A prophage encoded ribosomal RNA methyltransferase regulates the virulence of Shiga-toxin-producing Escherichia coli (STEC).

Shiga toxin (Stx) released by Shiga toxin producing Escherichia coli (STEC) causes life-threatening illness. Its production and release require induction of Stx-encoding prophage resident within the STEC genome. We identified two different STEC strains, PA2 and PA8, bearing Stx-encoding prophage whose sequences primarily differ by the position of an IS629 insertion element, yet differ in their abilities to kill eukaryotic cells and whose prophages differ in their spontaneous induction frequencies.

The Oligosaccharide Region of LPS Governs Predation of E. coli by the Bacterivorous Protist, Acanthamoeba castellanii.

Protozoan predation is a major cause of bacterial mortality. The first step of predation for phagocytic amoebae is the recognition of their prey. Lipopolysaccharide (LPS) is a major component of Gram-negative bacteria and is only present on the outer leaflet of the outer membrane lipid bilayer.

Small molecule MMRi62 targets MDM4 for degradation and induces leukemic cell apoptosis regardless of p53 status.

MDM2 and MDM4 proteins are key negative regulators of tumor suppressor p53. MDM2 and MDM4 interact their RING domains and form a heterodimer polyubiquitin E3 ligase essential for p53 degradation. MDM4 also forms heterodimer E3 ligases with MDM2 isoforms that lack p53-binding domains, which regulate p53 and MDM4 stability.

Carriage of Shiga toxin phage profoundly affects Escherichia coli gene expression and carbon source utilization.

Background: Enterohemorrhagic Escherichia coli (E. coli) are intestinal pathogenic bacteria that cause life-threatening disease in humans. Their cardinal virulence factor is Shiga toxin (Stx), which is encoded on lambdoid phages integrated in the chromosome.

Evolution of STEC virulence: Insights from the antipredator activities of Shiga toxin producing E. coli.

Shiga toxin-producing Escherichia coli (STEC) are a diverse group of strains that are implicated in over 270,000 cases of human illness annually in the United States alone. Shiga toxin (Stx), encoded by a resident temperate lambdoid bacteriophage, is the main STEC virulence factor. Although the population structure of E.

Molecular Mechanisms Governing "Hair-Trigger" Induction of Shiga Toxin-Encoding Prophages.

Shiga toxin (Stx)-encoding (STEC) strains are responsible for sporadic outbreaks of food poisoning dating to 1982, when the first STEC strain, O157:H7, was isolated. Regardless of STEC serotype, the primary symptoms of STEC infections are caused by Stx that is synthesized from genes resident on lambdoid prophage present in STEC. Despite similar etiology, the severity of STEC-mediated disease varies by outbreak.

Cheating, facilitation and cooperation regulate the effectiveness of phage-encoded exotoxins as antipredator molecules.

Temperate phage encoded Shiga toxin (Stx) kills the bacterivorous predator, Tetrahymena thermophila, providing Stx Escherichia coli with a survival advantage over Stx cells. Although bacterial death accompanies Stx release, since bacteria grow clonally the fitness benefits of predator killing accrue to the kin of the sacrificed organism, meaning Stx-mediated protist killing is a form of self-destructive cooperation. We show here that the fitness benefits of Stx production are not restricted to the kin of the phage-encoding bacteria.

Tetrahymena phagocytic vesicles as ecological micro-niches of phage transfer.

The microbial communities in natural environments such as soil, pond water, or animal rumens are composed of a diverse mixture of bacteria and protozoa including ciliates or flagellates. In such microbiomes, a major source of bacterial mortality is grazing by phagocytic protists. Many protists are omnivorous heterotrophs, feeding on a range of different bacterial species.

Mechanisms that Determine the Differential Stability of Stx⁺ and Stx(-) Lysogens.

Phages 933W, BAA2326, 434, and λ are evolutionarily-related temperate lambdoid phages that infect Escherichia coli. Although these are highly-similar phages, BAA2326 and 933W naturally encode Shiga toxin 2 (Stx⁺), but phage 434 and λ do not (Stx(-)). Previous reports suggest that the 933W Stx⁺ prophage forms less stable lysogens in E.

Determinants that govern the recognition and uptake of Escherichia coli O157 : H7 by Acanthamoeba castellanii.

Predation by phagocytic predators is a major source of bacterial mortality. The first steps in protozoan predation are recognition and consumption of their bacterial prey. However, the precise mechanisms governing prey recognition and phagocytosis by protists, and the identities of the molecular and cellular factors involved in these processes are, as yet, ill-characterized.

A Role for Autoinhibition in Preventing Dimerization of the Transcription Factor ETS1.

ETS1 is the archetype of the ETS transcription factor (TF) family. ETS TFs share a DNA-binding domain, the ETS domain. All ETS TFs recognize a core GGA(A/T) binding site, and thus ETS TFs are found to redundantly regulate the same genes.

Specific minor groove solvation is a crucial determinant of DNA binding site recognition.

The DNA sequence preferences of nearly all sequence specific DNA binding proteins are influenced by the identities of bases that are not directly contacted by protein. Discrimination between non-contacted base sequences is commonly based on the differential abilities of DNA sequences to allow narrowing of the DNA minor groove. However, the factors that govern the propensity of minor groove narrowing are not completely understood.

The Trojan Horse of the microbiological arms race: phage-encoded toxins as a defence against eukaryotic predators.

Phage-encoded Shiga toxin (Stx) acts as a bacterial defence against the eukaryotic predator Tetrahymena. To function as an effective bacterial anti-predator defence, Stx must kill a broad spectrum of predators. Consistent with that assertion, we show here that bacterially encoded Stx efficiently kills the bacteriovore Acanthamoeba castellanii in co-culture.

The microcosm mediates the persistence of shiga toxin-producing Escherichia coli in freshwater ecosystems.

Water is a major route for infection of humans by exotoxin-producing bacteria, including Shiga toxin-producing Escherichia coli (STEC). While STEC has the potential to be present in nearly every type of water source, its distribution is sporadic, and an understanding of factors that govern its emergence and persistence within water is lacking. In this study, we examined the influence of microbe content on STEC persistence in freshwater.

Bacteriophage 434 Hex protein prevents recA-mediated repressor autocleavage.

In a λ(imm434) lysogen, two proteins are expressed from the integrated prophage. Both are encoded by the same mRNA whose transcription initiates at the P(RM) promoter. One protein is the 434 repressor, needed for the establishment and maintenance of lysogeny.

Entry and killing of Tetrahymena thermophila by bacterially produced Shiga toxin.

Unlabelled: Phage-encoded Shiga toxin (Stx) acts as a bacterial defense against the eukaryotic predator Tetrahymena thermophila. It is unknown how Stx enters Tetrahymena protozoa or how it kills them. Tetrahymena protozoa are phagocytotic; hence, Stx could gain entry to the cytoplasm through the oral apparatus or via endocytosis.

Indirect readout of DNA sequence by p22 repressor: roles of DNA and protein functional groups in modulating DNA conformation.

The repressor of bacteriophage P22 (P22R) discriminates between its various DNA binding sites by sensing the identity of non-contacted base pairs at the center of its binding site. The "indirect readout" of these non-contacted bases is apparently based on DNA's sequence-dependent conformational preferences. The structures of P22R-DNA complexes indicate that the non-contacted base pairs at the center of the binding site are in the B' state.

Determinants of bacteriophage 933W repressor DNA binding specificity.

We reported previously that 933W repressor apparently does not cooperatively bind to adjacent sites on DNA and that the relative affinities of 933W repressor for its operators differ significantly from that of any other lambdoid bacteriophage. These findings indicate that the operational details of the lysis-lysogeny switch of bacteriophage 933W are unique among lambdoid bacteriophages. Since the functioning of the lysis-lysogeny switch in 933W bacteriophage uniquely and solely depends on the order of preference of 933W repressor for its operators, we examined the details of how 933W repressor recognizes its DNA sites.

Shiga toxin: expression, distribution, and its role in the environment.

In this review, we highlight recent work that has increased our understanding of the production and distribution of Shiga toxin in the environment. Specifically, we review studies that offer an expanded view of environmental reservoirs for Shiga toxin producing microbes in terrestrial and aquatic ecosystems. We then relate the abundance of Shiga toxin in the environment to work that demonstrates that the genetic mechanisms underlying the production of Shiga toxin genes are modified and embellished beyond the classical microbial gene regulatory paradigms in a manner that apparently "fine tunes" the trigger to modulate the amount of toxin produced.

The lysis-lysogeny decision of bacteriophage 933W: a 933W repressor-mediated long-distance loop has no role in regulating 933W P(RM) activity.

Our data show that unlike bacteriophage λ, repressor bound at O(L) of bacteriophage 933W has no role in regulation of 933W repressor occupancy of 933W O(R)3 or the transcriptional activity of 933W P(RM). This finding suggests that a cooperative long-range loop between repressor tetramers bound at O(R) and O(L) does not form in bacteriophage 933W. Nonetheless, 933W forms lysogens, and 933W prophage display a threshold response to UV induction similar to related lambdoid phages.

Sequence recognition of DNA by protein-induced conformational transitions.

The binding of proteins to specific sequences of DNA is an important feature of virtually all DNA transactions. Proteins recognize specific DNA sequences using both direct readout (sensing types and positions of DNA functional groups) and indirect readout (sensing DNA conformation and deformability). Previously we showed that the P22 c2 repressor N-terminal domain (P22R NTD) forces the central non-contacted 5'-ATAT-3' sequence of the DNA operator into the B' state, a state known to affect DNA hydration, rigidity and bending.

Characterization of GPIT-1 and GPIT-2, two auxiliary components of the Neurospora crassa GPI transamidase complex.

The glycosylphosphatidylinositol (GPI) transamidase contains five known subunits and functions in the lumen of the ER to produce GPI-anchored proteins. The transamidase cleaves proteins containing a GPI anchor attachment signal at their C terminus and generates an amide bond between the newly generated carboxyl terminus of the protein and a GPI anchor. We have identified and characterized GPIT-1 and GPIT-2, two of the transamidase subunits from Neurospora crassa.

Shiga toxin as a bacterial defense against a eukaryotic predator, Tetrahymena thermophila.

Bacterially derived exotoxins kill eukaryotic cells by inactivating factors and/or pathways that are universally conserved among eukaryotic organisms. The genes that encode these exotoxins are commonly found in bacterial viruses (bacteriophages). In the context of mammals, these toxins cause diseases ranging from cholera to diphtheria to enterohemorrhagic diarrhea.

P22 c2 repressor-operator complex: mechanisms of direct and indirect readout.

The P22 c2 repressor protein (P22R) binds to DNA sequence-specifically and helps to direct the temperate lambdoid bacteriophage P22 to the lysogenic developmental pathway. We describe the 1.6 A X-ray structure of the N-terminal domain (NTD) of P22R in a complex with a DNA fragment containing the synthetic operator sequence [d(ATTTAAGATATCTTAAAT)]2.

Effect of salt shock on stability of lambdaimm434 lysogens.

The affinities of the bacteriophage 434 repressor for its various binding sites depend on the type and/or concentration of monovalent cations. The ability of bacteriophage 434 repressor to govern the lysis-lysogeny decision depends on the DNA binding activities of the phage's cI repressor protein. We wished to determine whether changes in the intracellular ionic environment influence the lysis-lysogeny decision of the bacteriophage lambda(imm434).