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The herpes simplex virus 1 ORF UL41 encodes a protein (virion host shutoff or vhs) associated with selective degradation of mRNA early in infection. Some mRNAs, exemplified by GAPDH or β-actin mRNAs, are degraded rapidly. Others, for example IEX-1 mRNA, are degraded in two stages: whereas the 3′ domain disappears rapidly, a large 5′ domain fragment of the mRNA lingers for several hours. Still a third, exemplified by tristetraprolin mRNA, is not degraded, allowing its protein product to accumulate in infected cells. Here we report the following: (i) a GST-vhs protein produced in Escherichia coil, solubilized and purified to homogeneity acts as bona fide endoribonuclease when tested on in vitro transcribed IEX-1 probes. A GST-vhs protein in which three key vhs amino acids were replaced with alanines, solubilized and purified by the same protocol, had no enzymatic activity. (ii) The number of fragments generated by cleavage of a truncated IEX-1 RNA by vhs appears to be small; the cleavage sites are centered at or near the AU-rich elements located at the 3′ untranslated region of the mRNA. A truncated RNA containing only the IEX-1 coding domain was cleaved numerous times. (iii) In cells infected at high multiplicity and exposed to a large number of particles per cell, the vhs protein accumulated within 3 h after infection, in small uniform cytoplasmic granules raising the possibility that vhs colocalizes with tristerapolin, a protein induced after infection, in structures involved in degradation of RNA. [ABSTRACT FROM AUTHOR]

The UL41 protein of herpes simplex virus I has been reported to mediate the degradation of both viral and cellular mRNAs. Extensive studies on β-actin and some viral mRNAs were consonant with this conclusion. In earlier studies, we reported that the UL41- dependent degradation of cellular mRNAs up-regulated after infection was selective. One class of the up-regulated mRNAs, exemplified by the stress-inducible immediate-early 1 mRNA, is deadenylated, 3' to 5' degraded and is not translated. Another class of up-regulated mRNAs, exemplified by GADD45β, does not undergo this pattern of degradation and is translated. A puzzling feature of the earlier results is that the amounts of up-regulated mRNAs accumulating in the cytoplasm of ΔUL41 mutant virus-infected cells was lower than in WT virus-infected cells, a contra- diction, inasmuch as if the rates of accumulation were identical and degradation of the mRNAs were higher in WT virus-infected cells, the steady-state levels should have been higher in ΔUL41 mutant virus-infected cells. In this report, we show that in ΔUL41 mutant virus-infected cells, the rates of degradation of the stress-inducible immediate-early response gene I and other up-regulated mRNAs are approximately the same as those observed in mock-infected cells and are faster than in WT virus-infected cells. This is contrary to the observed UL41 -dependent degradation of β-actin and other mRNAs. The UL41 protein thus mediates two functions, i.e., it mediates rapid degradation of some mRNAs exemplified by β-actin and stabilizes or delays the degradation of other mRNAs exemplified by GADD45β, tristetraprolin, etc. A model unifying both activities of the UL41 protein is presented. [ABSTRACT FROM AUTHOR]

In cells infected with herpes simplex virus 1, the RNA encoded by the stress-inducible immediate early response gene IEX-1 was up-regulated immediately after infection. However, the accumulated RNA was degraded 3'-5', and the protein was detectable only at very early times after infection. The degradation was dependent on the UL41 gene encoding the virion host shutoff (vhs) protein and resulted in the accumulation of truncated RNA containing the 5'-end portion of the transcript. IEX-1 contains an AU-rich element (ARE) in its 3'-untranslated domains known to regulate negatively the RNA lifespan. To examine the rote of ARE in signaling the degradation, we compared the stability of several RNAs up-regulated dining infection to WT virus. These were ARE-containing RNAs encoding IEX-1, c-fos, and IκBα and the non-ARE-containing RNAs GADD45β and tristetraprolin. We report that the ARE-containing RNAs exemplified by IEX-1 RNA are deadenylated and cleaved in the ARE within the 3' UTR in a UL41-dependent manner. In contrast, Northern blot hybridizations and analyses of poly(A) tails revealed no evidence of degradation of GADD45β RNA. GADD45β protein was detected in WT virus-infected cells. These results indicate that the degradation of RNAs and the mechanism by which cellular RNAs are degraded are selective and may be sequence specific. The persistence of partially degraded ARE-containing RNAs may reflect specific targeting of the vhs proteins to the ARE and the modification of the RNA degradation machinery of the cell induced by the presence of the vhs protein. [ABSTRACT FROM AUTHOR]

Microarray data reported elsewhere indicated that herpes simplex virus I induces the up-regulation of nuclear factor &key;B (NF-&key;B)regulated genes, including that of its inhibitor, I&key;Bα, consistent with the reports that wild-type virus induces the activation of NF-&key;B. In this report we show that activation of NF-&key;B in infected cells is linked to the activation of protein kinase R (PKR). Specifically: (i) PKR is activated in infected cells although the effects of the activated enzyme on protein synthesis are negated by the viral gene 1,134.5, which encodes a protein phosphatase la accessory factor that enables the dephosphorylation of the a subunit of eukaryotic translation initiation factor 2. NF-&key;B is activated in wild-type murine embryonic fibroblasts but not in related PKR-null cells. (ii) In cells infected with a replication-competent δγ134.5 mutant (R5104), but carrying a Us11 gene expressed early in infection, eukaryotic translation initiation factor 2α is not phosphorylated, and in in vitro assays, PKR bound to the Us11 protein is not phosphorylated on subsequent addition of double-stranded RNA. Here we report that this mutant does not activate PKR, has no effect on the accumulation of I&key;Bα, and does not cause the translocation of NF-&key;B in infected cells. (iii) One hypothesis advanced for the activation of NF-&key;B is that it blocks apoptosis induced by viral gene products. The replication-competent R5104 mutant does not induce the programmed cell's death. We conclude that in herpes simplex virus 1-infected cells, activation of NF-&key;B depends on activation of PKR and that NF-&key;B is not required to block apoptosis in productively infected cells. [ABSTRACT FROM AUTHOR]

Focuses on the role of viral ORF byproduct VP22 in the transfer of the RNA from cell to cell for expression prior to viral infection. Identification of proteins capable of binding RNA; Physiologic role of RNA packaged in virions; Processes involved in herpes simplex virus replication in cultured cells.