An Inverted Repeat in the ospC Operator Is Required for Induction in Borrelia Burgdorferi

Borrelia burgdorferi, the spirochete that causes Lyme disease, differentially regulates synthesis of the outer membrane lipoprotein OspC to infect its host. OspC is required to establish infection but then repressed in the mammal to avoid clearance by the adaptive immune response. Inverted repeats (IR) upstream of the promoter have been implicated as an operator to regulate ospC expression. We molecularly dissected the distal inverted repeat (dIR) of the ospC operator by site-directed mutagenesis at its endogenous location on the circular plasmid cp26. We found that disrupting the dIR but maintaining the proximal IR prevented induction of OspC synthesis by DNA supercoiling, temperature, and pH. Moreover, the base-pairing potential of the two halves of the dIR was more important than the nucleotide sequence in controlling OspC levels. These results describe a cis-acting element essential for the expression of the virulence factor OspC

Live cell fluorescence microscopy to study microbial pathogenesis

Advances in microscopy and fluorescent probes provide new insight into the nanometer-scale biochemistry governing the interactions between eukaryotic cells and pathogens. When combined with mathematical modelling, these new technologies hold the promise of qualitative, quantitative and predictive descriptions of these pathways. Using the light microscope to study the spatial and temporal relationships between pathogens, host cells and their respective biochemical machinery requires an appreciation for how fluorescent probes and imaging devices function. This review summarizes how live cell fluorescence microscopy with common instruments can provide quantitative insight into the cellular and molecular functions of hosts and pathogens.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/72689/1/j.1462-5822.2009.01283.x.pd

International Energy and Natural Resources Law

içinde uyarıcı işlevler kazanır. Savaş öncesinde kabileyi coşturup yüreklendiren öğe, kuşaktan kuşağa destanlarla, baladlarla bilgi taşıyan araç haline dönüşür. Savaş çığlıkları giderek askeri marşlara dönüşmüş, tapınma sırasındaki ezgisel mırıltılar, dinsel ilahileri doğurmuştur. İlk ezgiler ya tanrıları kutlama törenlerine ilişkindir, ya da tanrılara yakın, güçlü kahramanlara övgüdür. Müzik sanatı, tarihin akışı içinde giderek düşünce ve imge gücünü uyarmış, her çağda, her kültürde kendine özgü bir üslup, o toplumun gereksiniminden doğan bir anlatım yolu bulmuştur. “Apollon ve Müzik” adlı bu tezde Apollon Tanrısı’nın Kimliklerinden Müzik Kimliğini ön plana çıkartarak antik müzik ve çalgıları irdelenmiştir. Bu çalışmanın sonunda da Klaros kazısında bulunmuş olan çok sayıdaki Lyralı Apollon Klarios pişmiş toprak figürinlerinin ışığında, Tanrı Apollon’un elindeki kaplumbağa kabuğundan lyra’nın tekrar yapımı amaçlanmıştır ve yapılıp icra edilmiştir. Antik müzik üzerine modern araştırmalar 19. yüzyılın sonlarında başlamıştır. Konu ile ilişkili yayınların tümü antik müziği ve müzik aletlerini incelerken Attika eserleri özellikle de seramikleri üzerindeki betimlerden yararlanmışlardır. Bu betimler, hem arkeologlar hem de müzikologlar açısından büyük bir şanstır. Bu resimler, müzik aletlerinin görünüşü, nasıl çalındığını ( belli bir dereceye kadar), hangi sosyal topluluk içinde ne amaçla çalındığını göstermektedir. Lyra çalgısının tekrar yapımında kaynak olarak, lyranın, heykel, kabartma, pişmiş toprak ve seramikler üzerindeki arkeolojik betimlerden yaralanıldı. Bazı betimlerin yeterince ayrıntılı olmamaları, bazılarında ise ayrıntıların ne kadar gerçekçi olduklarının tam olarak kestirilememesi nedeniyle sorunlar yaşadık. Betimlerin çoğunlukla sadece bir çalgıyı gerçekte olduğu şekilde resmetmek kaygısı ile yapılmadıkları açıktır. Tüm bu olumsuzluklara karşın arkeolojik veriler sayesinde antik dönem kültürlerin çalgılarına dair çıkarımda bulunmak mümkündür. Bu çalışmayı Telli Çalgıların Tarihçesi, Hellen Toplumunda Müziğin ve Lyra’nın Yeri, Çalgıların Sınıflandırılması, Lyra’nın Teknik incelenmesi son olarak ta Lyra ve Apollon İlişkisinin Klaros’taki İkonografik Yansımaları adlı bölümler oluşturmaktadır. Sonuç olarak, çalgı çalınır durumda döneminin müziğini çalar halde bize bir nebzede olsa geniş ufuklar açmasına sebep olmuştur. Yeni kazılarla ortaya çıkacak bulguların yanı sıra, antik dönemde birlikte icra edildiği çalgıların yapılarına ve müzikal niteliklerine ilişkin bilgilerinde artmasıyla, lyra bu çalgı toplulukları içinde veya solo icrasında nasıl bir müzikal işlev üstlenmiş olabileceğinin daha iyi anlaşılması beklenebilir

Ubiquitination of the bacterial inositol phosphatase, SopB, regulates its biological activity at the plasma membrane

The Salmonella type III effector, SopB, is an inositol polyphosphate phosphatase that modulates host cell phospholipids at the plasma membrane and the nascent Salmonella-containing vacuole (SCV). Translocated SopB persists for many hours after infection and is ubiquitinated but the significance of this covalent modification has not been investigated. Here we identify by mass spectrometry six lysine residues of SopB that are mono-ubiquitinated. Substitution of these six lysine residues with arginine, SopB-K6R, almost completely eliminated SopB ubiquitination. We found that ubiquitination does not affect SopB stability or membrane association, or SopB-dependent events in SCV biogenesis. However, two spatially and temporally distinct events are dependent on ubiquitination, downregulation of SopB activity at the plasma membrane and prolonged retention of SopB on the SCV. Activation of the mammalian pro-survival kinase Akt/PKB, a downstream target of SopB, was intensified and prolonged after infection with the SopB-K6R mutant. At later times, fewer SCV were decorated with SopB-K6R compared with SopB. Instead SopB-K6R was present as discrete vesicles spread diffusely throughout the cell. Altogether, our data show that ubiquitination of SopB is not related to its intracellular stability but rather regulates its enzymatic activity at the plasma membrane and intracellular localization

Lysophosphatidic acid acyltransferase 3 regulates Golgi complex structure and function

Recent studies have suggested that the functional organization of the Golgi complex is dependent on phospholipid remodeling enzymes. Here, we report the identification of an integral membrane lysophosphatidic acid–specific acyltransferase, LPAAT3, which regulates Golgi membrane tubule formation, trafficking, and structure by altering phospholipids and lysophospholipids. Overexpression of LPAAT3 significantly inhibited the formation of Golgi membrane tubules in vivo and in vitro. Anterograde and retrograde protein trafficking was slower in cells overexpressing LPAAT3 and accelerated in cells with reduced expression (by siRNA). Golgi morphology was also dependent on LPAAT3 because its knockdown caused the Golgi to become fragmented. These data are the first to show a direct role for a specific phospholipid acyltransferase in regulating membrane trafficking and organelle structure

Activation of Akt by the Bacterial Inositol Phosphatase, SopB, is Wortmannin Insensitive

Salmonella enterica uses effector proteins translocated by a Type III Secretion System to invade epithelial cells. One of the invasion-associated effectors, SopB, is an inositol phosphatase that mediates sustained activation of the pro-survival kinase Akt in infected cells. Canonical activation of Akt involves membrane translocation and phosphorylation and is dependent on phosphatidyl inositide 3 kinase (PI3K). Here we have investigated these two distinct processes in Salmonella infected HeLa cells. Firstly, we found that SopB-dependent membrane translocation and phosphorylation of Akt are insensitive to the PI3K inhibitor wortmannin. Similarly, depletion of the PI3K regulatory subunits p85α and p85ß by RNAi had no inhibitory effect on SopB-dependent Akt phosphorylation. Nevertheless, SopB-dependent phosphorylation does depend on the Akt kinases, PDK1 and rictor-mTOR. Membrane translocation assays revealed a dependence on SopB for Akt recruitment to Salmonella ruffles and suggest that this is mediated by phosphoinositide (3,4) P2 rather than phosphoinositide (3,4,5) P3. Altogether these data demonstrate that Salmonella activates Akt via a wortmannin insensitive mechanism that is likely a class I PI3K-independent process that incorporates some essential elements of the canonical pathway

Gene Regulation and Transcriptomics

Borrelia (Borreliella) burgdorferi, along with closely related species, is the etiologic agent of Lyme disease. The spirochete subsists in an enzootic cycle that encompasses acquisition from a vertebrate host to a tick vector and transmission from a tick vector to a vertebrate host. To adapt to its environment and persist in each phase of its enzootic cycle, B. burgdorferi wields three systems to regulate the expression of genes: the RpoN-RpoS alternative sigma factor cascade, the Hk1/Rrp1 two-component system and its product c-di-GMP, and the stringent response mediated by RelBbu and DksA. These regulatory systems respond to enzootic phase-specific signals and are controlled or fine- tuned by transcription factors, including BosR and BadR, as well as small RNAs, including DsrABb and Bb6S RNA. In addition, several other DNA-binding and RNA-binding proteins have been identified, although their functions have not all been defined. Global changes in gene expression revealed by high-throughput transcriptomic studies have elucidated various regulons, albeit technical obstacles have mostly limited this experimental approach to cultivated spirochetes. Regardless, we know that the spirochete, which carries a relatively small genome, regulates the expression of a considerable number of genes required for the transitions between the tick vector and the vertebrate host as well as the adaptation to each

Salmonella – the ultimate insider. Salmonella virulence factors that modulate intracellular survival

Salmonella enterica serovar Typhimurium is a common facultative intracellular pathogen that causes food-borne gastroenteritis in millions of people worldwide. Intracellular survival and replication are important virulence determinants and the bacteria can be found in a variety of phagocytic and non-phagocytic cells in vivo. Invasion of host cells and intracellular survival are dependent on two type III secretion systems, T3SS1 and T3SS2, each of which translocates a distinct set of effector proteins. However, other virulence factors including ion transporters, superoxide dismutase, flagella and fimbriae are also involved in accessing and utilizing the intracellular niche