Analysis of the Clonal Relationship of Serotype O26:H11 Enterohemorrhagic Escherichia coli Isolates from Cattle▿

Twelve cluster groups of Escherichia coli O26 isolates found in three cattle farms were monitored in space and time. Cluster analysis suggests that only some O26:H11 strains had the potential for long-term persistence in hosts and farms. As judged by their virulence markers, bovine enterohemorrhagic O26:H11 isolates may represent a considerable risk for human infection

Distinct Signal Transduction Pathways Downstream of the (P)RR Revealed by Microarray and ChIP-chip Analyses

The (pro)renin receptor ((P)RR) signaling is involved in different pathophysiologies ranging from cardiorenal end-organ damage via diabetic retinopathy to tumorigenesis. We have previously shown that the transcription factor promyelocytic leukemia zinc finger (PLZF) is an adaptor protein of the (P)RR. Furthermore, recent publications suggest that major functions of the (P)RR are mediated ligand-independently by its transmembrane and intracellular part, which acts as an accessory protein of V-ATPases. The transcriptome and recruitmentome downstream of the V-ATPase function and PLZF in the context of the (P)RR are currently unknown. Therefore, we performed a set of microarray and chromatin-immunoprecipitation (ChIP)-chip experiments using siRNA against the (P)RR, stable overexpression of PLZF, the PLZF translocation inhibitor genistein and the specific V-ATPase inhibitor bafilomycin to dissect transcriptional pathways downstream of the (P)RR. We were able to identify distinct and overlapping genetic signatures as well as novel real-time PCR-validated target genes of the different molecular functions of the (P)RR. Moreover, bioinformatic analyses of our data confirm the role of (P)R?s signal transduction pathways in cardiovascular disease and tumorigenesis

The (pro)renin receptor mediates constitutive PLZF-independent pro-proliferative effects which are inhibited by bafilomycin but not genistein

The (pro)renin receptor [(P)RR] is crucial for cardio-renal pathophysiology. The distinct molecular mechanisms of this receptor are still incompletely understood. The (P)RR is able to interact with different signalling proteins such as promyelocytic leukemia zinc finger protein (PLZF) and Wnt receptors. Moreover, domains of the (P)RR are essential for V-ATPase activity. V-ATPase- and Wnt-mediated effects imply constitutive, i.e., (pro)renin-independent functions of the (P)RR. Regarding ligand-dependent (P)RR signalling, the role of prorenin glycosylation is currently unknown. Therefore, the aim of this study was to analyse the contribution of constitutive (P)RR activity to its cellular effects and the relevance of prorenin glycosylation on its ligand activity. We were able to demonstrate that high glucose induces (P)RR signal transduction whereas deglycosylation of prorenin abolishes its intrinsic activity in neuronal and epithelial cells. By using siRNA against (P)RR or PLZF as well as the PLZF translocation blocker genistein and the specific V-ATPase inhibitor bafilomycin, we were able to dissect three distinct sub-pathways downstream of the (P)RR. The V-ATPase function is ligand-independently associated with strong pro-proliferative effects whereas prorenin causes moderate proliferation in vitro. In contrast, PLZF per se [i.e., in the absence of (pro)renin] does not interfere with cell number

The (pro)renin receptor ((P)RR) can act as a repressor of Wnt signalling

The (pro)renin receptor ((P)RR) and Wnt signalling are both involved in different diseases ranging from cardiac and renal end-organ damage to cancer. (P)RR function involves signalling via the transcription factor promyelocytic leukemia zinc finger protein (PLZF) as well as the furin-mediated generation of vacuolar proton-translocating ATPase (V-ATPase)-associated and soluble (P)RR isoforms. Recently, the (P)RR was described as adaptor protein of Wnt (co)receptors. The aim of this study was to analyse the contribution of these distinct (P)RR functions to Wnt signalling. Using Tcf/Lef reporter gene systems in HEK293T and HepG2 cells and quantification of endogenous axin2 mRNA and protein levels in HEK293T cells we were able to demonstrate that full-length (P)RR acts as a repressor of Wnt signalling in a system preactivated either by Wnt3a stimulation or by constitutively active ?-catenin. These repressive effects are mediated by Dvl but are independent of the mutation status of ?-catenin. Furthermore, the V-ATPase complex, but not PLZF translocation or renin enzymatic activity, is necessary for the induction of Tcf/Lef-responsive genes by Wnt3a. Our data indicate interference of (P)RR and Wnt cascades, a fact that has to be considered concerning pathophysiology of cardio-renal and oncological entities as well as in drug development programs targeting (P)RR or Wnt pathways

Moderate Correlations of in vitro versus in vivo Pharmacokinetics Questioning the Need of Early Microsomal Stability Testing

Putative in vitro-in vivo correlations of pharmacokinetic (PK) parameters are regarded as a prerequisite to filter hits derived from high-throughput screening (HTS) approaches for subsequent murine in vivo PK studies.In this study, we assessed stabilities in rat and human microsomes of 121 compounds from an early, academic drug discovery programme targeting the (pro)renin receptor and correlated the respective data with single-dose, in vivo PK parameters of 22 hits administered intravenously in rats.After transformation of in vitro half-lives to predicted in vivo hepatic clearances, r(2) regarding in vitro-in vivo clearance correlations were 0.31 and 0.27 for the rat and human species, respectively.Our data concerning structurally diverse real-world compounds indicate that microsomal stability testing is not a tool to triage early compounds for in vivo PK testing

Distinct Signal Transduction Pathways Downstream of the (P)RR Revealed by Microarray and ChIP-chip Analyses

<div><p>The (pro)renin receptor ((P)RR) signaling is involved in different pathophysiologies ranging from cardiorenal end-organ damage via diabetic retinopathy to tumorigenesis. We have previously shown that the transcription factor promyelocytic leukemia zinc finger (PLZF) is an adaptor protein of the (P)RR. Furthermore, recent publications suggest that major functions of the (P)RR are mediated ligand-independently by its transmembrane and intracellular part, which acts as an accessory protein of V-ATPases. The transcriptome and recruitmentome downstream of the V-ATPase function and PLZF in the context of the (P)RR are currently unknown. Therefore, we performed a set of microarray and chromatin-immunoprecipitation (ChIP)-chip experiments using siRNA against the (P)RR, stable overexpression of PLZF, the PLZF translocation inhibitor genistein and the specific V-ATPase inhibitor bafilomycin to dissect transcriptional pathways downstream of the (P)RR. We were able to identify distinct and overlapping genetic signatures as well as novel real-time PCR-validated target genes of the different molecular functions of the (P)RR. Moreover, bioinformatic analyses of our data confirm the role of (P)RŔs signal transduction pathways in cardiovascular disease and tumorigenesis.</p> </div

Experimental scheme of the ChIP-chips Cc1, Cc2 and Cc3 in combination with microarray Ma(PLZF; K).

<p>Pol II: RNA polymerase II binding sites in cells without intervention; pol IIⁱ: additional RNA polymerase II binding sites induced by PLZF overexpression.</p

Overview of microarray experiments.

<p>Overview of microarray experiments.</p