ID4 as a tumor suppressor: mechanism of action of ID4 in prostate cancer, 2013
Evans, Ashley L.
2010-2019
Id proteins are members of basic helix-loop-helix family. However, Id proteins lack the basic binding domain, which prevents DNA binding, and thereby regulates transcription. There are four members in the Id protein family termed Idl-4. In prostate cancer the expression of Idl and Id3 is high, whereas member Id4 expression is low. Decreased expression of Id4 is due to promoter hypermethylation in prostate cancer as well as many other cancers. This observation led us to hypothesize that Id4 acts as a tumor suppressor in prostate cancer. Furthermore, evidence suggests ectopic Id4 expression in metastatic prostate cancer cell line DU145 induced cell cycle arrest, apoptosis, and senescence. In this study, we expanded on these earlier studies to demonstrate that gain of Id4 attenuates cancer phenotype whereas loss of Id4 promotes cancer phenotype in prostate cancer cell lines DU145 and LNCaP, respectively. Upon over-expression of Id4 in DU145 cells (DU145+W4), there was an increase in apoptosis, due to decreased mitochondrial membrane potential (MMP) and increased expression of pro-apoptotic markers (PUMA, BAX, and p21). Inversely, silencing of Id4 in LNCaP cells (LNCaP-Id4) led to decreased apoptosis due to an intact mitochondrial membrane and decrease in the expression of pro-apoptotic markers (PUMA, BAX, and p21). Since BAX, PUMA, and p21 are direct transcriptional targets of p53, these results therefore prompted us to investigate the effect of Id4 on expression and activity of p53. LNCaP cells express wild-type p53. DU145 cells harbor mutant p53 (P223L and V274F), which lies within the DNA binding domain and abrogates p53 transcriptional activity. DU145 cells also express high levels of p53, due extended half-life. Surprisingly, there was decreased expression of p53 in DU145+Id4 cells associated with nuclear localization indicating enhanced transcriptional activity. We investigated p53 DNA binding and transcriptional activity. We determined that mutant p53 in DU145+Id4 cells was transcriptionally active evident by increased luciferase activity and binding of p53 to the promoters of its targets. In LNCaP-Id4, p53 expression was decreased which resulted in decreased p53 transcriptional activity and decreased DNA binding ability. The data suggested that Id4 can restore mutant p53 activity, which is a significant observation. Our results also suggest that Id4 promotes the assembly of a macromolecular complex involving CBP/p300 that results in acetylation of p53 at K373, a critical post-translational modification required for its biological activity. Loss of Id4 in LNCaP cells also abrogated wild type p53 DNA binding and transcriptional activity with concomitant loss of CBP/p300 requirement and decreased acetylation. In conclusion, we demonstrated that loss of Id4 promotes cancer phenotype in LNCaP cells. We also demonstrated that the tumor suppressor activity of Id4 is in part through regulation of CBP/p300 dependent acetylation and function of p53.
text
application/pdf
2013-07-01
dissertation
Doctor of Philosophy (PhD)
Clark Atlanta University
School of Arts and Sciences, Biological Sciences
Chaudhary, Jaideep Hinton, Cimona Khan, Shafiq
Georgia--Atlanta
http://hdl.handle.net/20.500.12322/cau.td:2013_evans_ashley_l
