MicroRNA (miR)-155 is upregulated in breast cancer cells and in sera of patients with breast cancer, but its clinical relevance remains uncertain. The objective of the present effort was to address the transcriptional regulation of miR-155. A bioinformatics analysis of public datasets validated upregulation of miR-155 in tumor cells of patients with breast cancer, particularly those who were at early stages and had triple-negative cancers. The expression profiling and clinical relevance of miR-155 in tumor cells and blood cells were characterized by TaqMan miR assays and, in plasma and exosomes, by nest-quantitative PCR analysis. There was a positive correlation between expression of FOXP3 and miR-155 in breast cancer cell lines and primary breast cancers. In breast cancer cells, FOXP3 induced miR-155 through transcriptional repression of BRCA1. Furthermore, in an Alabama cohort, blood and plasma samples were collected from 259 participants, including patients with breast cancer or benign breast tumors, members of breast cancer families, and matched healthy female controls. For patients with early stage or localized breast cancer, there were high levels of miR-155 in both plasma and blood cells. In cultured breast cancer cells, expression of miR-155 was induced by FOXP3 but was not significantly changed in culture medium or exosomes, suggesting that circulating miR-155 originated from blood cells. These findings reveal a transcriptional axis of FOXP3-BRCA1-miR-155 in breast cancer cells and show that plasma miR-155 may serve as a non-invasive biomarker for detection of early stage breast cancer.

The adenosine A1 receptor (A1R) is a key mediator of the neuroprotective effect by endogenous adenosine. Yet targeting this receptor for neuroprotection is challenging due to its broad expression throughout the body. A mechanistic understanding of the regulation of A1R signaling is necessary for the future design of therapeutic agents that can selectively enhance A1R-mediated responses in the nervous system. In this study, we demonstrate that A1R activation leads to a sustained localization of regulator of G protein signaling 4 (RGS4) at the plasma membrane, a process that requires neurabin (a neural tissue-specific protein). A1R and RGS4 interact with the overlapping regions of neurabin. In addition, neurabin domains required for oligomerization are essential for formation of the A1R/neurabin/RGS4 ternary complex, as well as for stable localization of RGS4 at the plasma membrane and attenuation of A1R signaling. Thus, A1R and RGS4 each likely interact with one neurabin molecule in a neurabin homo-oligomer to form a ternary complex, representing a novel mode of regulation of G protein-coupled receptor signaling by scaffolding proteins. Our mechanistic analysis of neurabin-mediated regulation of A1R signaling in this study will be valuable for the future design of therapeutic agents that can selectively enhance A1R-mediated responses in the nervous system.

The therapeutic mechanism of action underlying many psychopharmacological agents remains poorly understood, due largely to the extreme molecular promiscuity exhibited by these agents with respect to potential central nervous system targets. Agents of the tricyclic chemical class, including both antidepressants and antipsychotics, exhibit a particularly high degree of molecular promiscuity; therefore, any clarification of how these agents interact with specific central nervous system targets is of great potential significance to the field. Here, we present evidence demonstrating that tricyclic antipsychotics appear to segregate into three distinct groups based upon their molecular interactions with the centrally-important α2A adrenergic receptor (AR). Specifically, while the α2AAR binds all antipsychotics tested with similar affinities, and none of the agents are able to induce classical heterotrimeric G protein-mediated α2AAR signaling, significant differences are observed with respect to arrestin3 recruitment and receptor endocytosis. All antipsychotics tested induce arrestin3 recruitment to the α2AAR, but with differing strengths. Both chlorpromazine and clozapine drive significant α2AAR endocytosis, but via differing clathrin-dependent and lipid raft-dependent pathways, while fluphenazine does not drive a robust response. Intriguingly, in silico molecular modeling suggests that each of the three exhibits unique characteristics in interacting with the α2AAR ligand-binding pocket. In addition to establishing these three antipsychotics as novel arrestin-biased ligands at the α2AAR, our findings provide key insights into the molecular actions of these clinically-important agents.

Jun dimerization protein 2 (JDP2), a basic leucine zipper transcription factor, is involved in numerous biological and cellular processes such as cancer development and regulation, cell-cycle regulation, skeletal muscle and osteoclast differentiation, progesterone receptor signaling, and antibacterial immunity. Though JDP2 is widely expressed in mammalian tissues, its function in gonads and adrenals (such as regulation of steroidogenesis and adrenal development) is largely unknown. Herein, we find that JDP2 mRNA and proteins are expressed in mouse adrenal gland tissues. Moreover, overexpression of JDP2 in Y1 mouse adrenocortical cancer cells increases the level of melanocortin 2 receptor (MC2R) protein. Notably, Mc2r promoter activity is activated by JDP2 in a dose-dependent manner. Next, by mapping the Mc2r promoter, we show that cAMP response elements (between −1320 and −720-bp) are mainly required for Mc2r activation by JDP2 and demonstrate that −830-bp is the major JDP2 binding site by real-time chromatin immunoprecipitation (ChIP) analysis. Mutations of cAMP response elements on Mc2r promoter disrupts JDP2 effect. Furthermore, we demonstrate that removal of phosphorylation of JDP2 results in attenuated transcriptional activity of Mc2r. Finally, we show that JDP2 is a candidate for SUMOylation and SUMOylation affects JDP2-mediated Mc2r transcriptional activity. Taken together, JDP2 acts as a novel transcriptional activator of the mouse Mc2r gene, suggesting that JDP2 may have physiological functions as a novel player in MC2R-mediated steroidogenesis as well as cell signaling in adrenal glands.