Timeless noncoding DNA contains cell-type preferential enhancers important for proper Drosophila circadian regulation.

To address the contribution of transcriptional regulation to clock gene expression and to behavior, we generated a series of CRISPR-mediated deletions within two regions of the circadian gene (), an intronic E-box region and an upstream E-box region that are both recognized by the key transcription factor Clock (Clk) and its heterodimeric partner Cycle. The upstream deletions but not an intronic deletion dramatically impact expression in fly heads; the biggest upstream deletion reduces peak RNA levels and RNA cycling amplitude to about 15% of normal, and there are similar effects on protein (TIM). The cycling amplitude of other clock genes is also strongly reduced, in these cases due to increases in trough levels.

Widespread posttranscriptional regulation of cotransmission.

While neurotransmitter identity was once considered singular and immutable for mature neurons, it is now appreciated that one neuron can release multiple neuroactive substances (cotransmission) whose identities can even change over time. To explore the mechanisms that tune the suite of transmitters a neuron releases, we developed transcriptional and translational reporters for cholinergic, glutamatergic, and GABAergic signaling in . We show that many glutamatergic and GABAergic cells also transcribe cholinergic genes, but fail to accumulate cholinergic effector proteins.

Butt-seq: a new method for facile profiling of transcription.

A wide range of sequencing methods has been developed to assess nascent RNA transcription and resolve the single-nucleotide position of RNA polymerase genome-wide. These techniques are often burdened with high input material requirements and lengthy protocols. We leveraged the template-switching properties of thermostable group II intron reverse transcriptase (TGIRT) and developed Butt-seq (bulk analysis of nascent transcript termini sequencing), which can produce libraries from purified nascent RNA in 6 h and from as few as 10,000 cells-an improvement of at least 10-fold over existing techniques.

Widespread post-transcriptional regulation of co-transmission.

Unlabelled: While neurotransmitter identity was once considered singular and immutable for mature neurons, it is now appreciated that one neuron can release multiple neuroactive substances (co-transmission) whose identities can even change over time. To explore the mechanisms that tune the suite of transmitters a neuron releases, we developed transcriptional and translational reporters for cholinergic, glutamatergic, and GABAergic signaling in . We show that many glutamatergic and GABAergic cells also transcribe cholinergic genes, but fail to accumulate cholinergic effector proteins.

Loop-Mediated Isothermal Amplification Detection of SARS-CoV-2 and Myriad Other Applications.

As the second year of the COVID-19 pandemic begins, it remains clear that a massive increase in the ability to test for SARS-CoV-2 infections in a myriad of settings is critical to controlling the pandemic and to preparing for future outbreaks. The current gold standard for molecular diagnostics is the polymerase chain reaction (PCR), but the extraordinary and unmet demand for testing in a variety of environments means that both complementary and supplementary testing solutions are still needed. This review highlights the role that loop-mediated isothermal amplification (LAMP) has had in filling this global testing need, providing a faster and easier means of testing, and what it can do for future applications, pathogens, and the preparation for future outbreaks.

Development of a Saliva-Optimized RT-LAMP Assay for SARS-CoV-2.

Conventional reverse transcription quantitative polymerase chain reaction (RT-qPCR) technology has struggled to fulfill the unprecedented need for diagnostic testing created by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. Complexity and cost hinder access to testing, and long turnaround time decreases its utility. To ameliorate these issues, we focus on saliva and introduce several advances to colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) technology; RT-LAMP offers a minimal equipment alternative to RT-qPCR.

Long noncoding RNAs: a potent source of regulation in immunity and disease.

The discovery of functional long noncoding RNAs (lncRNAs) coupled with the ever-increasing accessibility of genomic and transcriptomic technology has led to an explosion of functional and mechanistic investigation and discovery into what was once dismissed as junk DNA. Over the past decade, a significant number of lncRNAs have been found to be involved in a diverse array of processes: from epigenetic modulation, both repressive and activating; to protein scaffolding; to miRNA sequestration; to competitive inhibition; and more. The broad character of these mechanisms means that lncRNAs have the potential for regulation across all biological processes-not least of which are immunity and disease.

Small molecule regulators of autophagy identified by an image-based high-throughput screen.

Autophagy is a lysosome-dependent cellular catabolic mechanism mediating the turnover of intracellular organelles and long-lived proteins. Reduction of autophagy activity has been shown to lead to the accumulation of misfolded proteins in neurons and may be involved in chronic neurodegenerative diseases such as Huntington's disease and Alzheimer's disease. To explore the mechanism of autophagy and identify small molecules that can activate it, we developed a series of high-throughput image-based screens for small-molecule regulators of autophagy.