Selma M. Soyal

Steroid receptor coactivator 2 is critical for progesterone-dependent uterine function and mammary morphogenesis in the mouse

ABSTRACT Although the essential involvement of the progesterone receptor (PR) in female reproductive tissues is firmly established, the coregulators preferentially enlisted by PR to mediate its physiological effects have yet to be fully delineated. To further dissect the roles of members of the steroid receptor coactivator (SRC)/p160 family in PR-mediated reproductive processes in vivo, state-of-the-art cre-loxP engineering strategies were employed to generate a mouse model (PRCre/+SRC-2flox/flox) in which SRC-2 function was abrogated only in cell lineages that express the PR. Fertility tests revealed that while ovarian activity was normal, PRCre/+SRC-2flox/flox mouse uterine function was severely compromised. Absence of SRC-2 in PR-positive uterine cells was shown to contribute to an early block in embryo implantation, a phenotype not shared by SRC-1 or -3 knockout mice. In addition, histological and molecular analyses revealed an inability of the PRCre/+SRC-2flox/flox mouse uterus to undergo the necessary cellular and molecular changes that precede complete P-induced decidual progression. Moreover, removal of SRC-1 in the PRCre/+SRC-2flox/flox mouse uterus resulted in the absence of a decidual response, confirming that uterine SRC-2 and -1 cooperate in P-initiated transcriptional programs which lead to full decidualization. In the case of the mammary gland, whole-mount and histological analysis disclosed the absence of significant ductal side branching and alveologenesis in the hormone-treated PRCre/+SRC-2flox/flox mammary gland, reinforcing an important role for SRC-2 in cellular proliferative changes that require PR. We conclude that SRC-2 is appropriated by PR in a subset of transcriptional cascades obligate for normal uterine and mammary morphogenesis and function.

Progesterone involvement in breast development and tumorigenesis--as revealed by progesterone receptor "knockout" and "knockin" mouse models.

In light of recent clinical trials, the debate concerning the risks and benefits of progestin-based postmenopausal hormone replacement therapy (HRT) has reached a renewed level of urgency. Irrespective of the position taken, the consensus is that more basic research needs to be performed to address progesterones fundamental role in mammary development and tumorigenesis. Towards this end, the progesterone receptor knockout (PRKO) mouse demonstrated that progesterone is essential for pregnancy-associated mammary gland ductal side-branching and alveologenesis and that these morphological changes are dependent on progesterone-induced mammary epithelial proliferation. Importantly, the PRKO mouse showed that the progesterone-proliferative signal significantly contributes to mammary tumor susceptibility in an established mammary tumor model. Insight into the cellular mechanism(s) by which progesterone affects mammary morphogenesis has been disclosed by a new PR-LacZ knockin mouse, which revealed that PRs spatial expression pattern undergoes precise choreographed distributional changes that precede key stages in postnatal mammary development. In the case of early pregnancy, the segregation of cells undergoing progesterone-induced proliferation from those that express PR implicates a paracrine mode of action for progesterone-induced mammary epithelial proliferation, whereas the preparturient decline of PR expression underscores the need to remove this signal for full functional differentiation of this tissue. Our findings support the proposal that the mammary glands normal response to the progesterone-signal is dependent upon specific spatial organizational patterns of PR expression and that derailment in these cellular processes may contribute to abnormal mammary development, including cancer. This review concludes by emphasizing the need to identify the downstream molecular targets that mediate progesterones effects in this tissue. Identification of such targets will not only enhance our mechanistic understanding of progesterones role in mammary development and cancer, but may also facilitate the formulation of new design strategies in breast cancer diagnosis and/or treatment.

Targeting RANKL to a specific subset of murine mammary epithelial cells induces ordered branching morphogenesis and alveologenesis in the absence of progesterone receptor expression

Despite support for receptor of activated NF‐kB ligand (RANKL) as a mediator of mammary progesterone action, the extent to which this cytokine can functionally contribute to established progesterone‐induced mammary morphogenetic responses in the absence of other presumptive effectors is still unclear. To address this uncertainty, we developed an innovative bigenic system for the doxycycline‐inducible expression of RANKL in the mammary epithelium of the progesterone receptor knockout (PRKO) mouse. In response to acute doxycycline exposure, RANKL is specifically expressed in the estrogen receptor a (ER) positive/progesterone receptor negative (ER+/PR−) cell type in the PRKO mammary epithelium, a cell type that is equivalent to the ER+/PR+ cell type in the wild‐type (WT) mammary epithelium. Notably, the ER+/PR+ mammary cell normally expresses RANKL in the WT mammary epithelium during pregnancy. In this PRKO bigenic system, acute doxycycline‐induced expression of RANKL results in ordered mammary ductal side branching and alveologenesis, morphological changes that normally occur in the parous WT mouse. This mammary epithelial expansion is accompanied by significant RANKL‐induced luminal epithelial proliferation, which is driven, in part, by indirect induction of cyclin D1. Collectively, our findings support the conclusion that RANKL represents a critical mediator of mammary PR action and that restricted expression of this effector to the ER+/PR+ mammary cell‐type is necessary for a spatially ordered morphogenetic response to progesterone.—Mukherjee, A., Soyal, S. M., Li, J., Ying, Y., He, B., DeMayo, F.J., Lydon, J. P. Targeting RANKL to a specific subset of murine mammary epithelial cells induces ordered branching morphogenesis and alveologenesis in the absence of progesterone receptor expression. FASEB J. 24, 4408–4419 (2010). www.fasebj.org

PGC-1α is a male-specific disease modifier of human and experimental amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a devastating, adult-onset neurodegenerative disorder of the upper and lower motor systems. It leads to paresis, muscle wasting and inevitably to death, typically within 3-5 years. However, disease onset and survival vary considerably ranging in extreme cases from a few months to several decades. The genetic and environmental factors underlying this variability are of great interest as potential therapeutic targets. In ALS, men are affected more often and have an earlier age of onset than women. This gender difference is recapitulated in transgenic rodent models, but no underlying mechanism has been elucidated. Here we report that SNPs in the brain-specific promoter region of the transcriptional co-activator PGC-1α, a master regulator of metabolism, modulate age of onset and survival in two large and independent ALS populations and this occurs in a strictly male-specific manner. In complementary animal studies, we show that deficiency of full-length (FL) Pgc-1α leads to a significantly earlier age of onset and a borderline shortened survival in male, but not in female ALS-transgenic mice. In the animal model, FL Pgc-1α-loss is associated with reduced mRNA levels of the trophic factor Vegf-A in males, but not in females. In summary, we indentify PGC-1α as a novel and clinically relevant disease modifier of human and experimental ALS and report a sex-dependent effect of PGC-1α in this neurodegenerative disorder.

Targeting SREBPs for treatment of the metabolic syndrome.

Over the past few decades, mortality resulting from cardiovascular disease (CVD) steadily decreased in western countries; however, in recent years, the decline has become offset by the increase in obesity. Obesity is strongly associated with the metabolic syndrome and its atherogenic dyslipidemia resulting from insulin resistance. While lifestyle treatment would be effective, drugs targeting individual risk factors are often required. Such treatment may result in polypharmacy. Novel approaches are directed towards the treatment of several risk factors with one drug. Studies in animal models and humans suggest a central role for sterol regulatory-element binding proteins (SREBPs) in the pathophysiology of the metabolic syndrome. Four recent studies targeting the maturation or transcriptional activities of SREBPs provide proof of concept for the efficacy of SREBP inhibition in this syndrome.

Progesterone receptors - animal models and cell signaling in breast cancer: Progesterone's role in mammary gland development and tumorigenesis as disclosed by experimental mouse genetics

The progesterone receptor knockout mouse demonstrated progesterones importance to parity-induced mammary tertiary branching and lobuloalveologenesis. Because early parity provides significant protection against breast cancer whereas prolonged exposure to premenopausal ovarian progesterone (or to postmenopausal supplementations thereof) has been linked to breast cancer risk, this steroid can be considered to exhibit contrasting roles in breast cancer etiology. This review describes the important mouse models that have contributed to our understanding of progesterones role in mammary gland development and neoplasia. We conclude by emphasising the urgent need to identify the molecular targets of the progesterone receptor, and to determine whether these targets are modulated differently by the progesterone receptor isoforms (A and B) during mammary morphogenesis and tumorigenesis.

Glucose acts as a regulator of serum iron by increasing serum hepcidin concentrations.

Mutual clinical and molecular interactions between iron and glucose metabolism have been reported. We aimed to investigate a potential effect of glucose on iron homeostasis. We found that serum iron concentrations gradually decreased over 180 min after the administration of 75 g of glucose from 109.8 ± 45.4 mg/L to 94.4 ± 40.4 mg/L (P<.001; N=40) but remained unchanged in control subjects receiving tap water (N=21). Serum hepcidin, the key iron regulatory hormone which is mainly derived from hepatocytes but also expressed in pancreatic β-cells, increased within 120 min after glucose ingestion from 19.7 ± 9.9 nmol/L to 31.4 ± 21.0 nmol/L (P<.001). In cell culture, glucose induced the secretion of hepcidin and insulin into the supernatant of INS-1E cultures, but did not change the amount of hepcidin detectable in the hepatocyte cell culture HepG2. We additionally confirmed the expression of hepcidin in a human islet cell preparation. These results suggest that glucose acts as a regulator of serum iron concentrations, most likely by triggering the release of hepcidin from β-cells.

A mouse model to dissect progesterone signaling in the female reproductive tract and mammary gland.

Considering the regulatory complexities of progesterone receptor (PR) action throughout the female reproductive axis and mammary gland, we generated a mouse model that enables conditional ablation of PR function in a spatiotemporal specific manner. Exon 2 of the murine PR gene was floxed to generate a conditional PR allele (PRflox) in mice. Crossing the PRflox/flox mouse with the ZP3‐cre transgenic demonstrated that the PRflox allele recombines to a PR null allele (PRd). Mice homozygous for the recombined null PR allele (PRd/d) exhibit uterine, ovarian, and mammary gland defects that phenocopy those of our previously described PR knockout (PRKO) model. Therefore, this conditional mouse model for PR ablation represents an invaluable resource with which to further define in a developmental and/or reproductive stage‐specific manner the individual and integrative roles of distinct PR populations resident in multiple progesterone‐responsive target sites. genesis 48:106–113, 2010.

Characterization of Novel Peroxisome Proliferator-activated Receptor γ Coactivator-1α (PGC-1α) Isoform in Human Liver

Peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) is a transcriptional coactivator that contributes to the regulation of numerous transcriptional programs including the hepatic response to fasting. Mechanisms at transcriptional and post-transcriptional levels allow PGC-1α to support distinct biological pathways. Here we describe a novel human liver-specific PGC-1α transcript that results from alternative promoter usage and is induced by FOXO1 as well as glucocorticoids and cAMP-response element-binding protein signaling but is not present in other mammals. Hepatic tissue levels of novel and wild-type transcripts were similar but were only moderately associated (p < 0.003). Novel mRNA levels were associated with a polymorphism located in its promoter region, whereas wild-type transcript levels were not. Furthermore, hepatic PCK1 mRNA levels exhibited stronger associations with the novel than with the wild-type transcript levels. Except for a deletion of 127 amino acids at the N terminus, the protein, termed L-PGC-1α, is identical to PGC-1α. L-PGC-1α was localized in the nucleus and showed coactivation properties that overlap with those of PGC-1α. Collectively, our data support a role of L-PGC-1α in gluconeogenesis, but functional differences predicted from the altered structure suggest that L-PGC-1α may have arisen to adapt PGC-1α to more complex metabolic pathways in humans.

Cholesteryl ester transfer protein and hepatic lipase gene polymorphisms: Effects on hepatic mRNA levels, plasma lipids and carotid atherosclerosis

OBJECTIVE HDL modifying effects of cholesteryl ester transfer protein (CETP) and hepatic lipase (LIPC) depend in part on each other. We studied associations of CETP-Taq1B and -514C>T-LIPC polymorphisms with hepatic mRNA levels, and their combined effects on plasma lipids and carotid atherosclerosis. METHODS We genotyped the CETP-Taq1B and the -514C>T-LIPC polymorphisms in 67 obese women in whom hepatic CETP and LIPC transcript levels were determined as well as in 1549 participants of the Salzburg Atherosclerosis Prevention Program in Subjects at High Individual Risk (SAPHIR). Carotid atherosclerosis was assessed by intima-media thickness and extent of plaques (B-score) of the carotid arteries. RESULTS In obese women, CETP-Taq1B and -514C>T-LIPC variant alleles were associated with reduced hepatic levels of CETP and LIPC mRNA, respectively. The CETP and LIPC polymorphisms accounted for 12.9 and 14.4% of the variability in respective transcripts. In the SAPHIR population, CETP-Taq1B showed independent effects on LDL diameter, HDL and LDL cholesterol, apolipoproteins AI and B and cholesterol/HDL cholesterol, while -514C>T-LIPC revealed independent effects on HDL cholesterol and apolipoprotein AI. The two polymorphisms displayed interactions at the level of HDL cholesterol. Compared to subjects carrying wild-type alleles at both loci, subjects homozygous for the CETP wild-type allele, but heterozygous for the LIPC polymorphism and subjects heterozygous for the CETP polymorphism, but homozygous for the LIPC wild-type allele showed an increased risk of carotid atherosclerosis (both P<0.05). CONCLUSIONS CETP and LIPC polymorphisms influence the respective hepatic transcript levels, demonstrate interactions on HDL cholesterol and suggest that imbalances between CETP and LIPC activities may modulate the risk of carotid atherosclerosis.