The Latency-Associated Nuclear Antigen Homolog of Herpesvirus Saimiri Inhibits Lytic Virus Replication

Herpesvirus saimiri (HVS), a T-lymphotropic tumor virus of neotropical primates, and the Kaposi's sarcoma-associated human herpesvirus 8 (KSHV) belong to the gamma-(2)-herpesvirus (Rhadinovirus) subfamily and share numerous features of genome structure and organization. The KSHV latency-associated nuclear antigen (LANA) protein appears to be relevant for viral persistence, latency, and transformation. It binds to DNA, colocalizes with viral episomal DNA, and presumably mediates efficient persistence of viral genomes. LANA further represses the transcriptional and proapoptotic activities of the p53 tumor suppressor protein. Here we report on the ORF73 gene of HVS strain C488, which is the positional and structural homolog of KSHV LANA. The ORF73 gene in OMK cells can encode a 62-kDa protein that localizes to the nucleus in a pattern similar to that of LANA. We show that the ORF73 gene product can regulate viral gene expression by acting as a transcriptional modulator of latent and lytic viral promoters. To define the HVS ORF73 function in the background of a replication-competent virus, we constructed a viral mutant that expresses ORF73 under the transcriptional control of a mifepristone (RU-486)-inducible promoter. The HVS ORF73 gene product efficiently suppresses lytic viral replication in permissive cells, indicating that it defines a critical control point between viral persistence and lytic replication

A Molecular Model for the Differential Activation of STAT3 and STAT6 by the Herpesviral Oncoprotein Tip

Constitutive STAT signaling provides growth promoting signals in many forms of malignancy. We performed molecular modeling and molecular dynamics studies of the interaction between the regulatory Src homology 2 (SH2) domains of STAT3 and 6 with phosphorylated peptides of the herpesviral oncoprotein Tip, which facilitates Src kinase mediated STATactivation and T cell proliferation. The studies give insight into the ligand binding specificity of the STAT SH2 domains and provide the first model for the differential activation of STAT3 or STAT6 by two distinct regions of the viral Tip protein. The biological relevance of the modeled interactions was then confirmed by activation studies using corresponding recombinant oncoproteins, and finally by respective recombinant viruses. The functional data give experimental validation of the molecular dynamics study, and provide evidence for the involvement of STAT6 in the herpesvirus induced T cel

Herpesvirus Ateles Tio Can Replace Herpesvirus Saimiri StpC and Tip Oncoproteins in Growth Transformation of Monkey and Human T Cells

Herpesvirus saimiri group C strains are capable of transforming human and simian T-lymphocyte populations to permanent antigen-independent growth. Two viral oncoproteins, StpC and Tip, that are encoded by a single bicistronic mRNA, act in concert to mediate this phenotype. A closely related New World monkey herpesvirus, herpesvirus ateles, transcribes a single spliced mRNA at an equivalent genome locus. The encoded protein, Tio, has sequence homologies to both StpC and Tip. We inserted the tio sequence of herpesvirus ateles strain 73 into a recombinant herpesvirus saimiri C488 lacking its own stpC/tip oncogene. Simian as well as human T lymphocytes were growth transformed by the chimeric Tio-expressing viruses. Thus, a single herpesvirus protein appears to be responsible for the oncogenic effects of herpesvirus ateles

Tip-Stat interactions detected in the molecular dynamics simulations.

<p>Only those interactions that were stable over more than 30% of the simulation time are reported. Residues of Tip are shown in bold and their interacting partners from Stat3/6 are listed in the column below. The letters ‘m’ and ‘s’ denote whether an interaction is formed by the main-chain or side-chain atoms, respectively (e.g. m-s denotes an interaction between the main-chain of the first residue (from Tip) and the side chain of the second residue (from STAT)). ‘hs’ denotes the special situation, in which only the hydrophobic atoms but not the polar part of a sidechain is involved in an interaction. In the top line, the three main interacting regions (I to III) are indicated. Region I comprises the phosphotyrosine itself and the adjacent residue pY+1. Region II comprises residues pY+2 and pY+3. The remaining C-terminal residues (pY+4 to pY+8) constitute region III.</p

Specific inhibition of Src-Tyrosine kinase reduces STAT6 activation.

<p>(A) Tip, Lck and STAT6 transfected 293T cells were treated with PP2 or the non-active control substance PP3 (10 µM in DMSO each). STAT6, pSTAT6, Tip and GAPDH were detected by fluorescent immunoblotting on the same membrane. (B) The ratio of pSTAT6 to total STAT6 was normalized to the GAPDH signal. The ratio of STAT6-pY641 (clone 18, detected with anti mouse Dylight-647), total STAT6 (polyclonal rb-anti-STAT6 (M-20) with anti-rabbit-Dylight-488) was calculated and normalized to the GAPDH loading control (biotinylated goat anti-GAPDH/Strepavidin-Alexa-555).</p

Details of the Tip-STAT interaction in the four investigated complexes.

<p>(A) Tip114-STAT3, (B) Tip127-STAT3, (C) Tip114-STAT6, and (D) Tip127-STAT6. For each complex, three panels are shown that highlight the interactions formed in region I, II, and III. The backbone topology of STAT is shown as cyan ribbon and the interacting residues are shown in ball-and-stick presentation. Tip those residues that bind to STAT are shown in stick presentation and labeled in italics. Polar interactions (hydrogen bonds, salt-bridges) are indicated as dotted red lines.</p