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The Ebola virus (EBOV) genome encodes a partly conserved 40-residue nonstructural polypeptide, called the delta peptide, that is produced in abundance during Ebola virus disease (EVD). The function of the delta peptide is unknown, but sequence analysis has suggested that delta peptide could be a viroporin, belonging to a diverse family of membrane-permeabilizing small polypeptides involved in replication and pathogenesis of numerous viruses. Full-length and conserved C-terminal delta peptide fragments permeabilize the plasma membranes of nucleated cells of rodent, dog, monkey, and human origin; increase ion permeability across confluent cell monolayers; and permeabilize synthetic lipid bilayers. Permeabilization activity is completely dependent on the disulfide bond between the two conserved cysteines. The conserved C-terminal portion of the peptide is biochemically stable in human serum, and most serum-stable fragments have full activity. Taken together, the evidence strongly suggests that Ebola virus delta peptide is a viroporin and that it may be a novel, targetable aspect of Ebola virus disease pathology.IMPORTANCE During the unparalleled West African outbreak of Ebola virus disease (EVD) that began in late 2013, the lack of effective countermeasures resulted in chains of serial infection and a high mortality rate among infected patients. A better understanding of disease pathology is desperately needed to develop better countermeasures. We show here that the Ebola virus delta peptide, a conserved nonstructural protein produced in large quantities by infected cells, has the characteristics of a viroporin. This information suggests a critical role for the delta peptide in Ebola virus disease pathology and as a possible target for novel countermeasures.

Nuclear hormone receptors, such as the estrogen receptors (ERs), are regulated by specific kinase signaling pathways. Here, we demonstrate that the p38 MAPK stimulates both ERalpha- and ERbeta-mediated transcription in MCF-7 breast carcinoma, Ishikawa endometrial adenocarcinoma, and human embryonic kidney 293 cells. Inhibition of this potentiation using the p38 inhibitor, RWJ67657, blocked estrogen-mediated transcription and proliferation. Activated ERs promote gene expression in part through the recruitment of the p160 class of coactivators. Because no direct p38 phosphorylation sites have been determined on either ERalpha or beta, we hypothesized that p38 could target the p160 class of coactivators. We show for the first time using pharmacological and molecular techniques that the p160 coactivator glucocorticoid receptor-interacting protein 1 (GRIP1) is phosphorylated and potentiated by the p38 MAPK signaling cascade in vitro and in vivo. S736 was identified as a necessary site for p38 induction of GRIP1 transcriptional activation. The C terminus of GRIP1 was also demonstrated to contain a p38-responsive region. Taken together, these results indicate that p38 stimulates ER-mediated transcription by targeting the GRIP1 coactivator.

Uterine leiomyomas, benign uterine smooth muscle tumors that affect 30% of reproductive-aged women, are a significant health concern. The initiation event for these tumors is unclear, but 17beta-estradiol (E2) is an established promoter of leiomyoma growth. E2 not only alters transcription of E2-regulated genes but also can rapidly activate signaling pathways. The aim of our study is to investigate the role of rapid E2-activated cytoplasmic signaling events in the promotion of leiomyomas. Western blot analysis revealed that E2 rapidly increases levels of phosphorylated protein kinase C alpha (PKC alpha) in both immortalized uterine smooth muscle (UtSM) and leiomyoma (UtLM) cell lines, but increases levels of phosphorylated ERK1/2 only in UtLM cells. Our studies demonstrate a paradoxical effect of molecular and pharmacological inhibition of PKC alpha on ERK1/2 activation and cellular proliferation in UtLM and UtSM cells. PKC alpha inhibition decreases levels of phosphorylated ERK1/2 and proliferation in UtLM cells but raises these levels in UtSM cells. cAMP-PKA signaling is rapidly activated only in UtSM cells with E2 and inhibits ERK1/2 activation and proliferation. We therefore propose a model whereby E2's rapid activation of PKC alpha and cAMP-PKA signaling plays a central role in the maintenance of a low proliferative index in normal uterine smooth muscle via its inhibition of the MAPK cascade and these pathways are altered in leiomyomas to promote MAPK activation and proliferation. These studies demonstrate that rapid E2-signaling pathways contribute to the promotion of leiomyomas.