Mechanism of transcriptional regulation of LRP16 gene expression by 17-beta estradiol in MCF-7 human breast cancer cells.
LRP16 gene expression is induced by 17-betaestradiol (E2) via estrogen receptor alpha (ERalpha) in MCF-7 human breast cancer cells. A previous study also demonstrated that ectopic expression of LRP16 gene promoted MCF-7 cell proliferation. To explore the mechanism of hormone-induced LRP16 gene expression, the LRP16 gene promoter region (-2600 to -24 bp upstream of the LRP16 gene translation starting site) was analyzed in the present study by using different 5'-truncated constructs, and a luciferase reporter. The 5'-flanking sequence of -676 to -24 bp (pGL3-S5) was found to be E2-responsive. After exchange of the fragment from -213 to -24 bp with the TK gene proximal promoter region in pGL3-S5, E2 still induced reporter gene activity in MCF-7 and HeLa cells. Sequence analysis showed that the pGL3-S6 (-676 to -214) sequence contains two motifs that may contribute to E2-induced transactivation; namely, an estrogen-responsive element (ERE) half-site/Sp1 at -246 to -227 bp and an E-box site at -225 to -219 bp. Further deletion and mutation analysis of these two motifs indicated that both the 1/2 ERE and Sp1 binding sites were required for E2 action, while E-box deletion did not affect the luciferase activity in MCF-7 and HeLa cells. The results of gel mobility shift and chromatin immunoprecipitation assays confirmed that both ERalphaand Sp1 were required for hormone-induced transactivation, which involved both ERalphaand Sp1 directly binding to DNA. Taken together, these findings suggest that ERalphaand Sp1 play a role in activation of the human LRP16 gene promoter.
Zhao
Y-L
YL
Department of Molecular Biology, Institute of Basic Medicine, Chinese PLA General Hospital, 28 Fu Xing Road, Beijing 100853, China.
Han
W-D
WD
Li
Q
Q
Mu
Y-M
YM
Lu
X-C
XC
Yu
L
L
Song
H-J
HJ
Li
X
X
Lu
J-M
JM
Pan
C-Y
CY
D001665:Binding Sites / Q000235:genetics; D001943:Breast Neoplasms / Q000188:drug therapy / Q000235:genetics*; D002265:Carboxylic Ester Hydrolases; D004958:Estradiol / Q000494:pharmacology*; D047628:Estrogen Receptor alpha / Q000378:metabolism; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects*; D006367:HeLa Cells; D006801:Humans; D009363:Neoplasm Proteins / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D016329:Sp1 Transcription Factor / Q000378:metabolism
D047628:Estrogen Receptor alpha; D009363:Neoplasm Proteins; D016329:Sp1 Transcription Factor; D004958:Estradiol; D002265:Carboxylic Ester Hydrolases; C476832:MACROD1 protein, human