Increased productivity of <Emphasis Type="Italic">Clostridium acetobutylicum</Emphasis> fermentation of acetone,butanol, and ethanol by pervaporation through supported ionic liquid membrane

Pervaporation proved to be one of the best methods to remove solvents out of a solvent producing Clostridium acetobutylicum culture. By using an ionic liquid (IL)-polydimethylsiloxane (PDMS) ultrafiltration membrane (pore size 60 nm), we could guarantee high stability and selectivity during all measurements carried out at 37°C. Overall solvent productivity of fermentation connected with continuous product removal by pervaporation was 2.34 g l⁻¹ h⁻¹. The supported ionic liquid membrane (SILM) was impregnated with 15 wt% of a novel ionic liquid (tetrapropylammonium tetracyano-borate) and 85 wt% of polydimethylsiloxane. Pervaporation, accomplished with the optimized SILM, led to stable and efficient removal of the solvents butan-1-ol and acetone out of a C. acetobutylicum culture. By pervaporation through SILM, we removed more butan-1-ol than C. acetobutylicum was able to produce. Therefore, we added an extra dose of butan-1-ol to run fermentation on limiting values where the bacteria would still be able to survive its lethal concentration (15.82 g/l). After pervaporation was switched off, the bacteria died from high concentration of butan-1-ol, which they produced.     

Accommodation of powdery mildew fungi in intact plant cells

Parasitic powdery mildew fungi have to overcome basic resistance and manipulate host cells to establish a haustorium as a functional feeding organ in a host epidermal cell. Currently, it is of central interest how plant factors negatively regulate basal defense or whether they even support fungal development in compatible interactions. Additionally, creation of a metabolic sink in infected cells may involve host activity. Here, we review the current progress in understanding potential fungal targets for host reprogramming and nutrient acquisition.     

Cloning and characterization of a salt stress-inducible small GTPase gene from the model grass species Lolium temulentum

A gene encoding a small guanosine triphosphate (GTP)-binding protein (smGTP) related to the Rab2 gene family of GTPases was identified during the analysis of a salt stress suppression subtractive hybridization (SSH) expression library from the model grass species Lolium temulentum L. (Darnel ryegrass). The smGTP gene was found to have a low-level constitutive expression and was strongly induced by salt stress in root, crown and leaf tissues. The expression pattern of the smGTP gene was compared against two additional stress genes identified in the SSH expression library, the well-characterized dehydration stress tolerance gene, delta 1-pyrroline-5-carboxylate synthetase (P5CS) encoding for a key enzyme in proline biosynthesis, and the cold response gene COR413. The genes were analyzed for their response to salinity as well as their responses to 7 different forms of abiotic stress in L. temulentum plants. The smGTP gene displayed an expression pattern similar to the P5CS gene, suggesting a role in dehydration stress. In contrast, the COR413 gene was found to be up-regulated in response to all stresses tested and has utility as a general stress marker in grass plants.     

Self-recognition by an intrinsically disordered protein

The intrinsically disordered translocation domain (T-domain) of the protein antibiotic colicin N binds to periplasmic receptors of target Escherichia coli cells in order to penetrate their inner membranes. We report here that the specific 27 consecutive residues of the T-domain of colicin N known to bind to the helper protein TolA in target cells also interacts intramolecularly with folded regions of colicin N. We suggest that this specific self-recognition helps intrinsically disordered domains to bury their hydrophobic recognition motifs and protect them against degradation, showing that an impaired self-recognition leads to increased protease susceptibility.     

Staphylococcal peptidoglycan initiates an inflammatory response and procoagulant activity in human vascular endothelial cells: a comparison with highly purified lipoteichoic acid and TSST-1

Staphylococcus aureus is one of the most significant pathogens in human sepsis and endocarditis. A hallmark of these endovascular S. aureus infections is that the coagulation system is triggered by a tissue factor (TF)-dependent pathway. This study demonstrates that highly purified S. aureus peptidoglycan, lipoteichoic acid (LTA) and TSST-1 increase TF mRNA and TF surface protein in human umbilical vein endothelial cells (ECs). Concomitantly, peptidoglycan- and LTA-activated ECs express significant TF-dependent procoagulant activity (TF PCA). In addition peptidoglycan, but not LTA or TSST-1, induced surface expression of the EC inflammation markers ICAM-1 and VCAM-1, which supported the adhesion of monocytes to these ECs. During the coculture of peptidoglycan-activated ECs and adherent monocytes, a marked additional increase of TF PCA was observed. Marginal increases in TF PCA were observed in cocultures of monocytes with LTA- or TSST-1-activated ECs. This study defines in particular staphylococcal peptidoglycan, previously known as a potent initiator of TF PCA in monocytes, as also being an activator of a coagulant response in human ECs that is further intensified by the presence of surface-bound monocytes.     

A dual requirement for Iroquois genes during Xenopus kidney development

The Iroquois (Irx) genes encode evolutionary conserved homeoproteins. We report that Xenopus genes Irx1 and Irx3 are expressed and required during different stages of Xenopus pronephros development. They are initially expressed during mid-neurulation in domains extending over most of the prospective pronephric territory. Expression onset takes place after kidney anlage specification, but before pronephric organogenesis occurs. Later, during nephron segmentation, expression becomes restricted to the intermediate tubule region of the proximal-distal axis. Loss- and gain-of-function analyses, performed with specific morpholinos and inducible wild-type and dominant-negative constructs, reveal a dual requirement for Irx1 and Irx3 during pronephros development. During neurula stages, these genes maintain the specification of the pronephric territory and define its size. This seems to occur, at least in part, through positive regulation of Bmp signalling. Subsequently, Irx genes are required for proper formation of the intermediate tubule. Finally, we find that retinoic acid signalling activates both Irx1 and Irx3 genes in the pronephros.     

Models of toxic beta-sheet channels of protegrin-1 suggest a common subunit organization motif shared with toxic alzheimer beta-amyloid ion channels

Antimicrobial peptides (AMPs) induce cytotoxicity by altering membrane permeability. The electrical properties of membrane-associated AMPs as well as their cellular effects have been extensively documented; however their three-dimensional structure is poorly understood. Gaining insight into channel structures is important to the understanding of the protegrin-1 (PG-1) and other AMP cytolytic mechanisms, and to antibiotics design. We studied the β-sheet channels morphology using molecular dynamics simulations. We modeled PG-1 channels as intrinsic barrel-stave and toroidal membrane pores, and simulated them in zwitterionic and anionic lipid bilayers. PG-1 channels consist of eight β-hairpins in a consecutive NCCN (N and C represent the β-hairpin's N- and C-termini) packing organization yielding antiparallel and parallel β-sheet channels. Both channels preserve the toroidal, but not the barrel-stave pores. The two lipid leaflets of the bilayer bend toward each other at the channels’ edges, producing a semitoroidal pore with the outward-pointing hydrophobic residues preventing the polar lipid headgroups from moving to the bilayer center. In all simulated lipid environments, PG-1 channels divide into four or five β-sheet subunits consisting of single or dimeric β-hairpins. The channel morphology with subunit organization is consistent with the four to five subunits observed by NMR in the POPE/POPG bilayer. Remarkably, a β-sheet subunit channel motif is in agreement with Alzheimer ion channels modeled using the universal U-shape β-strand-turn-β-strand structure, as well as with high resolution atomic force microscopy images of β-amyloid channels with four to six subunits. Consistent with the toxic β-amyloid channels that are ion-conducting, the PG-1 channels permeate anions.     

Small RNAs encoded within genetic islands of Salmonella typhimurium show host-induced expression and role in virulence

The emergence of pathogenic strains of enteric bacteria and their adaptation to unique niches are associated with the acquisition of foreign DNA segments termed ‘genetic islands’. We explored these islands for the occurrence of small RNA (sRNA) encoding genes. Previous systematic screens for enteric bacteria sRNAs were mainly carried out using the laboratory strain Escherichia coli K12, leading to the discovery of ~80 new sRNA genes. These searches were based on conservation within closely related members of enteric bacteria and thus, sRNAs, unique to pathogenic strains were excluded. Here we describe the identification and characterization of 19 novel unique sRNA genes encoded within the ‘genetic islands’ of the virulent strain Salmonella typhimurium. We show that the expression of many of the island-encoded genes is associated with stress conditions and stationary phase. Several of these sRNA genes are induced when Salmonella resides within macrophages. One sRNA, IsrJ, was further examined and found to affect the translocation efficiency of virulence-associated effector proteins into nonphagocytic cells. In addition, we report that unlike the majority of the E. coli sRNAs that are trans regulators, many of the island-encoded sRNAs affect the expression of cis-encoded genes. Our study suggests that the island encoded sRNA genes play an important role within the network that regulates bacterial adaptation to environmental changes and stress conditions and thus controls virulence.