Herpes simplex virus type 1 (HSV-1) mutants that fail to express the viral immediate-early protein ICP0 have a pronounced defect in viral gene expression and plaque formation in limited-passage human fibroblasts. ICP0 is a RING finger E3 ubiquitin ligase that induces the degradation of several cellular proteins. PML, the organizer of cellular nuclear substructures known as PML nuclear bodies or ND10, is one of the most notable proteins that is targeted by ICP0. Depletion of PML from human fibroblasts increases ICP0-null mutant HSV-1 gene expression, but not to wild-type levels. In this study, we report that depletion of Sp100, another major ND10 protein, results in a similar increase in ICP0-null mutant gene expression and that simultaneous depletion of both proteins complements the mutant virus to a greater degree. Although chromatin assembly and modification undoubtedly play major roles in the regulation of HSV-1 infection, we found that inhibition of histone deacetylase activity with trichostatin A was unable to complement the defect of ICP0-null mutant HSV-1 in either normal or PML-depleted human fibroblasts. These data lend further weight to the hypothesis that ND10 play an important role in the regulation of HSV-1 gene expression.Herpes simplex virus type 1 (HSV-1) is a common human pathogen that causes recurrent infections through its ability to establish a latent state in sensory ganglia after primary epithelial infections (for a general review, see reference 58). Lytic HSV-1 infection is characterized by abundant transcription from the entire viral genome in a temporal cascade of immediate-early (IE), early, and late gene products. The IE gene products regulate the expression of later classes of viral genes. In contrast, lytic cycle genes are repressed during latency, and only the latency-associated transcripts (LATs; derived from a single locus that lies countersense to the IE gene encoding ICP0) are expressed in readily detectable amounts (8,55,69). The IE protein ICP0 is a RING finger E3 ubiquitin ligase (4) that is required for efficient entry into the lytic cycle and which can induce reactivation of latent or quiescent genomes (reviewed in references 12, 14, 15, 29-31, and 55). ICP0 influences many cellular pathways, and one of its most prominent activities is its ability to localize to and disrupt nuclear substructures known as PML nuclear bodies (also known as ND10) (reviewed in references 10, 14, 16, and 43). This disruption occurs through ICP0-induced degradation of PML (17), the key component of ND10 which is required for assembly of these structures (34, 76). HSV-1 mutants that fail to express ICP0 or that express mutant ICP0 proteins that lack RING finger activity are unable to disrupt ND10 or to degrade PML (4,11,17,44,45). Such mutants have a profound defect in HSV-1 gene expression after infection of limited-passage human fibroblasts (9,21,62,63).The strong correlation between the effects of ICP0 on ND10and its requirement for lytic virus infection prompted the hypothesis that ND10 might ...

The development of treatments for neuropathic pain has been hindered by our limited understanding of the basic mechanisms underlying abnormalities in nociceptor hyperexcitability. We recently showed that the polymodal receptor transient receptor potential vanilloid 4 (TRPV4), a member of the transient receptor potential (TRP) family of ion channels, may play a role in inflammatory pain (Alessandri-Haber et al., 2003). The present study tested whether TRVP4 also contributes to neuropathic pain, using a rat model of Taxol-induced painful peripheral neuropathy. Taxol is the most widely used drug for the treatment of a variety of tumor types, but the dose of Taxol that can be tolerated is limited by the development of a small-fiber painful peripheral neuropathy.We found that Taxol treatment enhanced the nociceptive behavioral responses to both mechanical and hypotonic stimulation of the hind paw. Spinal administration of antisense oligodeoxynucleotides to TRPV4, which reduced the expression of TRPV4 in sensory nerve, abolished Taxol-induced mechanical hyperalgesia and attenuated hypotonic hyperalgesia by 42%. The enhancement of osmotic nociception involves sensitization of osmotransduction in primary afferents because osmotransduction was enhanced in cultured sensory neurons isolated from Taxol-treated rats. Taxol-induced TRPV4-mediated hyperalgesia and the enhanced osmotransduction in cultured nociceptors were dependent on integrin/Src tyrosine kinase signaling. These results suggest that TRPV4 plays a crucial role in a painful peripheral neuropathy, making it a very promising target for the development of a novel class of analgesics.

Toward the goal of defining new pharmacological targets for the treatment of chronic pain conditions, in previous studies we established a model, termed 'hyperalgesic priming,' in which an acute inflammatory stimulus causes a long-lasting latent susceptibility to hyperalgesia induced by subsequent exposures to the inflammatory mediator, prostaglandin E2 (PGE2). Those investigations suggested the hypothesis that priming induces a novel linkage between the PGE2-activated second messenger cascade and the epsilon isoform of protein kinase C (PKCepsilon). In the present study, comparison of dose-response relations for hyperalgesia produced by PGE2, forskolin, 8-Br-cAMP, or the protein kinase A (PKA) catalytic subunit, in primed versus normal animals, demonstrated that priming-induced enhancement of the PGE2-activated second messenger cascade occurs downstream to adenylate cyclase and upstream to PKA. Therefore, PGE2-induced hyperalgesia in the primed animal is enhanced by the recruitment of a novel cAMP/PKCepsilon signaling pathway in addition to the usual cAMP/PKA pathway. These observations suggest that pharmacological disruption of the novel interaction between cAMP and PKCepsilon might provide a route toward the development of highly specific methods to reverse cellular processes that underlie chronic pain states.

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