No abstract

The adenovirus VA1 RNA (VA1), a 160-nucleotide (nt)-long RNA transcribed by RNA polymerase III, is efficiently exported from the nucleus to the cytoplasm of infected cells, where it antagonizes the interferon-induced antiviral defense system. We recently reported that nuclear export of VA1 is mediated by a cis-acting RNA export motif, called minihelix, that comprises a double-stranded stem (>14 nt) with a base-paired 5 end and a 3-8-nt protruding 3 end. RNA export mediated by the minihelix motif is Ran-dependent, which indicates the involvement of a karyopherin-related factor (exportin) that remained to be determined. Here we show using microinjection in Xenopus laevis oocytes that VA1 is transported to the cytoplasm by exportin-5, a nuclear transport factor for double-stranded RNA binding proteins. Gel retardation assays revealed that exportin-5 directly interacts with VA1 RNA in a RanGTP-dependent manner. More generally, in vivo and in vitro competition experiments using various VA1-derived, but also artificial and cellular, RNAs lead to the conclusion that exportin-5 preferentially recognizes and transports minihelix motif-containing RNAs.Nucleo-cytoplasmic transport of most RNAs and proteins is dependent on soluble receptors called karyopherins that can dock at and translocate through the nuclear pore complex. Interaction between cargo and karyopherin ␤ is governed by the GTPase Ran. The asymmetric distribution of the Ran regulatory proteins provides a steep gradient of RanGDP (cytoplasmic)/RanGTP (nuclear) across the nuclear envelope that ensures the directionality of nuclear transport (1, 2). Nuclear import receptors unload their cargo upon binding to RanGTP in the nucleus, whereas RanGTP is used to assemble export complexes which are in turn destabilized by dissociation of RanGTP in the cytoplasm (3, 4).Our understanding of the nuclear export of RNAs has been greatly facilitated by the study of viral RNAs. For this reason, we focused our attention on the adenovirus VA1 RNA (VA1), a 160-nt 1 -long RNA transcribed by RNA polymerase III that massively accumulates in the cytoplasm of infected cells. It serves to antagonize the interferon-induced cellular antiviral defense system. Indeed, VA1 binds and inhibits the doublestranded RNA-dependent protein kinase R (PKR), which otherwise phosphorylates eIF2␣ and leads to the inhibition of protein synthesis (5, 6). Adenovirus VA1 RNA contains a new cis-acting RNA export motif that comprises a double-stranded stem (Ͼ14 nt) with a base-paired 5Ј end and a 3-8-nt protruding 3Ј end and that can tolerate some mismatches and bends (7). This export signal, called minihelix, is present not only in VA1 but in a large family of small viral and cellular RNAs transcribed by polymerase III. RNA export mediated by the minihelix motif is Ran-dependent, which indicates the involvement of a karyopherin-related factor (exportin). This exportin is distinct from Crm1 and exportin-t (7, 8). Therefore, we sought to identify cellular factors that bind to and mediate the export of miniheli...

An amplifying role for oral epithelial cells (ECs) in Epstein-Barr Virus (EBV) infection has been postulated to explain oral viral shedding. However, while lytic or latent EBV infections of oro/nasopharyngeal ECs are commonly detected under pathological conditions, detection of EBV-infected ECs in healthy conditions is very rare. In this study, a simple non-surgical tissue sampling procedure was used to investigate EBV infection in the periodontal epithelium that surrounds and attaches teeth to the gingiva. Surprisingly, we observed that the gingival ECs of the periodontium (pECs) are commonly infected with EBV and may serve as an important oral reservoir of latently EBV-infected cells. We also found that the basal level of epithelial EBV-infection is significantly increased in chronic periodontitis, a common inflammatory disease that undermines the integrity of tooth-supporting tissues. Moreover, the level of EBV infection was found to correlate with disease severity. In inflamed tissues, EBV-infected pECs appear to be prone to apoptosis and to produce larger amounts of CCL20, a pivotal inflammatory chemokine that controls tissue infiltration by immune cells. Our discovery that the periodontal epithelium is a major site of latent EBV infection sheds a new light on EBV persistence in healthy carriers and on the role of this ubiquitous virus in periodontitis. Moreover, the identification of this easily accessible site of latent infection may encourage new approaches to investigate and monitor other EBV-associated disorders.

Growth hormone (GH) is required for the terminal differentiation of preadipose Ob1771 cells that have entered the differentiation program as evidenced by the expression of early marker genes (pOb24 and lipoprotein lipase).Induction of c-os mRNA within 15 min and induction of insulin-like growth factor I mRNA within a few hours take place in response to GH. The role of GH is mediated, at least in part, by means ofthe activation ofprotein kinase C, as shown by the inhibition ofepidermal growth factor binding and by the expression of the c-fos gene, and is thus analogous to the action of prostaglandin F2. and 4l-phorbol-12,13-didecanoate in this respect. However, in contrast to that of the c-fos gene, the regulation ofinsulin-like growth factor I gene expression by GH is not mediated by means of the activation of protein kinase C, and, in line with this, prostaglandin F2, and 4f3-phorbol-12,13-didecanoate were ineffective. GH and prostaglandin F2.were able to stimulate the formation of diacylglycerol within a few seconds, but GH did not elicit an accumulation of inositol phosphates, in contrast to that generated by prostaglandin Fl.We conclude that the transduction signal of GH action in c-fos mRNA induction is the formation of diacylglycerol and that the mechanism whereby GH can activate protein kinase C is associated with a phospholipase C-mediated hydrolysis of glycerophospholipids other than inositol phospholipids.