Localization of PML (NLSˉ) Protein in NB4 Cells and Transplanted Tumors in Nude Mice

Abstract: To verify the existence and location of PML (NLS–) protein after PML-RARα cleaved by neutrophil elastase (NE)， the plasmid pCMV-HA-NE was electroporated into NB4 cells and NE protein was verified by Western blot. We extracted the cytoplasm protein of NB4-HA-NE cells and detected the PML (NLS–) protein by Western blot. Immunofluorescence assay and confocal laser microscopy were performed to localize the PML (NLS–) protein in the cytoplasm of NB4-HA-NE cells. Then， NB4， K562 and NB4 HA-NE cells were expressed in xenograft of nude mice. Western blot and immunohistochemistry were done to detect PML (NLS–) protein in the cytoplasm of the NB4-HA-NE cells. The results indicated that NE protein was detected in the NB4 -HA-NE cells and PML (NLS–) protein was verified in the NB4-HA-NE cells by Western blot， which demonstrated that PML-RARα was cut successfully by NE. Immunofluorescence assay and confocal laser microscopy showed that the PML (NLS–) protein was localized in the cytoplasm of NB4-HA-NE cells. Meanwhile， PML (NLS–) protein in nude mice was expressed and localized in the cytoplasm of NB4-HA-NE cells manifested by Western blot and immunohistochemistry， while PML protein was almost localized in the nucleus of NB4 and K562 cells in nude mice. In summary， the pCMV-HA-NE plasmid was successfully electransfered into NB4 cells and PML (NLS–) protein was verified to be in the cytoplasm of NB4-HA-NE cells with the use of Western blot， immunofluorescence， confocal laser microscopy and immunohistochemical techniques， which could provide new evidence for early diagnosis and treatment of acute promyelocytic leukemia.

CSTR: 32200.14.cjcb.2014.05.0004