Data information: s, S-layer; m, membrane; c, conical structure; a, archaella; r, ring. Scale bars, 100 nm; scale bar in (A) applies to (1-3); scale bar in (B) applies to (B-E). EMBO reportsª 2017 The Authors EMBO reportsStructure of an archaellar motor and cone Ariane Briegel et al EV1 Figure EV2. Double-cone structure observed in Thermococcus kodakaraensis.A tomographic slice through a side view shows two associated conical structures (c1 and c2), both associated with archaella (a). s, S-layer; m, membrane. Scale bar, 100 nm. A, B Tomographic slices through two cells, highlighting the association between the cone and the archaella. C, D 3D segmentations of the cells in (A) and (B), respectively, with cones in blue and archaella in red, embedded in tomographic slices. E Tomographic slices of individual archaella show the varying orientations of archaella with respect to the cell envelope, as well as apparent connections to the cone.Data information: Scale bars, 100 nm in (A) and (B), 50 nm in (E) (applies to all panels in E); segmentations not to scale. Figure EV4. Individual particles from the subtomogram average show heterogeneity in the L3 density and angle of cone density. EMBO reportsª 2017 The Authors EMBO reports Structure of an archaellar motor and cone Ariane Briegel et al EV3The L3 density appears as either two dots of similar (first two panels) or different intensity (third panel), a single dot (fourth panel), or a dot and an extended line (fifth panel). m, membrane; c, conical structure. Scale bar, 10 nm (applies to all panels).ª 2017 The Authors EMBO reportsAriane Briegel et al Structure of an archaellar motor and cone EMBO reports EV4

The sucrose nonfermenting 1 (SNF1) protein kinase of yeast plays a central role in the transcription of glucose-repressible genes in response to glucose starvation. In this study, we deleted an ortholog of SNF1 from Gibberella zeae to characterize its functions by using a gene replacement strategy. The mycelial growth of deletion mutants (⌬GzSNF1) was reduced by 21 to 74% on diverse carbon sources. The virulence of ⌬GzSNF1 mutants on barley decreased, and the expression of genes encoding cell-wall-degrading enzymes was reduced. The most distinct phenotypic changes were in sexual and asexual development. ⌬GzSNF1 mutants produced 30% fewer perithecia, which matured more slowly, and asci that contained one to eight abnormally shaped ascospores. Mutants in which only the GzSNF1 catalytic domain was deleted had the same phenotype changes as the ⌬GzSNF1 strains, but the phenotype was less extreme in the mutants with the regulatory domain deleted. In outcrosses between the ⌬GzSNF1 mutants, each perithecium contained ϳ70% of the abnormal ascospores, and ϳ50% of the asci showed unexpected segregation patterns in a single locus tested. The asexual spores of the ⌬GzSNF1 mutants were shorter and had fewer septa than those of the wild-type strain. The germination and nucleation of both ascospores and conidia were delayed in ⌬GzSNF1 mutants in comparison with those of the wild-type strain. GzSNF1 expression and localization depended on the developmental stage of the fungus. These results suggest that GzSNF1 is critical for normal sexual and asexual development in addition to virulence and the utilization of alternative carbon sources.

Antibiotic resistance and tolerance are increasing threats to global health as antibiotic-resistant bacteria can cause severe morbidity and mortality and can increase treatment cost 10-fold. Although several genes contributing to antibiotic tolerance among pneumococci have been identified, we report here that ClpL, a major heat shock protein, could modulate cell wall biosynthetic enzymes and lead to decreased penicillin susceptibility. On capsular type 1, 2, and 19 genetic backgrounds, mutants lacking ClpL were more susceptible to penicillin and had thinner cell walls than the parental strains, whereas a ClpL-overexpressing strain showed a higher resistance to penicillin and a thicker cell wall. Although exposure of Streptococcus pneumoniae D39 to penicillin inhibited expression of the major cell wall synthesis gene pbp2x, heat shock induced a ClpLdependent increase in the mRNA levels and protein synthesized by pbp2x. Inducible ClpL expression correlated with PBP2x expression and penicillin susceptibility. Fractionation and electron micrograph data revealed that ClpL induced by heat shock is localized at the cell wall, and the ⌬clpL showed significantly reduced net translocation of PBP2x into the cell wall. Moreover, coimmunoprecipitation with either ClpL or PBP2x antibody followed by reprobing with ClpL or PBP2x antibody showed an interaction between ClpL and PBP2x after heat stress. This interaction was confirmed by His tag pulldown assay with either ClpLHis 6 or PBP2xHis 6 . Thus, ClpL stabilized pbp2x expression, interacted with PBP2x, and facilitated translocation of PBP2x, a key protein of cell wall synthesis process, contributing to the decrease of antibiotic susceptibility in S. pneumoniae.

The 5' region of Potato virus X (PVX) RNA containing an AC-rich single-stranded region and stem-loop 1 (SL1) has been shown to be important for PVX replication (Miller, E.D., Plante, C.A., Kim, K.-H., Brown, J.W., Hemenway, C., 1998. Stem-loop structure in the 5' region of potato virus X genome required for plus-strand RNA accumulation. J. Mol. Biol. 284, 591-608.). Here, we describe the involvement of SL1 for binding to the PVX coat protein (CP) using an in vitro assembly system and various deletion mutants of the 5' region of PVX RNA. Internal and 5' terminal deletions of the 5'-nontranslated region of PVX RNA were assessed for their effects on formation of assembled virus-like particles (VLPs). Mutant RNAs that contain the top region of SL1 or sequences therein bound to CP to form VLPs. In contrast, transcripts of mutants that disrupt SL1 RNA structure were unable to form VLPs. SELEX was used to further confirm the specific RNA recognition of PVX CP using RNA transcripts containing randomized sequences of the upper portion of SL1. Wild-type (wt) sequences along with many other sequences that resemble SL1 structure were selected after fourth and fifth rounds of SELEX (27.0% and 44.4%, respectively). RNA transcripts from several SELEX winners that are predicted to form stable stem-loop structures very closely resembling wt PVX SL1 VLPs. RNA transcripts not predicted to form secondary structures similar to SL1 did not form VLPs in vitro. Taken together, our results suggest that RNA secondary structural elements within SL1 and/or sequences therein are crucial for formation of VLPs and are required for the specific recognition by the CP subunit.

^Äëíê~Åí= To test the impact of commercial nanomaterials on the environment and biological species, the potential eco-toxicity of nano sized silver powder was investigated with bacteria, Gram-negative bëÅÜÉêáÅÜá~=Åçäá and Gram-positive _~Åáääìë=ëìÄJ íáäáë, and yeast, p~ÅÅÜ~êçãóÅÉë=ÅÉêÉîáëá~É. When commercial silver nanopowder was dispersed in water, it was shown to contain both silver nanoparticles and silver ions. S. cerevisiae showed a higher survival rate than the other two species.To compare the antimicrobial activity as a quantitative parameter, the susceptibility constant was determined. The susceptibility constant of the silver ions were larger than those of the silver nanopowder. On average, the degree of susceptibility to silver decreased in the following order, bK=Åçäá=>=_K=ëìÄíáäáë=>=pK=ÅÉêÉîáëá~É. © KSBB hÉóïçêÇëW=ëáäîÉê=å~åçé~êíáÅäÉëI=ëáäîÉê=áçåëI=ÅóíçíçñáÅáíóI=~åíáÄ~ÅíÉêá~ä=~Åíáîáíó