TReP-132 is a novel progesterone receptor coactivator required for the inhibition of breast cancer cell growth and enhancement of differentiation by progesterone.

The sex steroid progesterone is essential for the proliferation and differentiation of the mammary gland epithelium during pregnancy. In relation to this, in vitro studies using breast carcinoma T47D cells have demonstrated a biphasic progesterone response, consisting of an initial proliferative burst followed by a sustained growth arrest. However, the transcriptional factors acting with the progesterone receptor (PR) to mediate the progesterone effects on mammary cell growth and differentiation remain to be determined. Recently, it has been demonstrated that the transcriptional regulating protein of 132 kDa (TReP-132), initially identified as a regulator of steroidogenesis, is also a cell growth suppressor. Similar to progesterone-bound PR, TReP-132 acts by inducing the gene expression of the G1 cyclin-dependent kinase inhibitors p21WAF1/Cip1 (p21) and p27Kip1 (p27). The putative interaction between TReP-132 and progesterone pathways in mammary cells was therefore analyzed in the present study. Our results show that TReP-132 interacts in vitro and in T47D cells with progesterone-activated PR. TReP-132 synergizes with progesterone-bound PR to trans activate the p21 and p27 gene promoters at proximal Sp1-binding sites. Moreover, TReP-132 overexpression and knockdown, respectively, increased or prevented the induction of p21 and p27 gene expression by progesterone. As a consequence, TReP-132 knockdown also resulted in the loss of the inhibitory effects of progesterone on pRB phosphorylation, G1/S cell cycle progression, and cell proliferation. Furthermore, the knockdown of TReP-132 expression also prevented the induction of both early and terminal markers of breast cell differentiation which had been previously identified as progesterone target genes. As well, the progesterone-induced accumulation of lipid vacuoles was inhibited in the TReP-132-depleted cells. Finally, TReP-132 gene expression levels increased following progesterone treatment, indicating the existence of a positive auto-regulatory loop between PR and TReP-132. Taken together, these data identify TReP-132 as a coactivator of PR mediating the growth-inhibitory and differentiation effects of progesterone on breast cancer cells.

Progesterone inhibits human breast cancer cell growth through transcriptional upregulation of the cyclin-dependent kinase inhibitor p27Kip1 gene.

The effects of progesterone derivatives on breast cancer development are still controversial, probably accounting for their biphasic, opposed effects on mammary cell-cycle regulation. Here, we demonstrate in vitro that the growth-inhibitory effects of progesterone on breast cancer T-47D cells require the transcriptional upregulation of the cyclin-dependent kinase inhibitor p27(Kip1) (p27) gene. A statistical analysis of human tumor biopsies further indicates that p27 mRNA levels correlate to progesterone receptor (PR) levels. Moreover, p27 gene expression is inversely associated with tumor aggressiveness, and is a prognostic factor of favorable disease outcome. Thus, progesterone derivatives selectively activating the p27 gene promoter could be promising drugs against breast cancer progression.

PPAR alpha inhibits vascular smooth muscle cell proliferation underlying intimal hyperplasia by inducing the tumor suppressor p16INK4a.

Vascular SMC proliferation is a crucial event in occlusive cardiovascular diseases. PPARalpha is a nuclear receptor controlling lipid metabolism and inflammation, but its role in the regulation of SMC growth remains to be established. Here, we show that PPARalpha controls SMC cell-cycle progression at the G1/S transition by targeting the cyclin-dependent kinase inhibitor and tumor suppressor p16(INK4a) (p16), resulting in an inhibition of retinoblastoma protein phosphorylation. PPARalpha activates p16 gene transcription by both binding to a canonical PPAR-response element and interacting with the transcription factor Sp1 at specific proximal Sp1-binding sites of the p16 promoter. In a carotid arterial-injury mouse model, p16 deficiency results in an enhanced SMC proliferation underlying intimal hyperplasia. Moreover, PPARalpha activation inhibits SMC growth in vivo, and this effect requires p16 expression. These results identify an unexpected role for p16 in SMC cell-cycle control and demonstrate that PPARalpha inhibits SMC proliferation through p16. Thus, the PPARalpha/p16 pathway may be a potential pharmacological target for the prevention of cardiovascular occlusive complications of atherosclerosis.

TReP-132 controls cell proliferation by regulating the expression of the cyclin-dependent kinase inhibitors p21WAF1/Cip1 and p27Kip1.

The transcriptional regulating protein of 132 kDa (TReP-132) has been identified in steroidogenic tissues, where it acts as a coactivator of steroidogenic factor 1 (SF-1). We show here that TReP-132 plays a role in the control of cell proliferation. In human HeLa cells, TReP-132 knockdown by using small interfering RNA resulted in increased G(1)-->S cell cycle progression. The growth-inhibitory effects of TReP-132 was further shown to be mediated by induction of G(1) cyclin-dependent kinase inhibitors p21(WAF1) (p21) and p27(KIP1) (p27) expression levels. As a consequence, G(1) cyclin/cyclin-dependent kinase activities and pRB phosphorylation were markedly reduced, and cell cycle progression was blocked in the G(1) phase. The stimulatory effect of TReP-132 on p21 and p27 gene transcription involved interaction of TReP-132 with the transcription factor Sp1 at proximal Sp1-binding sites in their promoters. Moreover, in different breast tumor cell lines, endogenous TReP-132 expression was positively related with a lower proliferation rate. In addition, TReP-132 knockdown resulted in enhanced cell proliferation and lowered p21 and p27 mRNA levels in the steroid-responsive and nonresponsive T-47D and MDA-MB-231 cell lines, respectively. Finally, a statistic profiling of human breast tumor samples highlighted that expression of TReP-132 is correlated with p21 and p27 levels and is associated with lower tumor incidence and aggressiveness. Together, these results identify TReP-132 as a basal cell cycle regulatory protein acting, at least in part, by interacting with Sp1 to activate the p21 and p27 gene promoters.