Invertebrate 7SK snRNAs

7SK RNA is a highly abundant noncoding RNA in mammalian cells whose function in transcriptional regulation has only recently been elucidated. Despite its highly conserved sequence throughout vertebrates, all attempts to discover 7SK RNA homologues in invertebrate species have failed so far. Here we report on a combined experimental and computational survey that succeeded in discovering 7SK RNAs in most of the major deuterostome clades and in two protostome phyla: mollusks and annelids. Despite major efforts, no candidates were found in any of the many available ecdysozoan genomes, however. The additional sequence data confirm the evolutionary conservation and hence functional importance of the previously described 3′ and 5′ stem-loop motifs, and provide evidence for a third, structurally well-conserved domain

Common Challenges and Identified Solutions for State Newborn Screening Programs during COVID-19 Pandemic

During the COVID-19 pandemic, state newborn screening programs faced challenges to ensure this essential public health program continued to function at a high level. In December 2020, the EveryLife Foundation for Rare Diseases held a workshop to discuss these common challenges and solutions. Newborn screening officials described challenges including short staffing across the entire program, collection and transport of specimens, interrupted follow-up activities, and pilot study recruitment. To address these challenges, state programs implemented a wide variety of solutions to maintain the high standards of newborn screening. To address staffing issues, newborn screening programs, public health laboratories, and hospitals all cross-trained personnel, worked to manage staff stress, and established essential functions. Other solutions included working with courier companies to ensure the timely pick-up of specimen, creating educational materials for hospital staff, and the creation of hybrid recruitment models for pilot studies. Implementing the lessons discussed throughout this paper can help to prepare for the next public health emergencies to ensure that a program that interacts with millions of families every year and saves the lives of thousands of children every year is minimally impacted

Er:YAG Laser for Metal and Ceramic Bracket Debonding: An In Vitro Study on Intrapulpal Temperature, SEM, and EDS Analysis.

OBJECTIVE To evaluate the effects of bracket removal using an erbium laser on the pulp temperature and enamel surface. BACKGROUND Removal of orthodontic brackets with conventional debonding pliers may result in enamel cracks. To avoid damage to the enamel surface and effectively remove metal or ceramic brackets, different types of lasers, such as Nd:YAG, CO, TM:YAP, diode laser, or Er:YAG, have been introduced for debonding. MATERIALS AND METHODS A total of 55 brackets (n = 55; 20 metal and 35 ceramic ones) were bonded to 55 caries-free premolars extracted for orthodontic indications. Brackets were irradiated with Er:YAG laser (Morita, Irvine, CA) with a wavelength of 2940 nm at a power of 3.4 W, energy 170 mJ, frequency 20 Hz, pulse duration 300 μs, tip diameter 0.8 mm, air/fluid cooling 3 mL/s, and time of irradiation: 6 sec. Debonding was made by scanning (n = 15; 6 sec irradiation at distance of 2 mm from the bracket with an "S" shape movement) and circular (n = 15; 6 sec irradiation at distance of 1 mm from the bracket) motion technique in ceramic brackets or the circular motion technique in metal brackets (n = 15). The number of 10 nonirradiated teeth with ceramic (n = 5) or metal brackets (n = 5) was used as a control in SEM test and EDS analysis. The damage in tooth enamel surface and the calcium percentage were analyzed by means of scanning electron microscope (JEOL 6610LV, JEOL, Japan) and energy dispersive X-ray spectroscopy (EDS, Oxford, United Kingdom). Temperature changes in the pulp were measured by K-type thermocouple. Evaluation of the Adhesive Remnant Index (ARI) on the enamel surface of each tooth was examined after bracket debonding. RESULTS The scanning method has caused significantly lower temperature increase (mean: 0.83°C) compared with circular motion technique around the ceramic brackets (mean: 1.78°C; p = 0.0001) or the metal brackets (mean: 1.29°C; p = 0.015). ARI score showed no differences between the study groups (p = 0.57). SEM analysis revealed no cracks on enamel surface after laser-assisted debonding in comparison with the control samples where cracks were found. EDS showed a higher mean percentage of the calcium (30.7-85.8%) for all test groups compared with control samples (mean: 7%; p = 0.0002). The amount of the calcium elements was higher for metal brackets in comparison with ceramic ones (p = 0.0002). CONCLUSIONS Er:YAG laser-assisted debonding causes a minor increase in the pulp temperature and reduced the risk of enamel damage compared with conventional bracket removal

Invertebrate 7SK snRNAs

7SK RNA is a highly abundant noncoding RNA in mammalian cells whose function in transcriptional regulation has only recently been elucidated. Despite its highly conserved sequence throughout vertebrates, all attempts to discover 7SK RNA homologues in invertebrate species have failed so far. Here we report on a combined experimental and computational survey that succeeded in discovering 7SK RNAs in most of the major deuterostome clades and in two protostome phyla: mollusks and annelids. Despite major efforts, no candidates were found in any of the many available ecdysozoan genomes, however. The additional sequence data confirm the evolutionary conservation and hence functional importance of the previously described 3´ and 5´ stemloop motifs, and provide evidence for a third, structurally well-conserved domain

Thienoquinolines as Novel Disruptors of the PKC epsilon/RACK2 Protein-Protein Interaction

Ten protein kinase C (PKC) isozymes play divergent roles in signal transduction. Because of sequence similarities, it is particularly difficult to generate isozyme-selective small molecule inhibitors. In order to identify such a selective binder, we derived a pharmacophore model from the peptide EAVSLKPT, a fragment of PKCε that inhibits the interaction of PKCε and receptor for activated C-kinase 2 (RACK2). A database of 330 000 molecules was screened in silico, leading to the discovery of a series of thienoquinolines that disrupt the interaction of PKCε with RACK2 in vitro. The most active molecule, N-(3-acetylphenyl)-9-amino-2,3-dihydro-1,4-dioxino[2,3-g]thieno[2,3-b]quinoline-8-carboxamide (8), inhibited this interaction with a measured IC50 of 5.9 μM and the phosphorylation of downstream target Elk-1 in HeLa cells with an IC50 of 11.2 μM. Compound 8 interfered with MARCKS phosphorylation and TPA-induced translocation of PKCε (but not that of PKCδ) from the cytosol to the membrane. The compound reduced the migration of HeLa cells into a gap, reduced invasion through a reconstituted basement membrane matrix, and inhibited angiogenesis in a chicken egg assay

Newborn Screening for Duchenne Muscular Dystrophy: First Year Results of a Population-Based Pilot

Advancements in therapies for Duchenne muscular dystrophy (DMD) have made diagnosis within the newborn period a high priority. We undertook a consortia approach to advance DMD newborn screening in the United States. This manuscript describes the formation of the Duchenne Newborn Screening Consortium, the development of the pilot protocols, data collection tools including parent surveys, and findings from the first year of a two-year pilot. The DMD pilot design is population-based recruitment of infants born in New York State. Data tools were developed to document the analytical and clinical validity of DMD NBS, capture parental attitudes, and collect longitudinal health information for diagnosed newborns. Data visualizations were updated monthly to inform the consortium on enrollment. After 12 months, 15,754 newborns were screened for DMD by the New York State Newborn Screening (NYS NBS) Program. One hundred and forty screened infants had borderline screening results, and sixteen infants were referred for molecular testing. Three male infants were diagnosed with dystrophinopathy. Data from the first year of a two-year NBS pilot for DMD demonstrate the feasibility of NBS for DMD. The consortia approach was found to be a useful model, and the Newborn Screening Translational Research Network’s data tools played a key role in describing the NBS pilot findings and engaging stakeholders

Thienoquinolines as Novel Disruptors of the PKCε/RACK2 Protein–Protein Interaction

Ten protein kinase C (PKC) isozymes play divergent roles in signal transduction. Because of sequence similarities, it is particularly difficult to generate isozyme-selective small molecule inhibitors. In order to identify such a selective binder, we derived a pharmacophore model from the peptide EAVSLKPT, a fragment of PKCε that inhibits the interaction of PKCε and receptor for activated C-kinase 2 (RACK2). A database of 330 000 molecules was screened in silico, leading to the discovery of a series of thienoquinolines that disrupt the interaction of PKCε with RACK2 in vitro. The most active molecule, <i>N</i>-(3-acetylphenyl)-9-amino-2,3-dihydro-1,4-dioxino­[2,3-<i>g</i>]­thieno­[2,3-<i>b</i>]­quinoline-8-carboxamide (<b>8</b>), inhibited this interaction with a measured IC₅₀ of 5.9 μM and the phosphorylation of downstream target Elk-1 in HeLa cells with an IC₅₀ of 11.2 μM. Compound <b>8</b> interfered with MARCKS phosphorylation and TPA-induced translocation of PKCε (but not that of PKCδ) from the cytosol to the membrane. The compound reduced the migration of HeLa cells into a gap, reduced invasion through a reconstituted basement membrane matrix, and inhibited angiogenesis in a chicken egg assay