[Regulation of NF-kappaB/P65 by MDM2 in acute lymphoblastic leukemia in childhood].
OBJECTIVE
MDM2 is considered a proto-oncogene due to its ability to inhibit P53 tumor-suppressor function. But, evidence showed that MDM2 might have a P53-independent role in tumorigenesis. MDM2 is over-expressed in human sarcoma and carcinoma. Recent studies showed that MDM2 might act as a transcriptional factor to modulate expressions of other genes involved in cell cycle regulation and transformation. In the present study, the investigators hypothesized that MDM2 directly affected NF-kappaB expression and function in a P53-independent manner.


METHODS
MDM2 was transfected to acute lymphoblastic leukemia (ALL) line EU-4 cells lacking P53 expression and expressing very low levels of MDM2. MDM2 and P65 expression in mRNA level and protein level were detected by Western blot and Northern blot after transfection. Since the expression of E-selectin is P65 dependent, E-selectin promoter-CAT construct and P65 and MDM2 expression plasmids were co-transfected to EU-4 cells. CAT activation was determined with ELISA. The effect of adriamycin (ADM) at the concentrations of 15 micro g/ml, 7.5 micro g/ml, 5 micro g/ml and 1 micro g/ml on MDM2-transfected EU-4 cells and the parent cells was detected by MTT assay.


RESULTS
The results showed that MDM2 up-regulated P65 expression at both mRNA and protein levels, and MDM2 increased P65-mediated transactivation of E-selectin promoter. Without P65, MDM2 had no effect on the transactivation of E-selectin. Moreover, MDM2 antisense could not change the transactivation of E-selectin. MTT results showed that the survival rate of MDM2 transfected EU-4 cells was higher than that of parental cells. The results suggested that MDM2 transfection increase drug resistance of EU-4 cells to ADM compared with parent cells.


CONCLUSIONS
(1) MDM2 up-regulated transcriptionally P65 expression. (2) MDM2 increased drug resistance of leukemia cells to ADM. (3) MDM2 elevated NF-kappaB activity in a P53-independent manner in childhood lymphoblastic leukemia cell line.
Hu
Qun
Q
Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Zhou
Mu-xiang
MX
Liu
Shuang-you
SY
Zhang
Liu-qing
LQ
Liu
Ai-quo
AQ
Guo
Yi-jie
YJ
Song
Yu
Y
D000903:Antibiotics, Antineoplastic / Q000627:therapeutic use; D015152:Blotting, Northern; D015153:Blotting, Western; D045744:Cell Line, Tumor / Q000187:drug effects; D002648:Child; D004317:Doxorubicin / Q000627:therapeutic use; D019008:Drug Resistance, Neoplasm / Q000502:physiology; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000502:physiology*; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000502:physiology*; D051736:Proto-Oncogene Proteins c-mdm2; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051996:Transcription Factor RelA; D014162:Transfection / Q000379:methods
D000903:Antibiotics, Antineoplastic; C000715470:MAS1 protein, human; D016328:NF-kappa B; D009687:Nuclear Proteins; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D051996:Transcription Factor RelA; D004317:Doxorubicin; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2