Coronavirus Gene 7 Counteracts Host Defenses and Modulates Virus Virulence

Transmissible gastroenteritis virus (TGEV) genome contains three accessory genes: 3a, 3b and 7. Gene 7 is only present in members of coronavirus genus a1, and encodes a hydrophobic protein of 78 aa. To study gene 7 function, a recombinant TGEV virus lacking gene 7 was engineered (rTGEV-Δ7). Both the mutant and the parental (rTGEV-wt) viruses showed the same growth and viral RNA accumulation kinetics in tissue cultures. Nevertheless, cells infected with rTGEV-Δ7 virus showed an increased cytopathic effect caused by an enhanced apoptosis mediated by caspase activation. Macromolecular synthesis analysis showed that rTGEV-Δ7 virus infection led to host translational shut-off and increased cellular RNA degradation compared with rTGEV-wt infection. An increase of eukaryotic translation initiation factor 2 (eIF2α) phosphorylation and an enhanced nuclease, most likely RNase L, activity were observed in rTGEV-Δ7 virus infected cells. These results suggested that the removal of gene 7 promoted an intensified dsRNA-activated host antiviral response. In protein 7 a conserved sequence motif that potentially mediates binding to protein phosphatase 1 catalytic subunit (PP1c), a key regulator of the cell antiviral defenses, was identified. We postulated that TGEV protein 7 may counteract host antiviral response by its association with PP1c. In fact, pull-down assays demonstrated the interaction between TGEV protein 7, but not a protein 7 mutant lacking PP1c binding motif, with PP1. Moreover, the interaction between protein 7 and PP1 was required, during the infection, for eIF2α dephosphorylation and inhibition of cell RNA degradation. Inoculation of newborn piglets with rTGEV-Δ7 and rTGEV-wt viruses showed that rTGEV-Δ7 virus presented accelerated growth kinetics and pathology compared with the parental virus. Overall, the results indicated that gene 7 counteracted host cell defenses, and modified TGEV persistence increasing TGEV survival. Therefore, the acquisition of gene 7 by the TGEV genome most likely has provided a selective advantage to the virus

Lymphocyte Chemotaxis Is Regulated by Histone Deacetylase 6, Independently of Its Deacetylase Activity

In this work, the role of HDAC6, a type II histone deacetylase with tubulin deacetylase activity, in lymphocyte polarity, motility, and transmigration was explored. HDAC6 was localized at dynamic subcellular structures as leading lamellipodia and the uropod in migrating T-cells. However, HDAC6 activity did not appear to be involved in the polarity of migrating lymphocytes. Overexpression of HDAC6 in freshly isolated lymphocytes and T-cell lines increased the lymphocyte migration mediated by chemokines and their transendothelial migration under shear flow. Accordingly, the knockdown of HDAC6 expression in T-cells diminished their chemotactic capability. Additional experiments with HDAC6 inhibitors (trichostatin, tubacin), other structural related molecules (niltubacin, MAZ-1391), and HDAC6 dead mutants showed that the deacetylase activity of HDAC6 was not involved in the modulatory effect of this molecule on cell migration. Our results indicate that HDAC6 has an important role in the chemotaxis of T-lymphocytes, which is independent of its tubulin deacetylase activity