Unique Immune Cell Coactivators Specify Locus Control Region Function and Cell Stage.

Locus control region (LCR) functions define cellular identity and have critical roles in diseases such as cancer, although the hierarchy of structural components and associated factors that drive functionality are incompletely understood. Here we show that OCA-B, a B cell-specific coactivator essential for germinal center (GC) formation, forms a ternary complex with the lymphoid-enriched OCT2 and GC-specific MEF2B transcription factors and that this complex occupies and activates an LCR that regulates the BCL6 proto-oncogene and is uniquely required by normal and malignant GC B cells. Mechanistically, through OCA-B-MED1 interactions, this complex is required for Mediator association with the BCL6 promoter. Densely tiled CRISPRi screening indicates that only LCR segments heavily bound by this ternary complex are essential for its function. Our results demonstrate how an intimately linked complex of lineage- and stage-specific factors converges on specific and highly essential enhancer elements to drive the function of a cell-type-defining LCR.

Multi-tiered Reorganization of the Genome during B Cell Affinity Maturation Anchored by a Germinal Center-Specific Locus Control Region.

During the humoral immune response, B cells undergo a dramatic change in phenotype to enable antibody affinity maturation in germinal centers (GCs). Using genome-wide chromosomal conformation capture (Hi-C), we found that GC B cells undergo massive reorganization of the genomic architecture that encodes the GC B cell transcriptome. Coordinate expression of genes that specify the GC B cell phenotype-most prominently BCL6-was achieved through a multilayered chromatin reorganization process involving (1) increased promoter connectivity, (2) formation of enhancer networks, (3) 5' to 3' gene looping, and (4) merging of gene neighborhoods that share active epigenetic marks. BCL6 was an anchor point for the formation of GC-specific gene and enhancer loops on chromosome 3. Deletion of a GC-specific, highly interactive locus control region upstream of Bcl6 abrogated GC formation in mice. Thus, large-scale and multi-tiered genomic three-dimensional reorganization is required for coordinate expression of phenotype-driving gene sets that determine the unique characteristics of GC B cells.

Redox signaling by glutathione peroxidase 2 links vascular modulation to metabolic plasticity of breast cancer.

In search of redox mechanisms in breast cancer, we uncovered a striking role for glutathione peroxidase 2 (GPx2) in oncogenic signaling and patient survival. GPx2 loss stimulates malignant progression due to reactive oxygen species/hypoxia inducible factor-α (HIF1α)/VEGFA (vascular endothelial growth factor A) signaling, causing poor perfusion and hypoxia, which were reversed by GPx2 reexpression or HIF1α inhibition. Ingenuity Pathway Analysis revealed a link between GPx2 loss, tumor angiogenesis, metabolic modulation, and HIF1α signaling. Single-cell RNA analysis and bioenergetic profiling revealed that GPx2 loss stimulated the Warburg effect in most tumor cell subpopulations, except for one cluster, which was capable of oxidative phosphorylation and glycolysis, as confirmed by coexpression of phosphorylated-AMPK and GLUT1. These findings underscore a unique role for redox signaling by GPx2 dysregulation in breast cancer, underlying tumor heterogeneity, leading to metabolic plasticity and malignant progression.

Protective role of chaperone-mediated autophagy against atherosclerosis.

Chaperone-mediated autophagy (CMA) contributes to regulation of energy homeostasis by timely degradation of enzymes involved in glucose and lipid metabolism. Here, we report reduced CMA activity in vascular smooth muscle cells and macrophages in murine and human arteries in response to atherosclerotic challenges. We show that in vivo genetic blockage of CMA worsens atherosclerotic pathology through both systemic and cell-autonomous changes in vascular smooth muscle cells and macrophages, the two main cell types involved in atherogenesis. CMA deficiency promotes dedifferentiation of vascular smooth muscle cells and a proinflammatory state in macrophages. Conversely, a genetic mouse model with up-regulated CMA shows lower vulnerability to proatherosclerotic challenges. We propose that CMA could be an attractive therapeutic target against cardiovascular diseases.

Epidural stimulation restores muscle synergies by modulating neural drives in participants with sensorimotor complete spinal cord injuries.

Multiple studies have corroborated the restoration of volitional motor control after motor-complete spinal cord injury (SCI) through the use of epidural spinal cord stimulation (eSCS), but rigorous quantitative descriptions of muscle coordination have been lacking. Six participants with chronic, motor and sensory complete SCI underwent a brain motor control assessment (BMCA) consisting of a set of structured motor tasks with and without eSCS. We investigated how muscle activity complexity and muscle synergies changed with and without stimulation. We performed this analysis to better characterize the impact of stimulation on neuromuscular control. We also recorded data from nine healthy participants as controls. Competition exists between the task origin and neural origin hypotheses underlying muscle synergies. The ability to restore motor control with eSCS in participants with motor and sensory complete SCI allows us to test whether changes in muscle synergies reflect a neural basis in the same task. Muscle activity complexity was computed with Higuchi Fractal Dimensional (HFD) analysis, and muscle synergies were estimated using non-negative matrix factorization (NNMF) in six participants with American Spinal Injury Association (ASIA) Impairment Score (AIS) A. We found that the complexity of muscle activity was immediately reduced by eSCS in the SCI participants. We also found that over the follow-up sessions, the muscle synergy structure of the SCI participants became more defined, and the number of synergies decreased over time, indicating improved coordination between muscle groups. Lastly, we found that the muscle synergies were restored with eSCS, supporting the neural hypothesis of muscle synergies. We conclude that eSCS restores muscle movements and muscle synergies that are distinct from those of healthy, able-bodied controls.

Loss of the RNA polymerase III repressor MAF1 confers obesity resistance.

MAF1 is a global repressor of RNA polymerase III transcription that regulates the expression of highly abundant noncoding RNAs in response to nutrient availability and cellular stress. Thus, MAF1 function is thought to be important for metabolic economy. Here we show that a whole-body knockout of Maf1 in mice confers resistance to diet-induced obesity and nonalcoholic fatty liver disease by reducing food intake and increasing metabolic inefficiency. Energy expenditure in Maf1(-/-) mice is increased by several mechanisms. Precursor tRNA synthesis was increased in multiple tissues without significant effects on mature tRNA levels, implying increased turnover in a futile tRNA cycle. Elevated futile cycling of hepatic lipids was also observed. Metabolite profiling of the liver and skeletal muscle revealed elevated levels of many amino acids and spermidine, which links the induction of autophagy in Maf1(-/-) mice with their extended life span. The increase in spermidine was accompanied by reduced levels of nicotinamide N-methyltransferase, which promotes polyamine synthesis, enables nicotinamide salvage to regenerate NAD(+), and is associated with obesity resistance. Consistent with this, NAD(+) levels were increased in muscle. The importance of MAF1 for metabolic economy reveals the potential for MAF1 modulators to protect against obesity and its harmful consequences.

Congenital Segmental Dilatation of Ascending Colon with Distal Microcolon: A Diagnostic Dilemma.

A 5-day-old male presented with bilious vomiting, a grossly distended abdomen, and passage of a small amount of stool. The anal opening was at a normal position. X-ray abdomen showed a large bowel loop with a single air-fluid level occupying more than half of the abdominal width. On laparotomy, the ascending colon was dilated to form a pouch-like structure, and the ileum and appendix were opening into it. Colon distal to pouch was present as microcolon. Histopathology of the dilated segment was suggestive of congenital segmental dilatation (CSD). In CSD, the distal bowel is of normal caliber. This is a rare case of CSD of ascending colon with distal microcolon.

Palladium-Catalyzed Direct C2-Biarylation of Indoles.

Biaryl and indole units are important structural motifs in several bioactive molecules and functional materials. We have accomplished straightforward access to C2-biarylated indole derivatives through palladium-catalyzed C-H activation strategy with a broad range of substrate scope in yields of 24 to 92%. Besides, the UV/visible absorption and fluorescence properties of the ensuing products were explored. The calculated higher dihedral angle and rotational barrier values for the selected C2-biarylated indoles show that these compounds may display atropisomerism at room temperature.

Improvement of Mineral Absorption and Nutritional Properties of Citrullus vulgaris Seeds Using Solid-State Fermentation.

Objective:Citrullus vulgaris seeds are the most concealed oil seeds consist of vital therapeutic potential. Apart from nutritional and therapeutic indices, these seeds contain numerous anti-nutritional components.Method: Hence, solid-state fermentation was used as an alternative biotechnological contrivance to enhance the nutritional and functional properties of seeds. Fungal strain Aspergillus awamori was used for solid-state fermentation and uncoated watermelon seeds were used as a substrate for 144h. The range of bulk density was obtained from 0.39 to 0.72 g/cm3, however, fermentation significantly decreased the bulk density of the seed flours. Based upon soluble nitrogen content (79.81%) and functional properties, fermented seed samples of 120h was selected for FTIR analysis and in-vitro mineral bioavailability through Caco-2 cells.Results: FTIR spectrum confirmed the presence of amide groups (1200-1100 cm-1) of the protein.Conclusion: Fermented seeds also unveiled significantly (p<0.05) higher iron (61.24%), zinc (62.36%), and calcium (61.89%) bioavailability and this significant increase in bioavailability confirmed significantly (p<0.05) higher cellular mineral uptake.

Base-promoted 1,6-conjugate addition of alkylazaarenes to para-quinone methides.

1,1,2-Triarylethanes embedded with an azaarene unit were prepared in a single step at ambient temperature via the sodium hexamethyldisilazide mediated 1,6-conjugate addition of unactivated alkylazaarenes on para-quinone methides (p-QMs). The extent of this methodology was investigated with a wide range of p-QMs and alkylazaarenes, and the respective products were obtained in moderate to excellent yields. The regioselective 1,6-conjugate addition of the trialkylazaarene precursor on para-quinone methide was also presented. Besides, we showcased the direct synthesis of the 1,1,2-triarylethane derivative from aldehyde via the in situ generation of para-quinone methide.

Bochdalek's hernia in an elderly male - A case report.

Diaphragmatic hernia is protrusion of the abdominal contents into the thoracic cavity. It is a congenital defect of the diaphragm. It is most commonly encountered in infancy but rarely in adults. Here, we would like to present a rare case of 75-year-old male with Bochdalek's hernia. The patient had presented for the first time in his life for his symptoms. This case report emphasises the rare presentation of Bochdalek's hernia in an elderly male. The patient was treated using minimal access surgery i.e., laparoscopic approach and hence had a better post-operative outcome.

Comparative Assessment of Facial Asymmetry in Malocclusion using Posteroanterior View.

AIM: The study was conducted to assess facial asymmetry in various dental malocclusions and to determine asymmetry in lower, mid, and upper face and jaws using posteroanterior cephalometric analysis. MATERIALS AND METHODS: Overall, 120 posteroanterior cepha-lograms were taken of individuals between 12 and 25 years of both sexes, and were divided into four groups: Angle's class I excellent occlusion, Angle's class I malocclusion, Angle's class II malocclusion, and Angle's class III malocclusion. These cepha-lograms were traced and Grummon's analysis was performed. RESULTS: In Angle's class I occlusion and Angle's class II malocclusion, the results obtained showed asymmetry present in the upper face. Correlation was found between occlusion, maloc-clusion, and facial asymmetry. CONCLUSION: Facial asymmetry was found in all dental occlusions whether excellent or malocclusion group, with maximum asymmetry having upward trend toward upper face starting from lower. CLINICAL SIGNIFICANCE: For the success of the orthodontic treatments in various types of malocclusion and to determine the facial asymmetry, posteroanterior view can be very helpful, as it gives additional information which can be utilized to augment treatment planning and improve prognosis in terms of relapse prevention.

Autophagy, lipophagy and lysosomal lipid storage disorders.

Autophagy is a catabolic process with an essential function in the maintenance of cellular and tissue homeostasis. It is primarily recognised for its role in the degradation of dysfunctional proteins and unwanted organelles, however in recent years the range of autophagy substrates has also been extended to lipids. Degradation of lipids via autophagy is termed lipophagy. The ability of autophagy to contribute to the maintenance of lipo-homeostasis becomes particularly relevant in the context of genetic lysosomal storage disorders where perturbations of autophagic flux have been suggested to contribute to the disease aetiology. Here we review recent discoveries of the molecular mechanisms mediating lipid turnover by the autophagy pathways. We further focus on the relevance of autophagy, and specifically lipophagy, to the disease mechanisms. Moreover, autophagy is also discussed as a potential therapeutic target in several key lysosomal storage disorders.

Autophagy and Lipid Droplets in the Liver.

Autophagy is a conserved quality-control pathway that degrades cytoplasmic contents in lysosomes. Autophagy degrades lipid droplets through a process termed lipophagy. Starvation and an acute lipid stimulus increase autophagic sequestration of lipid droplets and their degradation in lysosomes. Accordingly, liver-specific deletion of the autophagy gene Atg7 increases hepatic fat content, mimicking the human condition termed nonalcoholic fatty liver disease. In this review, we provide insights into the molecular regulation of lipophagy, discuss fundamental questions related to the mechanisms by which autophagosomes recognize lipid droplets and how ATG proteins regulate membrane curvature for lipid droplet sequestration, and comment on the possibility of cross talk between lipophagy and cytosolic lipases in lipid mobilization. Finally, we discuss the contribution of lipophagy to the pathophysiology of human fatty liver disease. Understanding how lipophagy clears hepatocellular lipid droplets could provide new ways to prevent fatty liver disease, a major epidemic in developed nations.

Autophagy in Myf5+ progenitors regulates energy and glucose homeostasis through control of brown fat and skeletal muscle development.

Macroautophagy (MA) regulates cellular quality control and energy balance. For example, loss of MA in aP2-positive adipocytes converts white adipose tissue (WAT) into brown adipose tissue (BAT)-like, enhancing BAT function and thereby insulin sensitivity. However, whether MA regulates early BAT development is unknown. We report that deleting Atg7 in myogenic Myf5+ progenitors inhibits MA in Myf5-cell-derived BAT and muscle. Knock out (KO) mice have defective BAT differentiation and function. Surprisingly, their body temperature is higher due to WAT lipolysis-driven increases in fatty acid oxidation in 'Beige' cells in inguinal WAT, BAT and muscle. KO mice also present impaired muscle differentiation, reduced muscle mass and glucose intolerance. Our studies show that ATG7 in Myf5+ progenitors is required to maintain energy and glucose homeostasis through effects on BAT and muscle development. Decreased MA in myogenic progenitors with age and/or overnutrition might contribute to the metabolic defects and sarcopenia observed in these conditions.

Autophagy regulates lipid metabolism.

The intracellular storage and utilization of lipids are critical to maintain cellular energy homeostasis. During nutrient deprivation, cellular lipids stored as triglycerides in lipid droplets are hydrolysed into fatty acids for energy. A second cellular response to starvation is the induction of autophagy, which delivers intracellular proteins and organelles sequestered in double-membrane vesicles (autophagosomes) to lysosomes for degradation and use as an energy source. Lipolysis and autophagy share similarities in regulation and function but are not known to be interrelated. Here we show a previously unknown function for autophagy in regulating intracellular lipid stores (macrolipophagy). Lipid droplets and autophagic components associated during nutrient deprivation, and inhibition of autophagy in cultured hepatocytes and mouse liver increased triglyceride storage in lipid droplets. This study identifies a critical function for autophagy in lipid metabolism that could have important implications for human diseases with lipid over-accumulation such as those that comprise the metabolic syndrome.

Autophagy and aging.

Autophagy is a critical quality control pathway that is conserved across diverse systems ranging from simple unicellular organisms like yeast to more complex systems, for instance mammals. Although, the fundamental role of autophagy is to maintain cellular quality control through lysosomal degradation of unwanted proteins and organelles, recent studies have mapped several new functions of this pathway that range from fuel utilization, cellular differentiation to protection against cell death. Given the importance of this pathway in maintaining cellular homeostasis, it has been considered that compromised autophagy could contribute to several of the commonly observed age-associated pathologies including neurodegeneration, reduction of muscle mass, cardiac malfunction, excessive lipid accumulation in tissues and glucose intolerance. The present chapter describes the two best-characterized autophagy pathways­macroautophagy and chaperone-mediated autophagy, and discusses how changes in these pathways associate with age-associated disorders. Understanding how to maintain "clean cells" by activation of autophagy could be an attractive strategy to maintain healthspan in aged individuals.

Diphthamide modification on eukaryotic elongation factor 2 is needed to assure fidelity of mRNA translation and mouse development.

To study the role of the diphthamide modification on eukaryotic elongation factor 2 (eEF2), we generated an eEF2 Gly(717)Arg mutant mouse, in which the first step of diphthamide biosynthesis is prevented. Interestingly, the Gly(717)-to-Arg mutation partially compensates the eEF2 functional loss resulting from diphthamide deficiency, possibly because the added +1 charge compensates for the loss of the +1 charge on diphthamide. Therefore, in contrast to mouse embryonic fibroblasts (MEFs) from OVCA1(-/-) mice, eEF2(G717R/G717R) MEFs retain full activity in polypeptide elongation and have normal growth rates. Furthermore, eEF2(G717R/G717R) mice showed milder phenotypes than OVCA1(-/-) mice (which are 100% embryonic lethal) and a small fraction survived to adulthood without obvious abnormalities. Moreover, eEF2(G717R/G717R)/OVCA1(-/-) double mutant mice displayed the milder phenotypes of the eEF2(G717R/G717R) mice, suggesting that the embryonic lethality of OVCA1(-/-) mice is due to diphthamide deficiency. We confirmed that the diphthamide modification is essential for eEF2 to prevent -1 frameshifting during translation and show that the Gly(717)-to-Arg mutation cannot rescue this defect.