The rockefeller university

The X-end processing machinery for metazoan replication-dependent histone precursor messenger RNAs (pre-mRNAs) contains the U7 small nuclear ribonucleoprotein and shares the key cleavage module with the canonical cleavage and polyadenylation machinery. We reconstituted an active human histone pre-mRNA processing machinery using 13 recombinant proteins and two RNAs and determined its structure by cryo-electron microscopy. The overall structure is highly asymmetrical and resembles an amphora with one long handle. We captured the pre-mRNA in the active site of the endonuclease, the 73-kilodalton subunit of the cleavage and polyadenylation specificity factor, poised for cleavage. The endonuclease and the entire cleavage module undergo extensive rearrangements for activation, triggered through the recognition of the duplex between the authentic pre-mRNA and U7 small nuclear RNA (snRNA). Our study also has notable implications for understanding canonical and snRNA 3-end processing.

Intestinal barrier function is required for the maintenance of mucosal homeostasis. Barrier dysfunction is thought to promote progression of both intestinal and systemic diseases. In many cases, this barrier loss reflects increased permeability of the paracellular tight junction as a consequence of myosin light chain kinase (MLCK) activation and myosin II regulatory light chain (MLC) phosphorylation. Although some details about MLCK activation remain to be defined, it is clear that this triggers perijunctional actomyosin ring (PAMR) contraction that leads to molecular reorganization of tight junction structure and composition, including occludin endocytosis. In disease states, this process can be triggered by pro-inflammatory cytokines including tumor necrosis factor-alpha (TNF), interleukin-1 beta (IL-1 beta), and several related molecules. Of these, TNF has been studied in the greatest detail and is known to activate long MLCK transcription, expression, enzymatic activity, and recruitment to the PAMR. Unfortunately, toxicities associated with inhibition of MLCK expression or enzymatic activity make these unsuitable as therapeutic targets. Recent work has, however, identified a small molecule that prevents MLCK1 recruitment to the PAMR without inhibiting enzymatic function. This small molecule, termed Divertin, restores barrier function after TNF-induced barrier loss and prevents disease progression in experimental chronic inflammatory bowel disease.

Understanding the molecular mechanisms of endogenous and environmental metabolites is crucial for basic biology and drug discovery. With the genome, proteome, and metabolome of many organisms being readily available, researchers now have the opportunity to dissect how key metabolites regulate complex cellular pathways in vivo. Nonetheless, characterizing the specific and functional protein targets of key metabolites associated with specific cellular phenotypes remains a major challenge. Innovations in chemical biology are now poised to address this fundamental limitation in physiology and disease. In this review, we highlight recent advances in chemoproteomics for targeted and proteome-wide analysis of metabolite-protein interactions that have enabled the discovery of unpredicted metabolite-protein interactions and facilitated the development of new small molecule therapeutics.

The brain is a vulnerable metabolic organ and must adapt to different fuel conditions to sustain function. Nerve terminals are a locus of this vulnerability, but how they regulate ATP synthesis as fuel conditions vary is unknown. We show that synapses can switch from glycolytic to oxidative metabolism, but to do so, they rely on activity-driven presynaptic mitochondrial Ca2+ uptake to accelerate ATP production. We demonstrate that, whereas mitochondrial Ca2+ uptake requires elevated extramitochondrial Ca2+ in non-neuronal cells, axonal mitochondria readily take up Ca2+ in response to small changes in external Ca2+. We identified the brain-specific protein MICU3 as a critical driver of this tuning of Ca2+ sensitivity. Ablation of MICU3 renders axonal mitochondria similar to non-neuronal mitochondria, prevents acceleration of local ATP synthesis, and impairs presynaptic function under oxidative conditions. Thus, presynaptic mitochondria rely on MICU3 to facilitate mitochondrial Ca2+ uptake during activity and achieve metabolic flexibility.

Background: Use of local corticosteroids, especially the inhaled types, has increasingly been associated with systemic uptake and consequent adverse effects. In this study, we assessed the associations between the use of different corticosteroid types with cognitive and neuropsychiatric adverse effects related to high glucocorticoid exposure. Methods: In 83,592 adults (mean age 44 years, 59% women) of the general population (Lifelines Cohort Study), we analyzed the relationship between corticosteroid use with executive cognitive functioning (Ruff Figural Fluency Test), and presence of mood and anxiety disorders (Mini-International Neuropsychiatric Interview survey). We performed additional exploration for effects of physical quality of life (QoL; RAND-36), and inflammation (high-sensitive C-reactive protein [CRP]). Results: Cognitive scores were lower among corticosteroid users, in particular of systemic and inhaled types, when compared to nonusers. Users of inhaled types showed lower cognitive scores irrespective of physical QoL, psychiatric disorders, and high-sensitive CRP. Overall corticosteroid use was also associated with higher likelihood for mood and anxiety disorders. Users of inhaled corticosteroids were more likely to have mood disorders (OR 1.40 [95% CI 1.19-1.65], p < 0.001) and anxiety disorders (OR 1.19 [95% CI 1.06-1.33], p = 0.002). These findings were independent of physical QoL. A higher likelihood for mood disorders was also found for systemic users whereas nasal and dermal corticosteroid users were more likely to have anxiety disorders. Conclusions: Commonly used local corticosteroids, in particular inhaled types, and systemic corticosteroids are associated with reduced executive cognitive functioning and a higher likelihood of mood and anxiety disorders in the general adult population.

High-resolution structures have not been reported for replicative helicases at a replication fork at atomic resolution, a prerequisite to understanding the unwinding mechanism. The eukaryotic replicative CMG (Cdc45, Mcm2-7, GINS) helicase contains a Mcm2-7 motor ring, with the N-tier ring in front and the C-tier motor ring behind. The N-tier ring is structurally divided into a zinc finger (ZF) sub-ring followed by the oligosaccharide/oligonucleotide-binding (OB) fold ring. Here we report the cryo-EM structure of CMG on forked DNA at 3.9 angstrom, revealing that parental DNA enters the ZF sub-ring and strand separation occurs at the bottom of the ZF sub-ring, where the lagging strand is blocked and diverted sideways by OB hairpin-loops of Mcm3, Mcm4, Mcm6, and Mcm7. Thus, instead of employing a specific steric exclusion process, or even a separation pin, unwinding is achieved via a "dam-and-diversion tunnel" mechanism that does not require specific protein-DNA interaction. The C-tier motor ring contains spirally configured PS1 and H2I loops of Mcms 2, 3, 5, 6 that translocate on the spirally-configured leading strand, and thereby pull the preceding DNA segment through the diversion tunnel for strand separation. The DNA duplex is known to be split apart in a steric exclusion manner during replication, but the specific mechanism has remained unclear. Here the authors present a cryo-EM structure of a eukaryotic replicative CMG helicase on forked DNA, revealing the mechanism of DNA unwinding.

Adult stem cells are rare, undifferentiated cells found in all tissues of the body. Although normally kept in a quiescent, nondividing state, these cells can proliferate and differentiate to replace naturally dying cells within their tissue and to repair its wounds in response to injury. Due to their proliferative nature and ability to regenerate tissue, adult stem cells have the potential to treat a variety of degenerative diseases as well as aging. In addition, since stem cells are often thought to be the source of malignant tumors, understanding the mechanisms that keep their proliferative abilities in check can pave the way for new cancer therapies. While adult stem cells have had limited practical and clinical applications to date, several clinical trials of stem cell-based therapies are underway. This report details recent research presented at the New York Academy of Sciences on March 14, 2019 on understanding the factors that regulate stem cell activity and differentiation, with the hope of translating these findings into the clinic.

Purpose Caspase-associated recruitment domain-9 (CARD9) deficiency is an inborn error of immunity that typically predisposes otherwise healthy patients to single fungal infections and the occurrence of multiple invasive fungal infections is rare. It has been described as the first known condition that predisposes to extrapulmonary Aspergillus infection with preserved lungs. We present a patient that expands the clinical variability of CARD9 deficiency. Materials and methods Genetic analysis was performed by Sanger sequencing. Neutrophils and mononuclear phagocyte response to fungal stimulation were evaluated through luminol-enhanced chemiluminescence and whole blood production of the proinflammatory mediator interleukin (IL)-6, respectively. Results We report a 56-year-old Argentinean woman, whose invasive Exophiala spinifera infection at the age of 32 years was unexplained and reported in year 2004. At the age of 49 years, she presented with chronic pulmonary disease due to Aspergillus nomius. After partial improvement following treatment with caspofungin and posaconazole, right pulmonary bilobectomy was performed. Despite administration of multiple courses of antifungals, sustained clinical remission could not be achieved. We recently found that the patient's blood showed an impaired production of IL-6 when stimulated with zymosan. We also found that she is homozygous for a previously reported CARD9 loss-of-function mutation (Q289*). Conclusions This is the first report of a patient with inherited CARD9 deficiency and chronic invasive pulmonary aspergillosis (IPA) due to A. nomius. Inherited CARD9 deficiency should be considered in otherwise healthy children and adults with one or more invasive fungal diseases.

Vibrio-related gastroenteritis in the United States is mostly associated with the consumption of raw or improperly cooked seafood. We describe a case of a stage IV lung adenocarcinoma patient who became ill after eating crab while visiting Upstate New York. Molecular testing and culture confirmed a coinfection with V. parahaemolyticus and a nontoxigenic strain V. cholera. (C) 2019 Elsevier Inc. All rights reserved.