Combined direct/indirect detection allows identification of DNA termini in diverse sequencing datasets and supports a multiple-initiation-site model for HIV plus-strand synthesis.

Replication of genetic material involves the creation of characteristic termini. Determining these termini is important to refine our understanding of the mechanisms involved in maintaining the genomes of cellular organisms and viruses. Here we describe a computational approach combining direct and indirect readouts to detect termini from next-generation short-read sequencing.

Transcription polymerase-catalyzed emergence of novel RNA replicons.

Transcription polymerases can exhibit an unusual mode of regenerating certain RNA templates from RNA, yielding systems that can replicate and evolve with RNA as the information carrier. Two classes of pathogenic RNAs (hepatitis delta virus in animals and viroids in plants) are copied by host transcription polymerases. Using in vitro RNA replication by the transcription polymerase of T7 bacteriophage as an experimental model, we identify hundreds of new replicating RNAs, define three mechanistic hallmarks of replication (subterminal de novo initiation, RNA shape-shifting, and interrupted rolling-circle synthesis), and describe emergence from DNA seeds as a mechanism for the origin of novel RNA replicons.

A Small RNA Isolation and Sequencing Protocol and Its Application to Assay CRISPR RNA Biogenesis in Bacteria.

Next generation high-throughput sequencing has enabled sensitive and unambiguous analysis of RNA populations in cells. Here, we describe a method for isolation and strand-specific sequencing of small RNA pools from bacteria that can be multiplexed to accommodate multiple biological samples in a single experiment. Small RNAs are isolated by polyacrylamide gel electrophoresis and treated with T4 polynucleotide kinase.

An Abundant Class of Non-coding DNA Can Prevent Stochastic Gene Silencing in the C. elegans Germline.

Cells benefit from silencing foreign genetic elements but must simultaneously avoid inactivating endogenous genes. Although chromatin modifications and RNAs contribute to maintenance of silenced states, the establishment of silenced regions will inevitably reflect underlying DNA sequence and/or structure. Here, we demonstrate that a pervasive non-coding DNA feature in Caenorhabditis elegans, characterized by 10-base pair periodic An/Tn-clusters (PATCs), can license transgenes for germline expression within repressive chromatin domains.

Translation readthrough mitigation.

A fraction of ribosomes engaged in translation will fail to terminate when reaching a stop codon, yielding nascent proteins inappropriately extended on their C termini. Although such extended proteins can interfere with normal cellular processes, known mechanisms of translational surveillance are insufficient to protect cells from potential dominant consequences. Here, through a combination of transgenics and CRISPR–Cas9 gene editing in Caenorhabditis elegans, we demonstrate a consistent ability of cells to block accumulation of C-terminal-extended proteins that result from failure to terminate at stop codons.

Suppression of insulin production and secretion by a decretin hormone.

Decretins, hormones induced by fasting that suppress insulin production and secretion, have been postulated from classical human metabolic studies. From genetic screens, we identified Drosophila Limostatin (Lst), a peptide hormone that suppresses insulin secretion. Lst is induced by nutrient restriction in gut-associated endocrine cells.

An Interdisciplinary Approach in Rehabilitation of an Adult Mutilated Dentition with Multiple Missing Permanent Teeth--A Case Report.

Adult orthodontics has been an area of increasing focus in recent times. Comprehensive treatment of adults is more challenging due to a myriad of problems that coexist in an adult that makes treatment planning more challenging. A large number of individuals seek dental care because of aesthetic reasons, that is, the desire to look more attractive by improving their smiles.

Interdisciplinary Approach for Management of Congenitally Missing Maxillary Lateral Incisors: A Case Report.

Maxillary lateral incisors are frequently found congenitally missing, and their replacement has to be done prosthodontically. However, there are a variety of treatment options; a justified solution after orthodontic correction is the use of Maryland Bridges. Following is a case report of congenitally missing maxillary lateral incisors and an impacted canine and their orthodontic correction followed by prosthetic replacement of the lateral incisors using Maryland bridges.

Direct assessment of hepatic mitochondrial oxidative and anaplerotic fluxes in humans using dynamic 13C magnetic resonance spectroscopy.

Despite the central role of the liver in the regulation of glucose and lipid metabolism, there are currently no methods to directly assess hepatic oxidative metabolism in humans in vivo. By using a new (13)C-labeling strategy in combination with (13)C magnetic resonance spectroscopy, we show that rates of mitochondrial oxidation and anaplerosis in human liver can be directly determined noninvasively. Using this approach, we found the mean rates of hepatic tricarboxylic acid (TCA) cycle flux (VTCA) and anaplerotic flux (VANA) to be 0.