A role for accessory genes rI.-1 and rI.1 in the regulation of lysis inhibition by bacteriophage T4

Lysis inhibition (LIN) is a known feature of the T-even family of bacteriophages. Despite its historical role in the development of modern molecular genetics, many aspects of this phenomenon remain mostly unexplained. The key element of LIN is an interaction between two phage-encoded proteins, the T holin and the RI antiholin. This interaction is stabilized by RIII. In this report, we demonstrate the results of genetic experiments which suggest a synergistic action of two accessory proteins of bacteriophage T4, RI.-1, and RI.1 with RIII in the regulation of LIN

Arterivirus Nsp1 Modulates the Accumulation of Minus-Strand Templates to Control the Relative Abundance of Viral mRNAs

The gene expression of plus-strand RNA viruses with a polycistronic genome depends on translation and replication of the genomic mRNA, as well as synthesis of subgenomic (sg) mRNAs. Arteriviruses and coronaviruses, distantly related members of the nidovirus order, employ a unique mechanism of discontinuous minus-strand RNA synthesis to generate subgenome-length templates for the synthesis of a nested set of sg mRNAs. Non-structural protein 1 (nsp1) of the arterivirus equine arteritis virus (EAV), a multifunctional regulator of viral RNA synthesis and virion biogenesis, was previously implicated in controlling the balance between genome replication and sg mRNA synthesis. Here, we employed reverse and forward genetics to gain insight into the multiple regulatory roles of nsp1. Our analysis revealed that the relative abundance of viral mRNAs is tightly controlled by an intricate network of interactions involving all nsp1 subdomains. Distinct nsp1 mutations affected the quantitative balance among viral mRNA species, and our data implicate nsp1 in controlling the accumulation of full-length and subgenome-length minus-strand templates for viral mRNA synthesis. The moderate differential changes in viral mRNA abundance of nsp1 mutants resulted in similarly altered viral protein levels, but progeny virus yields were greatly reduced. Pseudorevertant analysis provided compelling genetic evidence that balanced EAV mRNA accumulation is critical for efficient virus production. This first report on protein-mediated, mRNA-specific control of nidovirus RNA synthesis reveals the existence of an integral control mechanism to fine-tune replication, sg mRNA synthesis, and virus production, and establishes a major role for nsp1 in coordinating the arterivirus replicative cycle

Latrunculin alters the actin-monomer subunit interface to prevent polymerization

Latrunculin-A is a drug that is capable of rapidly, reversibly and specifically disrupting the actin cytoskeleton. The efficacy of its action has made it a compound of choice in many cell-biology laboratories, supplanting the classic actin-depolymerizing drug cytochalasin-D. One reason for this is that the mode of action of latrunculin seems to be less complex than that of cytochalasin. Whereas the latter affects the kinetics of actin-filament polymerization at both the barbed and pointed ends, latrunculin-A seems to associate only with actin monomers, thereby preventing them from repolymerizing into filaments. The association of latrunculin with monomeric, rather than filamentous, actin gave us the opportunity to further our understanding of this interaction by detailed structural analysis of actin monomers using crystallographic techniques. Here we show the first high-resolution structure of an actin-disrupting drug in association with actin and discuss how its interactions with actin, and the conformational changes that its binding causes, may explain its mode of action within the cell