Estrogen-mediated downregulation of CD24 in breast cancer cells.
We have previously reported on the relevance of the prevalence of CD44(+)/CD24(-/low) cells in primary breast tumors. To study regulation of CD24, we queried a number of publicly available expression array studies in breast cancer cells and found that CD24 was downregulated upon estrogen treatment. We confirmed this estrogen-mediated repression of CD24 mRNA by quantitative real-time PCR in MCF7, T47D and ZR75-1 cells. Repression was also seen at the protein level as measured by flow cytometry. CD24 was not downregulated in the ER alpha negative MDA-MB-231 cells suggesting that ER alpha was necessary. This was further confirmed by ER alpha silencing in MCF7 cells resulting in increased CD24 levels and by reintroduction of ER alpha into C4-12 cells resulting in decreased CD24 levels. Estrogen treatment did not alter half-life of CD24 mRNA and new protein synthesis was not essential for repression, suggesting a primary transcriptional effect. Histone deacetylase inhibition by Trichostatin A completely abolished the repression, but decrease of the ER alpha corepressors NCoR, LCoR, RIP140, silencing mediator of retinoid and thyroid hormone receptors, SAFB1 and SAFB2 by siRNA or overexpression of SAFB2, NCoR and silencing mediator of retinoid and thyroid hormone receptors had no effect. In silico promoter analyses led to the identification of two estrogen responsive elements in the CD24 promoter, one of which was able to bind ER alpha as shown by electrophoretic mobility shift assay and chromatin immunoprecipitation assay. Together, our results show that CD24 is repressed by estrogen and that this repression is a direct transcriptional effect depending on ER alpha and histone deacetylases.
Kaipparettu
Benny Abraham
BA
Division of Molecular Mechanisms of Origin and Treatment of Breast Cancer, Dr. Margarete Fischer-Bosch-Institute of Clinical Pharmacology, Stuttgart, Germany. kaippare@bcm.edu
Malik
Simeen
S
Konduri
Santhi D
SD
Liu
Wensheng
W
Rokavec
Matjaz
M
van der Kuip
Heiko
H
Hoppe
Reiner
R
Hammerich-Hille
Stephanie
S
Fritz
Peter
P
Schroth
Werner
W
Abele
Ina
I
Das
Gokul M
GM
Oesterreich
Steffi
S
Brauch
Hiltrud
H
D001943:Breast Neoplasms / Q000378:metabolism*; D051927:CD24 Antigen / Q000235:genetics* / Q000378:metabolism*; D045744:Cell Line, Tumor; D047369:Chromatin Immunoprecipitation; D015536:Down-Regulation; D024202:Electrophoretic Mobility Shift Assay; D004791:Enzyme Inhibitors / Q000494:pharmacology; D047628:Estrogen Receptor alpha / Q000378:metabolism; D004967:Estrogens / Q000378:metabolism* / Q000494:pharmacology; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D056572:Histone Deacetylase Inhibitors; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D014158:Transcription, Genetic
D051927:CD24 Antigen; C497889:CD24 protein, human; D004791:Enzyme Inhibitors; D047628:Estrogen Receptor alpha; D004967:Estrogens; D056572:Histone Deacetylase Inhibitors; D006877:Hydroxamic Acids; D012333:RNA, Messenger; C012589:trichostatin A