Transcriptional activation of the cyclin D1 gene (CCND1) plays a pivotal role in G 1 -phase progression, which is thereby controlled by multiple regulatory factors, including nuclear receptors (NRs). Appropriate CCND1 gene activity is essential for normal development and physiology of the mammary gland, where it is regulated by ovarian steroids through a mechanism(s) that is not fully elucidated. We report here that CCND1 promoter activation by estrogens in human breast cancer cells is mediated by recruitment of a c-Jun/c-Fos/estrogen receptor ␣ complex to the tetradecanoyl phorbol acetate-responsive element of the gene, together with Oct-1 to a site immediately adjacent. This process coincides with the release from the same DNA region of a transcriptional repressor complex including Yin-Yang 1 (YY1) and histone deacetylase 1 and is sufficient to induce the assembly of the basal transcription machinery on the promoter and to lead to initial cyclin D1 accumulation in the cell. Later on in estrogen stimulation, the cyclin D1/Cdk4 holoenzyme associates with the CCND1 promoter, where E2F and pRb can also be found, contributing to the long-lasting gene enhancement required to drive G 1 -phase completion. Interestingly, progesterone triggers similar regulatory events through its own NRs, suggesting that the gene regulation cascade described here represents a crossroad for the transcriptional control of G 1 -phase progression by different classes of NRs.Mammary gland morphogenesis and development result from the interplay of genetic and epigenetic pathways, controlled by hormones, growth factors, and other signaling molecules. Derangement of one or more of these regulatory pathways results in the abnormal growth and differentiation of mammary epithelial cells, leading to breast carcinogenesis. The ovarian hormones estrogen and progesterone promote mammary gland differentiation toward the female phenotype at the onset of puberty and control breast tropism and function throughout the reproductive life by affecting epithelial cell proliferation. Mammary gland cells are endowed with highaffinity receptors for these steroids (estrogen receptor ␣ [ER␣] and ER␤ and progesterone receptor A [PR-A] and PR-B, respectively), which belong to the nuclear receptor (NR) family of transcription factors (31

Phosphatidylinositol 4,5–biphosphate (PIP2) is a cell membrane phosphoinositide crucial for cell signaling and activation. Indeed, PIP2 is a pivotal source for second messenger generation and controlling the activity of several proteins regulating cytoskeleton reorganization. Despite its critical role in T cell activation, the molecular mechanisms regulating PIP2 turnover remain largely unknown. In human primary CD4+ T lymphocytes, we have recently demonstrated that CD28 costimulatory receptor is crucial for regulating PIP2 turnover by allowing the recruitment and activation of the lipid kinase phosphatidylinositol 4–phosphate 5–kinase (PIP5Kα). We also identified PIP5Kα as a key modulator of CD28 costimulatory signals leading to the efficient T cell activation. In this study, we extend these data by demonstrating that PIP5Kα recruitment and activation is essential for CD28-mediated cytoskeleton rearrangement necessary for organizing a complete signaling compartment leading to downstream signaling functions. We also identified Vav1 as the linker molecule that couples the C-terminal proline-rich motif of CD28 to the recruitment and activation of PIP5Kα, which in turn cooperates with Vav1 in regulating actin polymerization and CD28 signaling functions.

The effect of prostaglandin E 2 (PGE 2 ) in regulating the synthesis of the pro-inflammatory chemokine interleukin-8 (IL-8) in T lymphocytes is not yet defined, even though it may reduce or enhance IL-8 synthesis in other cell types. Here, we demonstrate that, in human T cells, PGE 2 induced IL-8 mRNA transcription through prostaglandin E 2 receptors 1-and 4-dependent signal transduction pathways leading to the activation of the transcription factor C/EBP homologous protein (CHOP), never before implicated in IL-8 transcription. Several kinases, including protein kinase C, Src family tyrosine kinases, phosphatidylinositol 3-kinase, and p38 MAPK, were involved in PGE 2 -induced CHOP activation and IL-8 production. The transactivation of the IL-8 promoter by CHOP was NF-B-independent. Our data suggest that PGE 2 acts as a potent pro-inflammatory mediator by inducing IL-8 gene transcription in activated T cells through different signal transduction pathways leading to CHOP activation. These findings show the complexity with which PGE 2 regulates IL-8 synthesis by inhibiting or enhancing its production depending on the cell types and environmental conditions.

CD28 superagonistic antibodies (CD28SAb) can preferentially activate and expand immunosuppressive regulatory T cells (Treg) in mice. However, pre-clinical trials assessing CD28SAbs for the therapy of autoimmune diseases reveal severe systemic inflammatory response syndrome in humans, thereby implying the existence of distinct signalling abilities between human and mouse CD28. Here, we show that a single amino acid variant within the C-terminal proline-rich motif of human and mouse CD28 (P212 in human vs. A210 in mouse) regulates CD28-induced NF-κB activation and pro-inflammatory cytokine gene expression. Moreover, this Y209APP212 sequence in humans is crucial for the association of CD28 with the Nck adaptor protein for actin cytoskeleton reorganisation events necessary for CD28 autonomous signalling. This study thus unveils different outcomes between human and mouse CD28 signalling to underscore the importance of species difference when transferring results from preclinical models to the bedside.