Rare paediatric case of agenesis of the vermiform appendix, ileal duplication and sickle cell disease.

This study reports an exceptional case of a 14-year-old girl with sickle cell disease that was diagnosed with agenesis of the vermiform appendix and ileal duplication. Both consist of extremely rare gastrointestinal malformations whose association has never been described. The preadolescent girl presented with abdominal pain and vomiting, and the ultrasound was suggestive of acute appendicitis. Surgical findings were agenesis of the vermiform appendix and a T-shaped ileal malformation with inflammatory changes. The patient underwent resection and ileal end-to-end anastomosis. Histopathological evaluation identified an ileal duplication, with small bowel and colonic mucosa, no communication to the adjacent ileum and ischaemic changes. At 8-month follow-up, the patient was asymptomatic.

Variable ring-shaped lithography for the fabrication of meso- and microscale binary optical elements.

This paper presents concept, optical design, and the implementation of a novel, to the best of our knowledge, lithographic exposure tool for the fabrication of rotationally symmetric meso- and microscale optical structures using a variable ring-shaped light distribution. Compared to the conventional lithographic technique of direct writing in Cartesian coordinates, which is intrinsically suboptimal for the fabrication of rotationally symmetric optical structures, this approach allows for fast exposure and avoids disturbing stitching effects. The diameter of the exposure ring varies between 1.6 and 6.5 mm, and the ring width measures ∼75µm full width at half-maximum for all diameters. The basic capabilities of the exposure tool are demonstrated by the fabrication of exemplary meso- and microscale structures such as diffractive axicon elements, phase rings, Fresnel zone plates and zone plate arrays.

PRISMA: Protein Interaction Screen on Peptide Matrix Reveals Interaction Footprints and Modifications- Dependent Interactome of Intrinsically Disordered C/EBPß.

CCAAT enhancer-binding protein beta (C/EBPß) is a pioneer transcription factor that specifies cell differentiation. C/EBPß is intrinsically unstructured, a molecular feature common to many proteins involved in signal processing and epigenetics. The structure of C/EBPß differs depending on alternative translation initiation and multiple post-translational modifications (PTM). Mutation of distinct PTM sites in C/EBPß alters protein interactions and cell differentiation, suggesting that a C/EBPß PTM indexing code determines epigenetic outcomes. Herein, we systematically explored the interactome of C/EBPß using an array technique based on spot-synthesized C/EBPß-derived linear tiling peptides with and without PTM, combined with mass spectrometric proteomic analysis of protein interactions. We identified interaction footprints of ∼1,300 proteins in nuclear extracts, many with chromatin modifying, chromatin remodeling, and RNA processing functions. The results suggest that C/EBPß acts as a multi-tasking molecular switchboard, integrating signal-dependent modifications and structural plasticity to orchestrate interactions with numerous protein complexes directing cell fate and function.

Functional interaction of CCAAT/enhancer-binding-protein-α basic region mutants with E2F transcription factors and DNA.

The transcription factor CCAAT/enhancer-binding protein α (C/EBPα) regulates cell cycle arrest and terminal differentiation of neutrophils and adipocytes. Mutations in the basic leucine zipper domain (bZip) of C/EBPα are associated with acute myeloid leukemia. A widely used murine transforming C/EBPα basic region mutant (BRM2) entails two bZip point mutations (I294A/R297A). BRM2 has been discordantly described as defective for DNA binding or defective for interaction with E2F. We have separated the two BRM2 mutations to shed light on the intertwined reciprocity between C/EBPα-E2F-DNA interactions. Both, C/EBPα I294A and R297A retain transactivation capacity and interaction with E2F-DP. The C/EBPα R297A mutation destabilized DNA binding, whereas the C/EBPα I294A mutation enhanced binding to DNA. The C/EBPα R297A mutant, like BRM2, displayed enhanced interaction with E2F-DP but failed to repress E2F-dependent transactivation although both mutants were readily suppressed by E2F1 for transcription through C/EBP cis-regulatory sites. In contrast, the DNA binding enhanced C/EBPα I294A mutant displayed increased repression of E2F-DP mediated transactivation and resisted E2F-DP mediated repression. Thus, the efficient repression of E2F dependent S-phase genes and the activation of differentiation genes reside in the balanced DNA binding capacity of C/EBPα.

Presentation and Clinical Outcomes of Choledochal Cysts in Children and Adults: A Multi-institutional Analysis.

IMPORTANCE: Choledochal cysts (CCs) are rare, with risk of infection and cancer. OBJECTIVE: To characterize the natural history, management, and long-term implications of CC disease. DESIGN, SETTING, AND PARTICIPANTS: A total of 394 patients who underwent resection of a CC between January 1, 1972, and April 11, 2014, were identified from an international multi-institutional database. Patients were followed up through September 27, 2014. Clinicopathologic characteristics, operative details, and outcome data were analyzed from May 1, 2014, to October 14, 2014. INTERVENTION: Resection of CC. MAIN OUTCOMES AND MEASURES: Management, morbidity, and overall survival. RESULTS: Among 394 patients, there were 135 children (34.3%) and 318 women (80.7%). Adults were more likely to present with abdominal pain (71.8% vs 40.7%; P < .001) and children were more likely to have jaundice (31.9% vs 11.6%; P < .001). Preoperative interventions were more commonly performed in adults (64.5% vs 31.1%; P < .001), including endoscopic retrograde pancreatography (55.6% vs 27.4%; P < .001), percutaneous transhepatic cholangiography (17.4% vs 5.9%; P < .001), and endobiliary stenting (18.1% vs 4.4%; P < .001)). Type I CCs were more often seen in children vs adults (79.7% vs 64.9%; P = .003); type IV CCs predominated in the adult population (23.9% vs 12.0%; P = .006). Extrahepatic bile duct resection with hepaticoenterostomy was the most frequently performed procedure in both age groups (80.3%). Perioperative morbidity was higher in adults (35.1% vs 16.3%; P < .001). On pathologic examination, 10 patients (2.5%) had cholangiocarcinoma. After a median follow-up of 28 months, 5-year overall survival was 95.5%. On follow-up, 13 patients (3.3%), presented with biliary cancer. CONCLUSIONS AND RELEVANCE: Presentation of CC varied between children and adults, and resection was associated with a degree of morbidity. Although concomitant cancer was uncommon, it occurred in 3.0% of the patients. Long-term surveillance is indicated given the possibility of future development of biliary cancer after CC resection.

[Persistence of left superior vena cava associated to esophageal atresia: presentation of 5 cases]. / Persistência da veia cava superior esquerda associada a atrésia do esófago: revisão de 5 casos.

BACKGROUND: Esophageal atresia is known to be associated with a variety of additional congenital anomalies in multiple organ systems. Persistent left superior vena cava is one of the most common venous thoracic anomaly, occurring in about 0,3% of the population. The aim of this study was to characterize persistent left superior vena cava in infants treated in Hospital Dona Estefânia with esophageal atresia. METHODS: A retrospective review of all children treated for esophageal atresia from January 2002 to December 2013 was undertaken. Charts were reviewed for gestational age, weight, type of atresia, preoperative echocardiogram, associated anomalies, surgical approach, eventual postoperative echocardiogram and angioresonance for the study of congenital venous anomaly. RESULTS: Of 107 children, five had persistent left superior vena cava. Of the five cases, one had prenatal diagnosis. Further investigation showed duodenal atresia in one, urologic malformation, coloboma and bilateral ear deformities in other. All five patients were operated on through right thoracotomy and primary anastomosis was performed after ligation of the trachea-esophageal fistula and underwent angioresonance to characterize the vascular anomaly. No operative or post-operative complications were registered. CONCLUSIONS: Inspite of the preoperative workup, the anomaly was only identified in one of the patients. Generally, diagnosis of PLSVC is incidentally found during routine left-sided central venous catheterizations. It is essential to characterize the pattern of cardiac venous return that places those patients at a risk for paradoxical embolic complications to the arterial system.

Crosstalk between C/EBPbeta phosphorylation, arginine methylation, and SWI/SNF/Mediator implies an indexing transcription factor code.

Cellular signalling cascades regulate the activity of transcription factors that convert extracellular information into gene regulation. C/EBPbeta is a ras/MAPkinase signal-sensitive transcription factor that regulates genes involved in metabolism, proliferation, differentiation, immunity, senescence, and tumourigenesis. The protein arginine methyltransferase 4 PRMT4/CARM1 interacts with C/EBPbeta and dimethylates a conserved arginine residue (R3) in the C/EBPbeta N-terminal transactivation domain, as identified by mass spectrometry of cell-derived C/EBPbeta. Phosphorylation of the C/EBPbeta regulatory domain by ras/MAPkinase signalling abrogates the interaction between C/EBPbeta and PRMT4/CARM1. Differential proteomic screening, protein interaction studies, and mutational analysis revealed that methylation of R3 constraines interaction with SWI/SNF and Mediator complexes. Mutation of the R3 methylation site alters endogenous myeloid gene expression and adipogenic differentiation. Thus, phosphorylation of the transcription factor C/EBPbeta couples ras signalling to arginine methylation and regulates the interaction of C/EBPbeta with epigenetic gene regulatory protein complexes during cell differentiation.

G9a-mediated lysine methylation alters the function of CCAAT/enhancer-binding protein-beta.

The functional capacity of the transcriptional regulatory CCAAT/enhancer-binding protein-beta (C/EBPbeta) is governed by protein interactions and post-translational protein modifications. In a proteome-wide interaction screen, the histone-lysine N-methyltransferase, H3 lysine 9-specific 3 (G9a), was found to directly interact with the C/EBPbeta transactivation domain (TAD). Binding between G9a and C/EBPbeta was confirmed by glutathione S-transferase pulldown and co-immunoprecipitation. Metabolic labeling showed that C/EBPbeta is post-translationally modified by methylation in vivo. A conserved lysine residue in the C/EBPbeta TAD served as a substrate for G9a-mediated methylation. G9a, but not a methyltransferase-defective G9a mutant, abrogated the transactivation potential of wild type C/EBPbeta. A C/EBPbeta TAD mutant that contained a lysine-to-alanine exchange was resistant to G9a-mediated inhibition. Moreover, the same mutation conferred super-activation of a chromatin-embedded, endogenous C/EBPbeta target gene. Our data identify C/EBPbeta as a direct substrate of G9a-mediated post-translational modification that alters the functional properties of C/EBPbeta during gene regulation.

A protein interaction network links GIT1, an enhancer of huntingtin aggregation, to Huntington's disease.

Analysis of protein-protein interactions (PPIs) is a valuable approach for characterizing proteins of unknown function. Here, we have developed a strategy combining library and matrix yeast two-hybrid screens to generate a highly connected PPI network for Huntington's disease (HD). The network contains 186 PPIs among 35 bait and 51 prey proteins. It revealed 165 new potential interactions, 32 of which were confirmed by independent binding experiments. The network also permitted the functional annotation of 16 uncharacterized proteins and facilitated the discovery of GIT1, a G protein-coupled receptor kinase-interacting protein, which enhances huntingtin aggregation by recruitment of the protein into membranous vesicles. Coimmunoprecipitations and immunofluorescence studies revealed that GIT1 and huntingtin associate in mammalian cells under physiological conditions. Moreover, GIT1 localizes to neuronal inclusions, and is selectively cleaved in HD brains, indicating that its distribution and function is altered during disease pathogenesis.