Distinct subcellular localization of transiently expressed types 1 and 2 iodothyronine deiodinases as determined by immunofluorescence confocal microscopy

We compared the subcellular localization of FLAG-epitope tagged Types 1 and 2 deiodinases (D1 and D2) transiently expressed in human embryonic kidney (HEK-293) and mouse neuroblastoma (NB2A) cells. D2 is an integral membrane protein based on resistance to extraction at pH 11 with the NH2 terminus in the endoplasmic reticulum (ER). Immunofluorescence confocal microscopy usi ng anti-FLAG and anti-GRP78/BiP antibodies showed the FLAG-DI signal was fo und in the periphery of the cells and not co-locatized with the ER specific marker GRP78/BiP. On the other hand, FLAG-D2 protein was found in the ER c o-localized with the GRP78/BiP protein. These differential distribution pat terns indicate subcellular sorting of D1 and D2 is determined by intrinsic protein sequence and can explain the ready access of D2-generated T3 to the nucleus.