Unexpected mode of engagement between enterovirus 71 and its receptor SCARB2

Enterovirus 71 (EV71) is a common cause of hand, foot and mouth disease—a disease endemic especially in the Asia-Pacific region1. Scavenger receptor class B member 2 (SCARB2) is the major receptor of EV71, as well as several other enteroviruses responsible for hand, foot and mouth disease, and plays a key role in cell entry2. The isolated structures of EV71 and SCARB2 are known3,4,5,6, but how they interact to initiate infection is not. Here, we report the EV71–SCARB2 complex structure determined at 3.4 Å resolution using cryo-electron microscopy. This reveals that SCARB2 binds EV71 on the southern rim of the canyon, rather than across the canyon, as predicted3,7,8. Helices 152–163 (α5) and 183–193 (α7) of SCARB2 and the viral protein 1 (VP1) GH and VP2 EF loops of EV71 dominate the interaction, suggesting an allosteric mechanism by which receptor binding might facilitate the low-pH uncoating of the virus in the endosome/lysosome. Remarkably, many residues within the binding footprint are not conserved across SCARB2-dependent enteroviruses; however, a conserved proline and glycine seem to be key residues. Thus, although the virus maintains antigenic variability even within the receptor-binding footprint, the identification of binding ‘hot spots’ may facilitate the design of receptor mimic therapeutics less likely to quickly generate resistance

Prevalence of Plasmid-Associated Tetracycline Resistance Genes in Multidrug-Resistant <i>Escherichia coli</i> Strains Isolated from Environmental, Animal and Human Samples in Panama

Antimicrobial resistance bacteria are nowadays ubiquitous. Its presence has been reported in almost every type of source, from water for agricultural and recreative use, water distribution pipes, and wastewater, to food, fomites, and clinical samples. Enterobacteriaceae, especially Escherichia coli, are not the exception, showing an increased resistance to several antibiotics, causing a global health and economic burden. Therefore, the monitoring of fecal microbiota is important because it is present in numerous reservoirs where gene transfer between commensal and virulent bacteria can take place, representing a potential source of resistant E. coli. In this work, antibiotic resistance profiles of 150 E. coli isolates from environmental, animal, and human samples, collected in three rural areas in Panama, were analyzed. A total of 116 isolates were resistant to at least one of the nine antibiotics tested. Remarkably, almost 100% of these exhibited resistance to tetracycline. Plasmid-associated tetA and tetB genes were detected in 42.86% of the isolates analyzed, tetA being the most prevalent. These results suggest that tetracycline resistance would be used as a convenient indicator of genetic horizontal transfer within a community

Prevalence of Plasmid-Associated Tetracycline Resistance Genes in Multidrug-Resistant Escherichia coli Strains Isolated from Environmental, Animal and Human Samples in Panama

Antimicrobial resistance bacteria are nowadays ubiquitous. Its presence has been reported in almost every type of source, from water for agricultural and recreative use, water distribution pipes, and wastewater, to food, fomites, and clinical samples. Enterobacteriaceae, especially Escherichia coli, are not the exception, showing an increased resistance to several antibiotics, causing a global health and economic burden. Therefore, the monitoring of fecal microbiota is important because it is present in numerous reservoirs where gene transfer between commensal and virulent bacteria can take place, representing a potential source of resistant E. coli. In this work, antibiotic resistance profiles of 150 E. coli isolates from environmental, animal, and human samples, collected in three rural areas in Panama, were analyzed. A total of 116 isolates were resistant to at least one of the nine antibiotics tested. Remarkably, almost 100% of these exhibited resistance to tetracycline. Plasmid-associated tetA and tetB genes were detected in 42.86% of the isolates analyzed, tetA being the most prevalent. These results suggest that tetracycline resistance would be used as a convenient indicator of genetic horizontal transfer within a community

Translation Initiation Factors eIF3 and HCR1 Control Translation Termination and Stop Codon Read-Through in Yeast Cells

Translation is divided into initiation, elongation, termination and ribosome recycling. Earlier work implicated several eukaryotic initiation factors (eIFs) in ribosomal recycling in vitro. Here, we uncover roles for HCR1 and eIF3 in translation termination in vivo. A substantial proportion of eIF3, HCR1 and eukaryotic release factor 3 (eRF3) but not eIF5 (a well-defined “initiation-specific” binding partner of eIF3) specifically co-sediments with 80S couples isolated from RNase-treated heavy polysomes in an eRF1-dependent manner, indicating the presence of eIF3 and HCR1 on terminating ribosomes. eIF3 and HCR1 also occur in ribosome- and RNA-free complexes with both eRFs and the recycling factor ABCE1/RLI1. Several eIF3 mutations reduce rates of stop codon read-through and genetically interact with mutant eRFs. In contrast, a slow growing deletion of hcr1 increases read-through and accumulates eRF3 in heavy polysomes in a manner suppressible by overexpressed ABCE1/RLI1. Based on these and other findings we propose that upon stop codon recognition, HCR1 promotes eRF3·GDP ejection from the post-termination complexes to allow binding of its interacting partner ABCE1/RLI1. Furthermore, the fact that high dosage of ABCE1/RLI1 fully suppresses the slow growth phenotype of hcr1? as well as its termination but not initiation defects implies that the termination function of HCR1 is more critical for optimal proliferation than its function in translation initiation. Based on these and other observations we suggest that the assignment of HCR1 as a bona fide eIF3 subunit should be reconsidered. Together our work characterizes novel roles of eIF3 and HCR1 in stop codon recognition, defining a communication bridge between the initiation and termination/recycling phases of translation

Activation mechanism of a noncanonical RNA-dependent RNA polymerase

Two lineages of viral RNA-dependent RNA polymerases (RDRPs) differing in the organization (canonical vs. noncanonical) of the palm subdomain have been identified. Phylogenetic analyses indicate that both lineages diverged at a very early stage of the evolution of the enzyme [Gorbalenya AE, Pringle FM, Zeddam JL, Luke BT, Cameron CE, Kalmakoff J, Hanzlik TN, Gordon KH, Ward VK (2002) J Mol Biol 324:47–62]. Here, we report the x-ray structure of a noncanonical birnaviral RDRP, named VP1, in its free form, bound to Mg2+ ions, and bound to a peptide representing the polymerase-binding motif of the regulatory viral protein VP3. The structure of VP1 reveals that the noncanonical connectivity of the palm subdomain maintains the geometry of the catalytic residues found in canonical polymerases but results in a partial blocking of the active site cavity. The VP1–VP3 peptide complex shows a mode of polymerase activation in which VP3 binding promotes a conformational change that removes the steric blockade of the VP1 active site, facilitating the accommodation of the template and incoming nucleotides for catalysis. The striking structural similarities between birnavirus (dsRNA) and the positive-stranded RNA picornavirus and calicivirus RDRPs provide evidence supporting the existence of functional and evolutionary relationships between these two virus groups

Structural and mechanistic insights into the association of PKCα-C2 domain to PtdIns(4,5)P2

C2 domains are widely-spread protein signaling motifs that in classical PKCs act as Ca2+-binding modules. However, the molecular mechanisms of their targeting process at the plasma membrane remain poorly understood. Here, the crystal structure of PKCα-C2 domain in complex with Ca2+, 1,2-dihexanoyl-sn-glycero-3-[phospho-l-serine] (PtdSer), and 1,2-diayl-sn-glycero-3-[phosphoinositol-4,5-bisphosphate] [PtdIns(4,5)P2] shows that PtdSer binds specifically to the calcium-binding region, whereas PtdIns(4,5)P2 occupies the concave surface of strands β3 and β4. Strikingly, the structure reveals a PtdIns(4,5)P2-C2 domain-binding mode in which the aromatic residues Tyr-195 and Trp-245 establish direct interactions with the phosphate moieties of the inositol ring. Mutations that abrogate Tyr-195 and Trp-245 recognition of PtdIns(4,5)P2 severely impaired the ability of PKCα to localize to the plasma membrane. Notably, these residues are highly conserved among C2 domains of topology I, and a general mechanism of C2 domain-membrane docking mediated by PtdIns(4,5)P2 is presented