Moving towards a cure in genetics : what is needed to bring somatic gene therapy to the clinic?

Clinical trials using somatic gene editing (e.g., CRISPR-Cas9) have started in Europe and the United States and may provide safe and effective treatment and cure, not only for cancers but also for some monogenic conditions. In a workshop at the 2018 European Human Genetics Conference, the challenges of bringing somatic gene editing therapies to the clinic were discussed. The regulatory process needs to be considered early in the clinical development pathway to produce the data necessary to support the approval by the European Medicines Agency. The roles and responsibilities for geneticists may include counselling to explain the treatment possibilities and safety interpretation.Peer reviewe

The Pseudomonas putida CsrA/RsmA homologues negatively affect c-di-GMP pools and biofilm formation through the GGDEF/EAL response regulator CfcR

Expression of cfcR, encoding the only GGDEF/EAL response regulator in Pseudomonas putida, is transcriptionally regulated by RpoS, ANR and FleQ, and the functionality of CfcR as a diguanylate cyclase requires the multisensor CHASE3/GAF hybrid histidine kinase named CfcA. Here an additional level of cfcR control, operating post-transcriptionally via the RNA-binding proteins RsmA, RsmE and RsmI, is unraveled. Specific binding of the three proteins to an Rsm binding motif (5’CANGGANG3’) encompassing the translational start codon of cfcR was confirmed. Although RsmA exhibited the highest binding affinity to the cfcR transcript, single deletions of rsmA, rsmE or rsmI, caused minor derepression in CfcR translation compared to a ∆rsmIEA triple mutant. RsmA also showed a negative impact on c-di-GMP levels in a double mutant ∆rsmIE through the control of cfcR, which is responsible for most of the free c-di-GMP during stationary phase in static conditions. In addition, a CfcR-dependent c-di-GMP boost was observed during this stage in ∆rsmIEA confirming the negative effect of Rsm proteins on CfcR translation and explaining the increased biofilm formation in this mutant compared to the wild type. Overall these results suggest that CfcR is a key player in biofilm formation regulation by the Rsm proteins in P. putida

Inhibiting N-acyl-homoserine lactone synthesis and quenching Pseudomonas quinolone quorum sensing to attenuate virulence

YesBacteria sense their own population size, tune the expression of responding genes, and behave accordingly to environmental stimuli by secreting signaling molecules. This phenomenon is termed as quorum sensing (QS). By exogenously manipulating the signal transduction bacterial population behaviors could be controlled, which may be done through quorum quenching (QQ). QS related regulatory networks have been proven their involvement in regulating many virulence determinants in pathogenic bacteria in the course of infections. Interfering with QS signaling system could be a novel strategy against bacterial infections and therefore requires more understanding of their fundamental mechanisms. Here we review the development of studies specifically on the inhibition of production of N-acyl-homoserine lactone (AHL), a common proteobacterial QS signal. The opportunistic pathogen, Pseudomonas aeruginosa, equips the alkylquinolone (AQ)-mediated QS which also plays crucial roles in its pathogenicity. The studies in QQ targeting on AQ are also discussed.University of Malaya High Impact Research Grants (UMC/625/1/HIR/MOHE/CHAN/01, A-000001-50001,and UMC/625/1/HIR/MOHE/CHAN/14/1, H-50001-A000027

Acidosis Potentiates the Host Proinflammatory Interleukin-1β Response to Pseudomonas Aeruginosa Infection

Infection by Pseudomonas aeruginosa, and bacteria in general, frequently promotes acidification of the local microenvironment, and this is reinforced by pulmonary exertion and exacerbation. However, the consequence of an acidic environment on the host inflammatory response to P. aeruginosa infection is poorly understood. Here we report that the pivotal cellular and host proinflammatory interleukin-1β (IL-1β) response, which enables host clearance of the infection but can produce collateral inflammatory damage, is increased in response to P. aeruginosa infection within an acidic environment. Synergistic mechanisms that promote increased IL-1β release in response to P. aeruginosa infection in an acidic environment are increased pro-IL-1β induction and increased caspase-1 activity, the latter being dependent upon a functional type III secretion system of the bacteria and the NLRC4 inflammasome of the host. Using an in vivo peritonitis model, we have validated that the IL-1β inflammatory response is increased in mice in response to P. aeruginosa infection within an acidic microenvironment. These data reveal novel insights into the regulation and exacerbation of inflammatory responses to P. aeruginosa

Surface sensing for biofilm formation in Pseudomonas aeruginosa

YesAggregating and forming biofilms on biotic or abiotic surfaces are ubiquitous bacterial behaviors under various conditions. In clinical settings, persistent presence of biofilms increases the risks of healthcare-associated infections and imposes huge healthcare and economic burdens. Bacteria within biofilms are protected from external damage and attacks from the host immune system and can exchange genomic information including antibiotic-resistance genes. Dispersed bacterial cells from attached biofilms on medical devices or host tissues may also serve as the origin of further infections. Understanding how bacteria develop biofilms is pertinent to tackle biofilm-associated infections and transmission. Biofilms have been suggested as a continuum of growth modes for adapting to different environments, initiating from bacterial cells sensing their attachment to a surface and then switching cellular physiological status for mature biofilm development. It is crucial to understand bacterial gene regulatory networks and decision-making processes for biofilm formation upon initial surface attachment. Pseudomonas aeruginosa is one of the model microorganisms for studying bacterial population behaviors. Several hypotheses and studies have suggested that extracellular macromolecules and appendages play important roles in bacterial responses to the surface attachment. Here, I review recent studies on potential molecular mechanisms and signal transduction pathways for P. aeruginosa surface sensing.This work is supported by University of Bradfor

RsmW, Pseudomonas aeruginosa small non-coding RsmA-binding RNA upregulated in biofilm versus planktonic growth conditions

BACKGROUND: Biofilm development, specifically the fundamentally adaptive switch from acute to chronic infection phenotypes, requires global regulators and small non-coding regulatory RNAs (sRNAs). This work utilized RNA-sequencing (RNA-seq) to detect sRNAs differentially expressed in Pseudomonas aeruginosa biofilm versus planktonic state. RESULTS: A computational algorithm was devised to detect and categorize sRNAs into 5 types: intergenic, intragenic, 5′-UTR, 3′-UTR, and antisense. Here we report a novel RsmY/RsmZ-type sRNA, termed RsmW, in P. aeruginosa up-transcribed in biofilm versus planktonic growth. RNA-Seq, 5’-RACE and Mfold predictions suggest RsmW has a secondary structure with 3 of 7 GGA motifs located on outer stem loops. Northern blot revealed two RsmW binding bands of 400 and 120 bases, suggesting RsmW is derived from the 3’-UTR of the upstream hypothetical gene, PA4570. RsmW expression is elevated in late stationary versus logarithmic growth phase in PB minimal media, at higher temperatures (37°C versus 28°C), and in both gacA and rhlR transposon mutants versus wild-type. RsmW specifically binds to RsmA protein in vitro and restores biofilm production and reduces swarming in an rsmY/rsmZ double mutant. PA4570 weakly resembles an RsmA/RsmN homolog having 49% and 51% similarity, and 16% and 17% identity to RsmA and RsmN amino acid sequences, respectively. PA4570 was unable to restore biofilm and swarming phenotypes in ΔrsmA deficient strains. CONCLUSION: Collectively, our study reveals an interesting theme regarding another sRNA regulator of the Rsm system and further unravels the complexities regulating adaptive responses for Pseudomonas species

Art in the Open: Milwaukee, Murals, and Community Engagement

A photo essay detailing mural culture in Milwaukee with an overview of its history, conversation with local muralists, and the benefits and consequences of muralism.https://epublications.marquette.edu/dslgallery/1006/thumbnail.jp

The feast of Lucy

In late summer of 1975, the body of a man is pulled from the Kinnickinnic River in Milwaukee. Among the neighbors that come to witness the scene is the Dobosz family, who must confront their own connections to the victim, his ties to an underground parish gambling ring, and what the discovery of his body means for them. As each member of the family turns to the others with suspicion, old and well-kept secrets threaten to come to light