Laboratory Management of Mammalian Hosts for -Host-Pathogen Interaction Studies.

Due to their hematophagous life cycle, hard-bodied ticks including the genus are a potential vector for numerous pathogenic organisms including bacteria, protozoa, viruses, and infectious prions. The natural geographic range of several hard tick species, includig , has expanded over recent decades. Consequently, there is an ongoing need to maintain, feed, and propagate ticks for host-pathogen interaction studies to better understand and mitigate their impact on human and animal health.

Mechanisms and Functions of the RNA Polymerase II General Transcription Machinery during the Transcription Cycle.

Central to the development and survival of all organisms is the regulation of gene expression, which begins with the process of transcription catalyzed by RNA polymerases. During transcription of protein-coding genes, the general transcription factors (GTFs) work alongside RNA polymerase II (Pol II) to assemble the preinitiation complex at the transcription start site, open the promoter DNA, initiate synthesis of the nascent messenger RNA, transition to productive elongation, and ultimately terminate transcription. Through these different stages of transcription, Pol II is dynamically phosphorylated at the C-terminal tail of its largest subunit, serving as a control mechanism for Pol II elongation and a signaling/binding platform for co-transcriptional factors.

Single molecule studies characterize the kinetic mechanism of tetrameric p53 binding to different native response elements.

The transcriptional activator p53 is a tumor suppressor protein that controls cellular pathways important for cell fate decisions, including cell cycle arrest, senescence, and apoptosis. It functions as a tetramer by binding to specific DNA sequences known as response elements (REs) to control transcription via interactions with co-regulatory complexes. Despite its biological importance, the mechanism by which p53 binds REs remains unclear.

Short-term exposure to ethanol induces transcriptional changes in nontumorigenic breast cells.

Breast cancer is a leading cause of cancer-related deaths in women. Many genetic and behavioral risk factors can contribute to the initiation and progression of breast cancer, one being alcohol consumption. Numerous epidemiological studies have established a positive correlation between alcohol consumption and breast cancer; however, the molecular basis for this link remains ill defined.

Multiple RNA- and DNA-binding proteins exhibit direct transfer of polynucleotides with implications for target-site search.

We previously demonstrated that the polycomb repressive complex 2 chromatin-modifying enzyme can directly transfer between RNA and DNA without a free-enzyme intermediate state. Simulations suggested that such a direct transfer mechanism may be generally necessary for RNA to recruit proteins to chromatin, but the prevalence of direct transfer capability is unknown. Herein, we used fluorescence polarization assays and observed direct transfer for several well-characterized nucleic acid-binding proteins: three-prime repair exonuclease 1, heterogeneous nuclear ribonucleoprotein U, Fem-3-binding factor 2, and MS2 bacteriophage coat protein.

Differences in UV-C LED Inactivation of Serogroups in Drinking Water.

(Lp) is an opportunistic pathogen that causes respiratory infections primarily through inhalation of contaminated aerosols. Lp can colonize premise plumbing systems due to favorable growth conditions (e.g.

Interactions of HMGB Proteins with the Genome and the Impact on Disease.

High Mobility Group Box (HMGB) proteins are small architectural DNA binding proteins that regulate multiple genomic processes such as DNA damage repair, nucleosome sliding, telomere homeostasis, and transcription. In doing so they control both normal cellular functions and impact a myriad of disease states, including cancers and autoimmune diseases. HMGB proteins bind to DNA and nucleosomes to modulate the local chromatin environment, which facilitates the binding of regulatory protein factors to the genome and modulates higher order chromosomal organization.

The Herpes Simplex Virus 1 Protein ICP4 Acts as both an Activator and a Repressor of Host Genome Transcription during Infection.

Infection by herpes simplex virus 1 (HSV-1) impacts nearly all steps of host cell gene expression. The regulatory mechanisms by which this occurs, and the interplay between host and viral factors, have yet to be fully elucidated. We investigated how the occupancy of RNA polymerase II (Pol II) on the host genome changes during HSV-1 infection and is impacted by the viral immediate early protein ICP4.

Emergency response to stormwater contamination: A framework for containment and treatment.

This paper presents a Stormwater Emergency Response Framework (SERF) for use in the containment and treatment of stormwater runoff following a hazardous material release. The framework consists of four high level process steps and a decision tree. These resources are intended to assist stormwater managers in fulfilling their emergency response responsibilities within the United States' National Incident Management System.

Single molecule studies reveal that p53 tetramers dynamically bind response elements containing one or two half sites.

The tumor suppressor protein p53 is critical for cell fate decisions, including apoptosis, senescence, and cell cycle arrest. p53 is a tetrameric transcription factor that binds DNA response elements to regulate transcription of target genes. p53 response elements consist of two decameric half-sites, and data suggest one p53 dimer in the tetramer binds to each half-site.

Release of Human TFIIB from Actively Transcribing Complexes Is Triggered upon Synthesis of 7- and 9-nt RNAs.

RNA polymerase II (Pol II) and its general transcription factors assemble on the promoters of mRNA genes to form large macromolecular complexes that initiate transcription in a regulated manner. During early transcription, these complexes undergo dynamic rearrangement and disassembly as Pol II moves away from the start site of transcription and transitions into elongation. One step in disassembly is the release of the general transcription factor TFIIB, although the mechanism of release and its relationship to the activity of transcribing Pol II is not understood.

Periventricular White Matter Alterations From Explosive Blast in a Large Animal Model: Mild Traumatic Brain Injury or "Subconcussive" Injury?

The neuropathology of mild traumatic brain injury in humans resulting from exposure to explosive blast is poorly understood as this condition is rarely fatal. A large animal model may better reflect the injury patterns in humans. We investigated the effect of explosive blasts on the constrained head minimizing the effects of whole head motion.

miR-206 knockout shows it is critical for myogenesis and directly regulates newly identified target mRNAs.

The muscle specific miRNA, miR-206, is important for the process of myogenesis; however, studying the function of miR-206 in muscle development and differentiation still proves challenging because the complement of mRNA targets it regulates remains undefined. In addition, miR-206 shares close sequence similarity to miR-1, another muscle specific miRNA, making it hard to study the impact of miR-206 alone in cell culture models. Here we used CRISPR/Cas9 technology to knockout miR-206 in C2C12 muscle cells.

Dynamic Thermal Mapping of Localized Therapeutic Hypothermia in the Brain.

Although whole body cooling is used widely to provide therapeutic hypothermia for the brain, there are undesirable clinical side effects. Selective brain cooling may allow for rapid and controllable neuroprotection while mitigating these undesirable side effects. We evaluated an innovative cerebrospinal fluid (CSF) cooling platform that utilizes chilled saline pumped through surgically implanted intraventricular catheters to induce hypothermia.

Monitoring transcriptional activity by RNA polymerase II in vitro using single molecule co-localization.

RNA polymerase II (Pol II) transcribes eukaryotic mRNA genes. To initiate transcription, pre-initiation complexes (PICs) containing Pol II and general transcription factors (GTFs) form on the core promoters of target genes. In cells this process is regulated by transcriptional activators, co-activators, and chromatin modifying complexes.

Understanding the Impact of Mesh on Tank Overflow System Capacity.

A 2016 incident that resulted in damage to a water storage tank's roof motivated pilot-scale experiments to be conducted to determine the impact of mesh on tank overflow capacity. A clean mesh installed near the outlet of an overflow system did not reduce the capacity during the weir dominated flow regime. The impact of a mesh was found to be a reduction in the area available to flow, which was found to lower the achievable capacity through the system.

DETENTION OUTLET RETROFIT IMPROVES THE FUNCTIONALITY OF EXISTING DETENTION BASINS BY REDUCING EROSIVE FLOWS IN RECEIVING CHANNELS.

By discharging excess stormwater at rates that more frequently exceed the critical flow for stream erosion, conventional detention basins often contribute to increased channel instability in urban and suburban systems that can be detrimental to aquatic habitat and water quality, as well as adjacent property and infrastructure. However, these ubiquitous assets, valued at approximately $600,000 per km in a representative suburban watershed, are ideal candidates to aid in reversing such cycles of channel degradation because improving their functionality would not necessarily require property acquisition or heavy construction. The objective of this research was to develop a simple, cost-effective device that could be installed in detention basin outlets to reduce the erosive power of the relatively frequent storm events (~ < two-year recurrence) and provide a passive bypass to maintain flood control performance during infrequent storms (such as the 100-year recurrence).

Heat Shock Causes a Reversible Increase in RNA Polymerase II Occupancy Downstream of mRNA Genes, Consistent with a Global Loss in Transcriptional Termination.

Cellular transcriptional programs are tightly controlled but can profoundly change in response to environmental challenges or stress. Here we describe global changes in mammalian RNA polymerase II (Pol II) occupancy at mRNA genes in response to heat shock and after recovery from the stress. After a short heat shock, Pol II occupancy across thousands of genes decreased, consistent with widespread transcriptional repression, whereas Pol II occupancy increased at a small number of genes in a manner consistent with activation.

Reaction products of hexamethylene diisocyanate vapors with "self" molecules in the airways of rabbits exposed via tracheostomy.

1. Hexamethylenediisocyanate (HDI) is a widely used aliphatic diisocyanate and a well-recognized cause of occupational asthma. 2.

Finding the start site: redefining the human initiator element.

Transcription by RNA polymerase II (Pol II) is dictated in part by core promoter elements, which are DNA sequences flanking the transcription start site (TSS) that help direct the proper initiation of transcription. Taking advantage of recent advances in genome-wide sequencing approaches, Vo ngoc and colleagues (pp. 6-11) identified transcripts with focused sites of initiation and found that many were transcribed from promoters containing a new consensus sequence for the human initiator (Inr) core promoter element.

Neuronal and glial changes in the brain resulting from explosive blast in an experimental model.

Mild traumatic brain injury (mTBI) is the signature injury in warfighters exposed to explosive blasts. The pathology underlying mTBI is poorly understood, as this condition is rarely fatal and thus postmortem brains are difficult to obtain for neuropathological studies. Here we report on studies of an experimental model with a gyrencephalic brain that is exposed to single and multiple explosive blast pressure waves.

The HMGB1 C-Terminal Tail Regulates DNA Bending.

High mobility group box protein 1 (HMGB1) is an architectural protein that facilitates the formation of protein-DNA assemblies involved in transcription, recombination, DNA repair, and chromatin remodeling. Important to its function is the ability of HMGB1 to bend DNA non-sequence specifically. HMGB1 contains two HMG boxes that bind and bend DNA (the A box and the B box) and a C-terminal acidic tail.

Single molecule microscopy reveals mechanistic insight into RNA polymerase II preinitiation complex assembly and transcriptional activity.

Transcription by RNA polymerase II (Pol II) is a complex process that requires general transcription factors and Pol II to assemble on DNA into preinitiation complexes that can begin RNA synthesis upon binding of NTPs (nucleoside triphosphate). The pathways by which preinitiation complexes form, and how this impacts transcriptional activity are not completely clear. To address these issues, we developed a single molecule system using TIRF (total internal reflection fluorescence) microscopy and purified human transcription factors, which allows us to visualize transcriptional activity at individual template molecules.

Noncoding RNAs: Regulators of the Mammalian Transcription Machinery.

Transcription by RNA polymerase II (Pol II) is required to produce mRNAs and some noncoding RNAs (ncRNAs) within mammalian cells. This coordinated process is precisely regulated by multiple factors, including many recently discovered ncRNAs. In this perspective, we will discuss newly identified ncRNAs that facilitate DNA looping, regulate transcription factor binding, mediate promoter-proximal pausing of Pol II, and/or interact with Pol II to modulate transcription.