Cytochromes P4502C1/2 and P450 2E1 are retained in the endoplasmic reticulum membrane by different mechanisms

Cytochrome P450 (P450) 2C1/2 contains redundant endoplasmic reticulum (ER) retention signals and is excluded from the recycling pathway. Other P450s, such as P450 2E1, have been detected in the plasma membrane and Golgi appar atus. To examine whether the mechanisms of ER retention might differ for P4 50 2C1/2 and P450 2E1, chimeras of green flourescent protein and the full-l ength proteins, N-terminal signal/anchor sequences, or the cytoplasmic cata lytic domains from these proteins have been expressed in COS1 cells. Chimer as with either the N-terminal signal/anchor sequence or the cytoplasmic dom ain of P450 2C1/2 were retained in the ER and the distribution was not alte red by treatment with nocodazole. A chimera with full-length P450 2E1 was l ocated in the ER, but in contrast to P450 2C1/2, treatment with no codazole resulted in redistribution to a vesicular pattern, which suggested that th is protein was retained in the ER by a retrieval mechanism. In support of t his possibility, the P450 2E1 chimera, but not the P450 2C1/2 chimera, was included in transport vesicles generated in an in vitro budding assay. A ch imera with only the N-terminal signal/anchor sequence of P450 2E1 fused to green fluorescent protein was located in the ER and nocodazole treatment al tered its distribution, whereas a chimera with only the cytoplasmic domain of P450 2E1 was not efficiently retained in the ER and accumulated primaril y in the Gels region. These results demonstrate that the mechanisms for ret ention in the ER of two closely related members of the P450 superfamily are different and that the N-terminal signal/anchor sequence contains the domi nant retention signal. (C) 2000 Academic Press.