An operon of three transcriptional regulators controls horizontal gene transfer of the integrative and conjugative element ICEclc in Pseudomonas knackmussii B13.

The integrative and conjugative element ICEclc is a mobile genetic element in Pseudomonas knackmussii B13, and an experimental model for a widely distributed group of elements in Proteobacteria. ICEclc is transferred from specialized transfer competent cells, which arise at a frequency of 3-5% in a population at stationary phase. Very little is known about the different factors that control the transfer frequency of this ICE family. Here we report the discovery of a three-gene operon encoded by ICEclc, which exerts global control on transfer initiation. The operon consists of three consecutive regulatory genes, encoding a TetR-type repressor MfsR, a MarR-type regulator and a LysR-type activator TciR. We show that MfsR autoregulates expression of the operon, whereas TciR is a global activator of ICEclc gene expression, but no clear role was yet found for MarR. Deletion of mfsR increases expression of tciR and marR, causing the proportion of transfer competent cells to reach almost 100% and transfer frequencies to approach 1 per donor. mfsR deletion also caused a two orders of magnitude loss in population viability, individual cell growth arrest and loss of ICEclc. This indicates that autoregulation is an important feature maintaining ICE transfer but avoiding fitness loss. Bioinformatic analysis showed that the mfsR-marR-tciR operon is unique for ICEclc and a few highly related ICE, whereas tciR orthologues occur more widely in a large variety of suspected ICE among Proteobacteria

Prospective multicentre study using high intensity focused ultrasound (HIFU) for the focal treatment of prostate cancer: Safety outcomes and complications

Purpose To investigate focal therapy using High Intensity Focused Ultrasound (HIFU) for the treatment of localized prostate cancer (CaP), we analyzed the safety and complications of this procedure. Methods Patients (pts) eligible for this multicenter prospective cohort study suffered from low to intermediate risk localized CaP with no prior treatment. After tumor identification on multiparametric MRI and in prostate biopsy, the lesions were treated with HIFU observing a safety margin of 8 to 10 mm. Adverse events (AE) after 30 and 90 days, as well as the required interventions were assessed and stratified for treatment localizations. Results Of the 98 men included in the study in two European centers, 35 (35.7%) experienced AEs in the first 30 days after HIFU intervention with Clavien-Dindo grade ≤ II: 15 pts (15.3%) had a postoperative urinary tract infection and 26 pts (26.5%) a urinary retention. Four pts (4.1%) underwent subsequent intervention (Clavien-Dindo grade IIIa/b). The number of late postoperative complications occurring between 30 and 90 days after intervention was low (2.0%). The highest complication rate was associated with tumors located at the anterior base (50.0%). The inclusion of the urethra in the ablation zone led to AEs in 20 out of 41 cases (48.8%) and represented a significant risk factor for complications within 30 days (odds ratio = 2.53; 95% confidence interval: 1.08–5.96; P = 0.033). Conclusions Focal therapy of CaP lesions with a robotic HIFU-probe is safe and renders an acceptable rate of minor early AEs. The inclusion of the urethra in the ablation zone leads to an increase in early complications and should be avoided whenever possible

Face valid phenotypes in a mouse model of the most common mutation in EEF1A2 related neurodevelopmental disorder, E122K

De novo heterozygous missense mutations in EEF1A2, encoding neuromuscular translation-elongation factor eEF1A2, are associated with developmental and epileptic encephalopathies. We used CRISPR/Cas9 to recapitulate the most common mutation, E122K, in mice. Although E122K heterozygotes were not observed to have convulsive seizures, they exhibited frequent electrographic seizures and EEG abnormalities, transient early motor deficits and growth defects. Both E122K homozygotes and Eef1a2-null mice developed progressive motor abnormalities, with E122K homozygotes reaching humane endpoints by P31. The null phenotype is driven by progressive spinal neurodegeneration; however, no signs of neurodegeneration were observed in E122K homozygotes. The E122K protein was relatively stable in neurons yet highly unstable in skeletal myocytes, suggesting that the E122K/E122K phenotype is instead driven by loss of function in muscle. Nevertheless, motor abnormalities emerged far earlier in E122K homozygotes than in nulls, suggesting a toxic gain of function and/or a possible dominant-negative effect. This mouse model represents the first animal model of an EEF1A2 missense mutation with face-valid phenotypes and has provided mechanistic insights needed to inform rational treatment design.</p

External Validation and Comparison of Prostate Cancer Risk Calculators Incorporating Multiparametric Magnetic Resonance Imaging for Prediction of Clinically Significant Prostate Cancer

PURPOSE: To externally validate recently published prostate cancer risk calculators (PCa-RCs) incorporating multiparametric magnetic resonance imaging (mpMRI) for the prediction of clinically significant prostate cancer (csPCa) and compare their performance to mpMRI-naïve PCa-RCs. MATERIAL AND METHODS: Men without previous PCa diagnosis undergoing transperineal template saturation prostate biopsy with fusion-guided targeted biopsy between 11/2014 and 03/2018 in our academic tertiary referral center were identified. Any Gleason pattern ≥4 was defined to be csPCa. Predictors (age, PSA, DRE, prostate volume, family history, previous prostate biopsy and highest region of interest according to PIRADS) were retrospectively collected. Four mpMRI-PCa-RCs and two mpMRI-naïve PCa-RCs were evaluated for their discrimination, calibration and clinical net benefit using a ROC analysis, calibration plots and a decision curve analysis, respectively. RESULTS: Out of 468 men, 193 (41%) were diagnosed with csPCa. Three mpMRI-PCa-RCs showed similar discrimination with area-underneath-the-receiver-operating-characteristic-curves (AUC) from 0.83 to 0.85, which was significantly higher than the other PCa-RCs (AUCs: 0.69-0.74). Calibration-in-the-large showed minimal deviation from the true amount of csPCa by 2% for two mpMRI-PCa-RCs, while the other PCa-RCs showed worse calibration (11-27%). A clinical net benefit could only be observed for three mpMRI-PCa-RCs at biopsy thresholds ≥15%, while none of the six investigated PCa-RCs demonstrated clinical utility against a biopsy all strategy at thresholds <15%. CONCLUSIONS: Performance of the mpMRI-PCa-RCs varies, but they generally outperform mpMRI-naïve PCa-RCs in regard to discrimination, calibration and clinical usefulness. External validation in other biopsy settings is highly encouraged

GABAergic Projections from the Medial Septum Selectively Inhibit Interneurons in the Medial Entorhinal Cortex

The medial septum (MS) is required for theta rhythmic oscillations and grid cell firing in the medial entorhinal cortex (MEC). While GABAergic, glutamatergic, and cholinergic neurons project from the MS to the MEC, their synaptic targets are unknown. To investigate whether MS neurons innervate specific layers and cell types in the MEC, we expressed channelrhodopsin-2 in mouse MS neurons and used patch-clamp recording in brain slices to determine the response to light activation of identified cells in the MEC. Following activation of MS axons, we observed fast monosynaptic GABAergic IPSPs in the majority (>60%) of fast-spiking (FS) and low-threshold-spiking (LTS) interneurons in all layers of the MEC, but in only 1.5% of nonstellate principal cells (NSPCs) and in no stellate cells. We also observed fast glutamatergic responses to MS activation in a minority (<5%) of NSPCs, FS, and LTS interneurons. During stimulation of MS inputs at theta frequency (10 Hz), the amplitude of GABAergic IPSPs was maintained, and spike output from LTS and FS interneurons was entrained at low (25–60 Hz) and high (60–180 Hz) gamma frequencies, respectively. By demonstrating cell type-specific targeting of the GABAergic projection from the MS to the MEC, our results support the idea that the MS controls theta frequency activity in the MEC through coordination of inhibitory circuits