Abstract
In this study, we investigated the functional role of early growth response-1 (Egr1 gene) in the regulation of radiation-induced clonogenic inhibition and apoptosis in p53 wild-type and mutant prostate cancer cells 22Rv1 and DU145, respectively. 22Rv1 cells were more sensitive to irradiation compared with DU145 cells, and the sensitivity was enhanced by overexpression of EGR-1 in both cells. Dominant-negative EGR-1 mutant (dnEGR-1) or repressor of EGR-1, NGFIA binding protein 1 (NAB1), increased radioresistance of these cells. Significant activation of caspases 3 and 9 and Bcl2-associated X (Bax) with increased poly(ADP-ribose) polymerase (PARP) cleavage and cytochrome c release was observed in radiation-exposed EGR-1 overexpressing cells. Gel shift analysis and chloramphenicol acetyl transferase (CAT) reporter assays indicate that EGR-1 transactivates the promoter of the Bax gene. Interaction of EGR-1 and Yes kinase-associated protein 1 (YAP-1) through the WW domain of YAP-1 enhances the transcriptional activity of EGR-1 on the Bax promoter as shown by chromatin immunoprecipitation and reporter assays. Irradiation of PC3 cell xenografts that were treated with adenoviral EGR-1 showed significant regression in tumor volume. These findings establish the radiation-induced pro-apoptotic action of EGR-1, in a p53-independent manner, by directly transactivating Bax, and prove that alters the B-cell CLL/lymphoma 2 (Bcl-2)/Bax ratio as one of the mechanisms resulting in significant activation of caspases, leading to cell death through the novel interaction of EGR-1 with YAP-1.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Adaptor Proteins, Signal Transducing / genetics
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Adaptor Proteins, Signal Transducing / metabolism*
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Animals
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Apoptosis / physiology
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Apoptosis / radiation effects*
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Blotting, Western
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Caspase 3 / metabolism
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Chloramphenicol O-Acetyltransferase / metabolism
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Colony-Forming Units Assay
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Cytochromes c / metabolism
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Early Growth Response Protein 1 / genetics
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Early Growth Response Protein 1 / metabolism*
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Electrophoretic Mobility Shift Assay
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Humans
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Immunoprecipitation
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Male
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Mice
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Mice, Inbred BALB C
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Mice, Nude
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Phosphoproteins / genetics
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Phosphoproteins / metabolism*
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Poly(ADP-ribose) Polymerases / metabolism
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Promoter Regions, Genetic
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Prostatic Neoplasms / genetics
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Prostatic Neoplasms / metabolism*
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Prostatic Neoplasms / pathology*
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Protein Array Analysis
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Proto-Oncogene Proteins c-bcl-2 / metabolism
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RNA, Messenger / genetics
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RNA, Messenger / metabolism
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Radiation Tolerance
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Repressor Proteins / metabolism
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Reverse Transcriptase Polymerase Chain Reaction
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Transcription Factors
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Transcriptional Activation
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Transfection
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Transplantation, Heterologous
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Tumor Cells, Cultured
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Up-Regulation
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X-Rays
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YAP-Signaling Proteins
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bcl-2-Associated X Protein / genetics
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bcl-2-Associated X Protein / metabolism*
Substances
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Adaptor Proteins, Signal Transducing
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BAX protein, human
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EGR1 protein, human
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Early Growth Response Protein 1
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NAB1 protein, human
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Phosphoproteins
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Proto-Oncogene Proteins c-bcl-2
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RNA, Messenger
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Repressor Proteins
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Transcription Factors
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YAP-Signaling Proteins
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YAP1 protein, human
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bcl-2-Associated X Protein
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Cytochromes c
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Chloramphenicol O-Acetyltransferase
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Poly(ADP-ribose) Polymerases
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Caspase 3