KCL TEST: an open-source inspired asymptomatic SARS-CoV-2 surveillance programme in an academic institution.

Rapid and accessible testing was paramount in the management of the COVID-19 pandemic. Our university established KCL TEST: a SARS-CoV-2 asymptomatic testing programme that enabled sensitive and accessible PCR testing of SARS-CoV-2 RNA in saliva. Here, we describe our learnings and provide our blueprint for launching diagnostic laboratories, particularly in low-resource settings.

Non-coding RNAs associated with Prader-Willi syndrome regulate transcription of neurodevelopmental genes in human induced pluripotent stem cells.

Mutations and aberrant gene expression during cellular differentiation lead to neurodevelopmental disorders, such as Prader-Willi syndrome (PWS), which results from the deletion of an imprinted locus on paternally inherited chromosome 15. We analyzed chromatin-associated RNA in human induced pluripotent cells (iPSCs) upon depletion of hybrid small nucleolar long non-coding RNAs (sno-lncRNAs) and 5' snoRNA capped and polyadenylated long non-coding RNAs (SPA-lncRNAs) transcribed from the locus deleted in PWS. We found that rapid ablation of these lncRNAs affects transcription of specific gene classes.

Homebrew: Protocol for glassmilk-based nucleic-acid extraction for SARS-CoV-2 diagnostics.

The gold standard protocol for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection detection remains reverse transcription quantitative polymerase chain reaction (qRT-PCR), which detects viral RNA more sensitively than any other approach. Here, we present Homebrew, a low-cost protocol to extract RNA using widely available reagents. Homebrew is as sensitive as commercially available RNA extraction kits.

Resilient SARS-CoV-2 diagnostics workflows including viral heat inactivation.

There is a worldwide need for reagents to perform SARS-CoV-2 detection. Some laboratories have implemented kit-free protocols, but many others do not have the capacity to develop these and/or perform manual processing. We provide multiple workflows for SARS-CoV-2 nucleic acid detection in clinical samples by comparing several commercially available RNA extraction methods: QIAamp Viral RNA Mini Kit (QIAgen), RNAdvance Blood/Viral (Beckman) and Mag-Bind Viral DNA/RNA 96 Kit (Omega Bio-tek).

Resilient SARS-CoV-2 diagnostics workflows including viral heat inactivation.

There is a worldwide need for reagents to perform SARS-CoV-2 detection. Some laboratories have implemented kit-free protocols, but many others do not have the capacity to develop these and/or perform manual processing. We provide multiple workflows for SARS-CoV-2 nucleic acid detection in clinical samples by comparing several commercially available RNA extraction methods: QIAamp Viral RNA Mini Kit (QIAgen), RNAdvance Blood/Viral (Beckman) and Mag-Bind Viral DNA/RNA 96 Kit (Omega Bio-tek).

p38γ is essential for cell cycle progression and liver tumorigenesis.

The cell cycle is a tightly regulated process that is controlled by the conserved cyclin-dependent kinase (CDK)-cyclin protein complex. However, control of the G0-to-G1 transition is not completely understood. Here we demonstrate that p38 MAPK gamma (p38γ) acts as a CDK-like kinase and thus cooperates with CDKs, regulating entry into the cell cycle.

Nuclear fate of yeast snoRNA is determined by co-transcriptional Rnt1 cleavage.

Small nucleolar RNA (snoRNA) are conserved and essential non-coding RNA that are transcribed by RNA Polymerase II (Pol II). Two snoRNA classes, formerly distinguished by their structure and ribonucleoprotein composition, act as guide RNA to target RNA such as ribosomal RNA, and thereby introduce specific modifications. We have studied the 5'end processing of individually transcribed snoRNA in S.

Cell-Cycle Modulation of Transcription Termination Factor Sen1.

Many non-coding transcripts (ncRNA) generated by RNA polymerase II in S. cerevisiae are terminated by the Nrd1-Nab3-Sen1 complex. However, Sen1 helicase levels are surprisingly low compared with Nrd1 and Nab3, raising questions regarding how ncRNA can be terminated in an efficient and timely manner.

Disengaging polymerase: terminating RNA polymerase II transcription in budding yeast.

Termination of transcription by RNA polymerase II requires two distinct processes: The formation of a defined 3' end of the transcribed RNA, as well as the disengagement of RNA polymerase from its DNA template. Both processes are intimately connected and equally pivotal in the process of functional messenger RNA production. However, research in recent years has elaborated how both processes can additionally be employed to control gene expression in qualitative and quantitative ways.

Gene loops enhance transcriptional directionality.

Eukaryotic genomes are extensively transcribed, forming both messenger RNAs (mRNAs) and noncoding RNAs (ncRNAs). ncRNAs made by RNA polymerase II often initiate from bidirectional promoters (nucleosome-depleted chromatin) that synthesize mRNA and ncRNA in opposite directions. We demonstrate that, by adopting a gene-loop conformation, actively transcribed mRNA encoding genes restrict divergent transcription of ncRNAs.

Yeast Sen1 helicase protects the genome from transcription-associated instability.

Sen1 of S. cerevisiae is a known component of the NRD complex implicated in transcription termination of nonpolyadenylated as well as some polyadenylated RNA polymerase II transcripts. We now show that Sen1 helicase possesses a wider function by restricting the occurrence of RNA:DNA hybrids that may naturally form during transcription, when nascent RNA hybridizes to DNA prior to its packaging into RNA protein complexes.

Fail-safe transcriptional termination for protein-coding genes in S. cerevisiae.

Transcription termination of RNA polymerase II (Pol II) on protein-coding genes in S. cerevisiae relies on pA site recognition by 3' end processing factors. Here we demonstrate the existence of two alternative termination mechanisms that rescue polymerases failing to disengage from the template at pA sites.

Budding yeast RNA polymerases I and II employ parallel mechanisms of transcriptional termination.

Both RNA polymerase I and II (Pol I and Pol II) in budding yeast employ a functionally homologous "torpedo-like" mechanism to promote transcriptional termination. For two well-defined Pol II-transcribed genes, CYC1 and PMA1, we demonstrate that both Rat1p exonuclease and Sen1p helicase are required for efficient termination by promoting degradation of the nascent transcript associated with Pol II, following mRNA 3' end processing. Similarly, Pol I termination relies on prior Rnt1p cleavage at the 3' end of the pre-rRNA 35S transcript.

Small nuclear RNAs encoded by Herpesvirus saimiri upregulate the expression of genes linked to T cell activation in virally transformed T cells.

Seven small nuclear RNAs of the Sm class are encoded by Herpesvirus saimiri (HVS), a gamma Herpesvirus that causes aggressive T cell leukemias and lymphomas in New World primates and efficiently transforms T cells in vitro. The Herpesvirus saimiri U RNAs (HSURs) are the most abundant viral transcripts in HVS-transformed, latently infected T cells but are not required for viral replication or transformation in vitro. We have compared marmoset T cells transformed with wild-type or a mutant HVS lacking the most highly conserved HSURs, HSURs 1 and 2.

Actinomycin D induces histone gamma-H2AX foci and complex formation of gamma-H2AX with Ku70 and nuclear DNA helicase II.

Formation of gamma-H2AX foci is a P. O.cellular response to genotoxic stress, such as DNA double strand breaks or stalled replication forks.

The Herpesvirus saimiri small nuclear RNAs recruit AU-rich element-binding proteins but do not alter host AU-rich element-containing mRNA levels in virally transformed T cells.

Herpesvirus saimiri (HVS) encodes seven Sm-class small nuclear RNAs, called HSURs (for Herpesvirus saimiri U RNAs), that are abundantly expressed in HVS-transformed, latently infected marmoset T cells but are of unknown function. HSURs 1, 2, and 5 have highly conserved 5'-end sequences containing the AUUUA pentamer characteristic of AU-rich elements (AREs) that regulate the stability of many host mRNAs, including those encoding most proto-oncogenes and cytokines. To test whether the ARE-containing HSURs act to sequester host proteins that regulate the decay of these mRNAs, we demonstrate their in vivo interaction with the ARE-binding proteins hnRNP D and HuR in HVS-transformed T cells using a new cross-linking assay.