3Outer membrane proteins (OMPs) play important roles in Gram-negative bacteria, mitochondria and chloroplasts in nutrition transport, protein import, secretion, and other fundamental biological processes [1][2][3] . Dysfunction of mitochondria outer membrane proteins are linked to disorders such as diabetes, Parkinsons and other neurodegenerative diseases 4,5 . The OMPs are inserted and folded correctly into the outer membrane (OM) by the conserved OMP85 family proteins [6][7][8] , suggesting that similar insertion mechanisms may be used in Gram-negative bacteria, mitochondria and chloroplasts.In Gram-negative bacteria, OMPs are synthesized in the cytoplasm, and are transported across the inner membrane by SecYEG into the periplasm 8,9 . The seventeen kilodalton (kDa) protein (Skp) and the survival factor A (SurA) chaperones escort the unfolded OMPs across the periplasm to the β-barrel assembly machinery (BAM), which is responsible for insertion and assembly of OMPs into the OM 10-12 . InEscherichia coli, the BAM complex consists of BamA and four lipoprotein subunits, BamB, BamC, BamD and BamE. BamA is comprised of five N-terminal polypeptide transport-associated (POTRA) domains and a C-terminal OMP transmembrane barrel, while the four lipoproteins are affixed to the membrane by N-terminal lipid-modified cysteines. Of these subunits, BamA and BamD are essential 3,6 . One copy of each of these five proteins is required to form the BAM complex with an approximate molecular weight of 200 kDa (Extended Data Fig. 1). In vitro reconstitution of the E.coli BAM complex and functional assays showed that all five subunits are required to obtain the maximum activity of BAM [13][14][15][16] . Furthermore, comparison of the two complexes reveals that the periplasmic units are rotated with respect to the barrel, which appears to be linked to significant conformational changes in the β-strands β1C-β6C of the barrel. Taken together this suggests a novel insertion mechanism whereby rotation of the BAM periplasmic ring promotes insertion of OMPs into the OM. To our knowledge, this is the first reported crystal structure of an intramembrane barrel with a lateral-open conformation.Unique architecture of two E. coli BAM complexes X-ray diffraction data of selenomethionine labelled crystals were collected to 3.9Ångström (Å) resolution and the BAM structure was determined by singlewavelength anomalous dispersion (SAD) and manual molecular replacement (Methods, Extended Data Table 1). The first structure contained four proteins: BamA, BamC, BamD and BamE (Fig. 1a-c), with the electron density and crystal packing indicating that the BamB is absent in the complex. This was confirmed by SDS-PAGE analysis of the crystals (Extended Data Fig. 1 and Supplementary Data Fig. S1). In this model, BamA, BamC, BamD and BamE contain residues E22-I806, C25-K344, E26-S243, and C20-E110, respectively. The machinery is approximately 115 Å in length, 84 Å in width and 132 Å in height (Fig. 1a). 5The architecture of BamACDE resembles a top hat with a...

SUMMARY Several lines of evidence suggest that mitochondrial dysfunction plays a critical role in the pathogenesis of microvascular complications of diabetes, including diabetic nephropathy. However, the signaling pathways by which hyperglycemia leads to mitochondrial dysfunction are not fully understood. Here we examined the role of Rho-associated coiled-coil containing protein kinase 1 (ROCK1) on mitochondrial dynamics by generating two diabetic mouse models with targeted deletions of ROCK1, and an inducible podocyte-specific knock-in mouse expressing a constitutively active (cA) mutant of ROCK1. Our findings suggest that ROCK1 mediates hyperglycemia-induced mitochondrial fission by promoting dynamin-related protein-1 (Drp1) recruitment to the mitochondria. Deletion of ROCK1 in diabetic mice prevented mitochondrial fission, whereas podocyte-specific cA-ROCK1 mice exhibited increased mitochondrial fission. Importantly, we found that ROCK1 triggers mitochondrial fission by phosphorylating Drp1 at Serine 600 residue. These findings provide insights into the unexpected role of ROCK1 in a signaling cascade that regulates mitochondrial dynamics.

Summary Lassa fever virus (LASV) causes thousands of deaths yearly and is a biological threat agent, for which there is no vaccine and limited therapy1. The nucleoprotein (NP) of LASV plays essential roles in viral RNA synthesis and immune suppression2-6, the molecular mechanisms of which are poorly understood. Here, we report the crystal structure of LASV NP at 1.80 Angstrom resolution, which reveals N- and C-domains with structures unlike any of the reported viral NPs7-10. The N domain folds into a novel structure with a deep cavity for binding the m7GpppN cap structure that is required for viral RNA transcription, whereas the C domain contains 3′-5′ exoribonuclease activity involved in suppressing interferon induction. This is the first X-ray crystal structure solved for an arenaviral NP, which reveals its unexpected functions and suggests unique mechanisms in cap binding and immune evasion. These findings provide great potential for vaccine and drug development.

SummaryHow ion channels are gated to regulate ion flux in and out of cells is the subject of intense interest. The E. coli mechanosensitive channel, MscS, opens to allow rapid ion efflux, relieving the turgor pressure that would otherwise destroy the cell. We present a 3.45 Å resolution structure for the MscS channel in an open conformation. This structure has a pore diameter of ~13 Å created by substantial rotational re-arrangement of the three transmembrane helices. The structure suggests a molecular mechanism that underlies MscS gating and its decay of conductivity during prolonged activation. Support for this mechanism is provided by single channel analysis of mutants with altered gating characteristics.