domain of Secl2p also has an ability to localize the protein to the ER. The efficiency of the ER localization attained by the cytoplasmic domain appears to be lower than that of the TMD because the secretion of a-factor from the Mfalp fusion is not completely prevented by this domain alone. The effect of the cytoplasmic domain does not require the presence of Rerlp. This raises an intriguing possibility that the retention by the cytoplasmic domain may be due to the static mechanism at the exit from the ER. It should also be noted that, in the absence of Rerlp, the cytoplasmic domain is not the only region that causes the retention. As seen in Table II , the Secl2p TMD also has an effect in the Arerl cells (see DSSm and DSDm). This Rerlp-independent mechanism may involve both the cytoplasmic and TMD regions.