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Background: The past decade has seen the emergence of several molecular tools that render possible modification of cellular functions through accurate and easy addition, removal, or exchange of genomic DNA sequences. Among these technologies, transcription activator-like effectors (TALE) has turned out to be one of the most versatile and incredibly robust platform for generating targeted molecular tools as demonstrated by fusion to various domains such as transcription activator, repressor and nucleases., Results: In this study, we generated a novel nuclease architecture based on the transcription activator-like effector scaffold. In contrast to the existing Tail to Tail (TtT) and head to Head (HtH) nuclease architectures based on the symmetrical association of two TALE DNA binding domains fused to the C-terminal (TtT) or N-terminal (HtH) end of FokI, this novel architecture consists of the asymmetrical association of two different engineered TALE DNA binding domains fused to the N- and C-terminal ends of FokI (TALE::FokI and FokI::TALE scaffolds respectively). The characterization of this novel Tail to Head (TtH) architecture in yeast enabled us to demonstrate its nuclease activity and define its optimal target configuration. We further showed that this architecture was able to promote substantial level of targeted mutagenesis at three endogenous loci present in two different mammalian cell lines., Conclusion: Our results demonstrated that this novel functional TtH architecture which requires binding to only one DNA strand of a given endogenous locus has the potential to extend the targeting possibility of FokI-based TALE nucleases.

Diatoms, a major group of photosynthetic microalgae, have a high biotechnological potential that has not been fully exploited because of the paucity of available genetic tools. Here we demonstrate targeted and stable modifications of the genome of the marine diatom Phaeodactylum tricornutum, using both meganucleases and TALE nucleases. When nuclease-encoding constructs are co-transformed with a selectable marker, high frequencies of genome modifications are readily attained with 56 and 27% of the colonies exhibiting targeted mutagenesis or targeted gene insertion, respectively. The generation of an enhanced lipid-producing strain (45-fold increase in triacylglycerol accumulation) through the disruption of the UDP-glucose pyrophosphorylase gene exemplifies the power of genome engineering to harness diatoms for biofuel production.

The last few years have seen the increasing development of new DNA targeting molecular tools and strategies for precise genome editing. However, opportunities subsist to either improve or expand the current toolbox and further broaden the scope of possible biotechnological applications. Here we report the discovery and the characterization of BurrH, a new modular DNA binding protein from Burkholderia rhizoxinica that is composed of highly polymorphic DNA targeting modules. We also engineered this scaffold to create a new class of designer nucleases that can be used to efficiently induce in vivo targeted mutagenesis and targeted gene insertion at a desired locus.

Objectives: To evaluate the performance of diffusion-weighted imaging (DWI) against the reference standard of gadolinium-enhanced T1-weighted imaging (Gd-T1-WI) in children., Methods: Thirty-nine consecutive patients (mean age 5.7 years) with suspected acute pyelonephritis underwent magnetic resonance imaging (MRI) including DWI and (the reference standard) Gd-T1-WI. Each study was read in double-blinded fashion by two radiologists. Each kidney was graded as normal or abnormal. Sensitivity and specificity of DWI were computed. Agreement between sequences and interobserver reproducibility were calculated (Cohen κ statistic and the McNemar tests)., Results: Thirty-two kidneys (41 %) had hypo-enhancing areas on Gd-T1-W images. The sensitivity and specificity of DWI were 100 % (32/32) and 93.5 % (43/46). DWI demonstrated excellent agreement (κ = 0.92,) with Gd-T1-W, with no significant difference (P = 0.25) in detection of abnormal lesions. Interobserver reproducibility was excellent with DWI (κ = 0.79)., Conclusion: DWI enabled similar detection of abnormal areas to Gd-T1-WI and may provide an injection-free means of evaluation of acute pyelonephritis., Key Points: • Diffusion weighted magnetic resonance imaging (DWI) can confirm acute pyelonepritis. • DWI provided comparable results to gadolinium enhanced T1-W MRI in acute pyelonepritis. • Contrast medium injection could be avoided for diagnosing acute pyelonephritis by MRI. • MRI with T2-WI and DWI provide a fast and comprehensive diagnostic tool.

Introduction: The aim of this study was to evaluate the medium-term results and complications of open inguinal varicocelectomy, including vein ligation, intraoperative venography, and antegrade sclerotherapy., Materials and Methods: Sixty-four children were treated between 2000 and 2009 for idiopathic varicocele. Fifty children were examined 6 months after surgery. In 2010, 22 patients were recalled for testicular ultrasound scans (US) to evaluate the medium-term results of the technique., Results: The mean age of the children was 12.8 years at first consultation. Of the 50 cases, 35 children were asymptomatic, 13 experienced pain, 3 suffered from discomfort, and 1 had testicular asymmetry. Thirteen children had delayed left testicular growth compared with the right testis. The mean age at surgery was 13.3 years, and follow-up duration was 8.3 months ± 13.9. Thirty-eight patients achieved good results postsurgery; there was varicocele recurrence in 3, testicular hypotrophy in 7, and complete testicular atrophy in 2 patients., Conclusion: Naked eye inguinal surgical ligation does not appear to be safe enough to treat young adolescents, with the theoretical risk of a decrease in fertility in the future. In teams which are untrained in microsurgical or laparoscopic varicocelectomy, we suggest referring adolescent patients to a radiologist for embolization., (Georg Thieme Verlag KG Stuttgart · New York.)

Transcription activator-like effector nucleases are readily targetable 'molecular scissors' for genome engineering applications. These artificial nucleases offer high specificity coupled with simplicity in design that results from the ability to serially chain transcription activator-like effector repeat arrays to target individual DNA bases. However, these benefits come at the cost of an appreciably large multimeric protein complex, in which DNA cleavage is governed by the nonspecific FokI nuclease domain. Here we report a significant improvement to the standard transcription activator-like effector nuclease architecture by leveraging the partially specific I-TevI catalytic domain to create a new class of monomeric, DNA-cleaving enzymes. In vivo yeast, plant and mammalian cell assays demonstrate that the half-size, single-polypeptide compact transcription activator-like effector nucleases exhibit overall activity and specificity comparable to currently available designer nucleases. In addition, we harness the catalytic mechanism of I-TevI to generate novel compact transcription activator-like effector nuclease-based nicking enzymes that display a greater than 25-fold increase in relative targeted gene correction efficacy.

An 11-year-old Caucasian girl was investigated for a clitoromegaly that had increased in size over 5 weeks. Clitoromegaly is a rare condition in childhood. Among nonhormonal causes are tumours, both benign and malignant. Evaluation of the adrenal glands and ovaries was performed by US. An epidermoid cyst was suggested by MRI including diffusion-weighted imaging, and this was confirmed histopathologically.

We report a case of Herlyn-Werner-Wunderlich syndrome diagnosed in the neonatal period. US revealed the classic association of a uterus didelphys with blind hemivagina and no ipsilateral kidney. The diagnosis was established by postnatal US and confirmed by MRI. Differential diagnoses are discussed. A trans-hymeneal resection of the vaginal septum was performed at 1 month of age. Intra operative endoscopy revealed no left hemitrigone but showed an atretic orifice in the ipsilateral blind hemivagina, probably corresponding to the insertion of an ectopic ureter. Follow-up was unremarkable.

Engrailed transcription factors regulate the expression of guidance cues that pattern retinal axon terminals in the dorsal midbrain. They also act directly to guide axon growth in vitro. We show here that an extracellular En gradient exists in the tectum along the anterior-posterior axis. Neutralizing extracellular Engrailed in vivo with antibodies expressed in the tectum causes temporal axons to map aberrantly to the posterior tectum in chick and Xenopus. Furthermore, posterior membranes from wild-type tecta incubated with anti-Engrailed antibodies or posterior membranes from Engrailed-1 knockout mice exhibit diminished repulsive activity for temporal axons. Since EphrinAs play a major role in anterior-posterior mapping, we tested whether Engrailed cooperates with EphrinA5 in vitro. We find that Engrailed restores full repulsion to axons given subthreshold doses of EphrinA5. Collectively, our results indicate that extracellular Engrailed contributes to retinotectal mapping in vivo by modulating the sensitivity of growth cones to EphrinA.

Homeogenes encode homeoprotein transcription factors that have fundamental roles in development. They are key players in genetic networks that lay out the body plan and also determine morphology and physiology at the cellular and multicellular level. However, homeoproteins share activities that extend beyond transcription, including translation regulation and signalling. For example, homeoproteins participate in the definition of territories in the neuroepithelium and also have a function in axonal guidance. Based on these examples, we propose that homeoproteins are not only morphogenetic transcription factors, but also morphogens themselves.