Stress dramatically exacerbates pain in diseases such as fibromyalgia and rheumatoid arthritis, but the underlying mechanisms are unknown. We tested the hypothesis that stress causes generalized hyperalgesia by enhancing pronociceptive effects of immune mediators. Rats exposed to nonhabituating sound stress exhibited no change in mechanical nociceptive threshold, but showed a marked increase in hyperalgesia evoked by local injections of prostaglandin E 2 or epinephrine. This enhancement, which developed more than a week after exposure to stress, required concerted action of glucocorticoids and catecholamines at receptors located in the periphery on sensory afferents. The altered response to pronociceptive mediators involved a switch in coupling of their receptors from predominantly stimulatory to inhibitory G-proteins (G s to G i ), and for prostaglandin E 2 , emergence of novel dependence on protein kinase C. Thus, an important mechanism in generalized pain syndromes may be stress-induced coactivation of the hypothalamo-pituitary-adrenal and sympathoadrenal axes, causing a long-lasting alteration in intracellular signaling pathways, enabling normally innocuous levels of immune mediators to produce chronic hyperalgesia.
Many of the events required for productive herpes simplex virus type 1 (HSV-1) infection occur within globular nuclear domains called replication compartments, whose formation appears to depend on interactions with cellular nuclear domains 10 (ND10). We have previously demonstrated that the formation of HSV-1 replication compartments involves progression through several stages, including the disruption of intact ND10 (stage I to stage II) and the formation of PML-associated prereplicative sites (stage III) and replication compartments (stage IV) (J. Burkham, D. M. Coen, and S. K. Weller, J. Virol. 72:10100-10107, 1998). In this paper, we show that some, but not all, PML isoforms are recruited to stage III foci and replication compartments. Genetic experiments showed that the recruitment of PML isoforms to stage III prereplicative sites and replication compartments requires the localization of the HSV-1 polymerase protein (UL30) to these foci but does not require polymerase catalytic activity. We also examined the stages of viral infection under conditions affecting ND10 integrity. Treatment with factors that increase the stability of ND10, arsenic trioxide and the proteasome inhibitor MG132, inhibited viral disruption of ND10, formation of replication compartments, and production of progeny virus. These results strengthen the previously described correlation between ND10 disruption and productive viral infection.Herpes simplex virus type 1 (HSV-1) carries out gene expression, DNA replication, and DNA encapsidation in globular nuclear domains designated replication compartments (53, 55). These domains contain the essential viral DNA replication proteins (the origin-binding protein, the single-stranded-DNAbinding protein, the helicase-primase subunits, and the polymerase subunits [34,36,55]) and are usually visualized by antibodies either against ICP8, the single-stranded-DNA-binding protein, or UL42, the polymerase processivity subunit. The formation of replication compartments is mediated in part by interactions with nuclear structures called ND10 (nuclear domains 10), promyelocytic leukemia bodies, or PODs (17). The function of ND10 has not yet been defined for cellular or viral growth. Proteins found in ND10 have been associated with the control of cellular growth, cell cycle regulation, transcription, and apoptosis (11,12,24,27,46,71). In the case of the herpesviruses, viral DNA is deposited at ND10 and immediateearly transcripts can be detected at sites adjacent to ND10 (42). Furthermore, replication compartments formed after transfection with the seven essential HSV-1 replication proteins localize adjacent to ND10 (36, 74).ND10 are dynamic structures which are disrupted during mitosis and respond to environmental stimuli including interferon treatment, heat shock, treatment with heavy metals, and viral infection (44,64,65). The most extensively studied ND10 protein, PML, is expressed as a fusion with retinoic acid receptor ␣ in individuals with acute promyelocytic leukemia (31,56). In this disease, disrupti...
When herpes simplex virus type 1 (HSV-1) DNA replication is blocked by viral polymerase inhibitors, such as phosphonoacetic acid (PAA) or acyclovir (ACV), UL29 (ICP8) localizes to numerous punctate nuclear foci which are called prereplicative sites. Since this pattern can form in cells infected with mutants which are defective in UL5, UL8, UL9, or UL52 in the presence of polymerase inhibitors (C.
Herpes simplex virus type 1 (HSV-1) infection results in the disruption of ND10 (also called nuclear bodies, PODs, or PML-associated bodies), which are nuclear matrix domains of unknown function present in mammalian cells. After ND10 disruption, viral transcription and DNA replication occur in globular nuclear domains called replication compartments. In this report we define four stages of infection by using antibodies to ICP8 (also called SSB and UL29) and the ND10 antigen PML. Immediately after infection, cells contain intact ND10 as detected by staining for PMLs (stage I); within 1 hour, however, ND10 are disrupted and cells begin to exhibit diffuse staining for the major viral DNA binding protein, ICP8 (stage II). After all ND10 have been disrupted, foci which resemble but are not equivalent to ND10 appear, containing both PML and ICP8 (stage III). Cells infected with mutants defective in the helicase-primase or origin binding protein are unable to form stage III foci. Cells infected with a mutant that is null for the polymerase catalytic subunit, however, form stage III-like ICP8 foci which do not contain PML. Thus, stage III foci recruit the cellular PML protein in the presence but not the absence of HSV polymerase. PML was recruited to stage III foci in some but not all cells infected with a mutant defective in the polymerase accessory protein, UL42. Thus, UL42 is not required for the recruitment of PML to viral foci. In wild-type infection, stage III cells are quickly replaced by cells containing replication compartments (stage IV). PML and ICP8 staining are both observed within replication compartments, indicating a potential role for PML in HSV-1 replication. Models for the role of ND10 proteins in the formation of replication compartments are discussed.
We have tritium labeled two nucleic acid molecules, an 8 kDa DNA oligomer and a 20 kDa 'hammer-head' RNA for tritium NMR investigations. The DNA sequence studied has been previously used in homonuclear studies of DNA-bound water molecules and tritium NMR was expected to facilitate these investigations by eliminating the need to suppress the water resonance in tritium-detected 3H-1H NOESY experiments. We observed the anticipated through-space interactions found in B-form DNA in the NOESY experiments and an unexpected 'antiphase' cross-peak at the water frequency. T1 measurements on the tritiated DNA molecule indicated that relaxation rates were also accelerated for tritium and protons. Tritium NMR spectra of the hammerhead RNA molecule indicated conformational dynamics in the conserved region of the molecule in the absence of Mg2+ and spermine, two components necessary for cleavage. The dynamics were also investigated by 15N-correlated 1H spectroscopy and persisted after the addition of Mg2+ and spermine.
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