Abstract CTCF is an essential chromatin regulator implicated in important nuclear processes including in nuclear organization and transcription. Herpes Simplex Virus-1 (HSV-1) is a ubiquitous human pathogen, which enters productive infection in human epithelial and many other cell types. CTCF is known to bind several sites in the HSV-1 genome during latency and reactivation, but its function has not been defined. Here, we report that CTCF interacts extensively with the HSV-1 DNA during lytic infection by ChIP-seq, and its knockdown results in the reduction of viral transcription, viral genome copy number and virus yield. CTCF knockdown led to increased H3K9me3 and H3K27me3, and a reduction of RNA pol II occupancy on viral genes. Importantly, ChIP-seq analysis revealed that there is a higher level of CTD Ser2P modified RNA Pol II near CTCF peaks relative to the Ser5P form in the viral genome. Consistent with this, CTCF knockdown reduced the Ser2P but increased Ser5P modified forms of RNA Pol II on viral genes. These results suggest that CTCF promotes HSV-1 lytic transcription by facilitating the elongation of RNA Pol II and preventing silenced chromatin on the viral genome.
A hallmark of the research experience is encountering difficulty and working through those challenges to achieve success. This ability is essential to being a successful scientist, but replicating such challenges in a teaching setting can be difficult. The Genomics Education Partnership (GEP) is a consortium of faculty who engage their students in a genomics Course-Based Undergraduate Research Experience (CURE). Students participate in genome annotation, generating gene models using multiple lines of experimental evidence. Our observations suggested that the students’ learning experience is continuous and recursive, frequently beginning with frustration but eventually leading to success as they come up with defendable gene models. In order to explore our “formative frustration” hypothesis, we gathered data from faculty via a survey, and from students via both a general survey and a set of student focus groups. Upon analyzing these data, we found that all three datasets mentioned frustration and struggle, as well as learning and better understanding of the scientific process. Bioinformatics projects are particularly well suited to the process of iteration and refinement because iterations can be performed quickly and are inexpensive in both time and money. Based on these findings, we suggest that a dynamic of “formative frustration” is an important aspect for a successful CURE.
Abstract The Muller F element (4.2 Mb, ~80 protein-coding genes) is an unusual autosome of Drosophila melanogaster; it is mostly heterochromatic with a low recombination rate. To investigate how these properties impact the evolution of repeats and genes, we manually improved the sequence and annotated the genes on the D. erecta, D. mojavensis, and D. grimshawi F elements and euchromatic domains from the Muller D element. We find that F elements have greater transposon density (25–50%) than euchromatic reference regions (3–11%). Among the F elements, D. grimshawi has the lowest transposon density (particularly DINE-1: 2% vs. 11–27%). F element genes have larger coding spans, more coding exons, larger introns, and lower codon bias. Comparison of the Effective Number of Codons with the Codon Adaptation Index shows that, in contrast to the other species, codon bias in D. grimshawi F element genes can be attributed primarily to selection instead of mutational biases, suggesting that density and types of transposons affect the degree of local heterochromatin formation. F element genes have lower estimated DNA melting temperatures than D element genes, potentially facilitating transcription through heterochromatin. Most F element genes (~90%) have remained on that element, but the F element has smaller syntenic blocks than genome averages (3.4–3.6 vs. 8.4–8.8 genes per block), indicating greater rates of inversion despite lower rates of recombination. Overall, the F element has maintained characteristics that are distinct from other autosomes in the Drosophila lineage, illuminating the constraints imposed by a heterochromatic milieu.
There is widespread agreement that science, technology, engineering, and mathematics programs should provide undergraduates with research experience. Practical issues and limited resources, however, make this a challenge. We have developed a bioinformatics project that provides a course-based research experience for students at a diverse group of schools and offers the opportunity to tailor this experience to local curriculum and institution-specific student needs. We assessed both attitude and knowledge gains, looking for insights into how students respond given this wide range of curricular and institutional variables. While different approaches all appear to result in learning gains, we find that a significant investment of course time is required to enable students to show gains commensurate to a summer research experience. An alumni survey revealed that time spent on a research project is also a significant factor in the value former students assign to the experience one or more years later. We conclude: 1) implementation of a bioinformatics project within the biology curriculum provides a mechanism for successfully engaging large numbers of students in undergraduate research; 2) benefits to students are achievable at a wide variety of academic institutions; and 3) successful implementation of course-based research experiences requires significant investment of instructional time for students to gain full benefit.
Polycomb group (PcG) and trithorax group (trxG) proteins act in an epigenetic fashion to maintain active and repressive states of expression of the Hox and other target genes by altering their chromatin structure. Genetically, mutations in trxG and PcG genes can antagonize each other's function, whereas mutations of genes within each group have synergistic effects. Here, we show in Drosophila that multiple trxG and PcG proteins act through the same or juxtaposed sequences in the maintenance element (ME) of the homeotic gene Ultrabithorax. Surprisingly, trxG or PcG proteins, but not both, associate in vivo in any one cell in a salivary gland with the ME of an activated or repressed Ultrabithorax transgene, respectively. Among several trxG and PcG proteins, only Ash1 and Asx require Trithorax in order to bind to their target genes. Together, our data argue that at the single-cell level, association of repressors and activators correlates with gene silencing and activation, respectively. There is, however, no overall synergism or antagonism between and within the trxG and PcG proteins and, instead, only subsets of trxG proteins act synergistically.
