Karl Nightingale

King's College London | KCL

Supplementary lecture capture is widely used in higher education as the recordings generated are highly valued by students. Here we used a between-subjects, mixed methods study to evaluate whether this approach can support the learning of students disclosing Specific Learning Difficulties (SpLDs). We used a ‘Lecture – Independent study – Exam’ desi...

Background The KMT2A/MLL1 lysine methyltransferase complex is an epigenetic regulator of selected developmental genes, in part through the SET domain-catalysed methylation of H3K4. It is essential for normal embryonic development and haematopoiesis and frequently mutated in cancer. The catalytic properties and targeting of KMT2A/MLL1 depend on the...

Lecture capture, or 'podcasting' technology offers a new and engaging format of learning materials that can be used to increase the flexibility and interactivity of learning and teaching environments. Here we discuss different ways that these recordings can be incorporated into STEM discipline teaching, and the impact this can have on students' lea...

Epigenetics is one of the fastest moving fields in drug discovery, with almost every large pharmaceutical company and a substantial number of biotechnology companies targeting epigenetic processes to treat diseases ranging from cancer to Huntington’s disease and from inflammation to sickle cell anaemia. The book is structured in three main sections...

Background Histone deacetylase inhibitors (HDACi) cause histone hyperacetylation and H3K4 hypermethylation in various cell types. They find clinical application as anti-epileptics and chemotherapeutic agents, but the pathways through which they operate remain unclear. Surprisingly, changes in gene expression caused by HDACi are often limited in ext...

Lecture recordings are increasingly used to supplement lecture attendance within higher education, but their impact on student learning remains unclear. Here we describe a study to evaluate student use of lecture recordings and quantify their impact on academic performance. Questionnaire responses and online monitoring of student's access to record...

Histone deacetylase inhibitors (HDACi) are increasingly used as therapeutic agents, but the mechanisms by which they alter cell behaviour remain unclear. Here we use microarray expression analysis to show that only a small proportion of genes (∼9%) have altered transcript levels after treating HL60 cells with different HDACi (valproic acid, Trichos...

Primers used to assess changes in gene expression. Forward (F) and reverse (R) sequences are listed along with their melting temperatures (Tm). For some genes, more than one primer pair was used. (DOC)

Primers used for chromatin immunoprecipitation analysis. Forward (F) and reverse (R) primer sequences are listed along with melting temperatures (Tm). (DOC)

Histone acetylation plays an integral role in the epigenetic regulation of gene expression. Transcriptional activity reflects the recruitment of opposing classes of enzymes to promoter elements; histone acetyltransferases (EC 2.3.1.48) that deposit acetyl marks at a subset of histone residues and histone deacetylases that remove them. Many histone...

Histone deacetylase inhibitors (HDIs) are emerging as valuable new agents in the treatment of acute myeloid leukaemia (AML). However, since response rates to these agents alone are low, we sought to identify markers associated with responsiveness. In a trial of 20 patients treated with the HDI sodium valproate (VPA) in combination with all trans re...

Catalytic promiscuity: The human histone acetyltransferase P/CAF can confer propionyl instead of acetyl marks onto peptides that mimic the H3 histone tail. Catalysis of propionyl transfer occurs with a kcat of 8.8 min−1 (versus a kcat of 12 min−1 observed for the transfer of an acetyl group) and with identical specificity for individual lysine resi...

Histones are subject to a wide variety of post-translational modifications that play a central role in gene activation and silencing. We have used histone modification-specific antibodies to demonstrate that two histone modifications involved in gene activation, histone H3 acetylation and H3 lysine 4 methylation, are functionally linked. This inter...

Targeted chromatin remodelling is essential for many nuclear processes, including the regulation of V(D)J recombination. ATP-dependent nucleosome remodelling complexes are important players in this process whose activity must be tightly regulated. We show here that histone acetylation regulates nucleosome remodelling complex activity to boost RAG c...

The genetic code epitomises simplicity, near universality and absolute predictive power. By contrast, epigenetic information, in the form of histone modifications, is characterised by complexity, diversity and an overall tendency to respond to changes in genomic function rather than to predict them. Perhaps the transient changes in histone modifica...

ERK and p38 MAP kinases, acting through the downstream mitogen- and stress-activated kinase 1/2 (MSK1/2), elicit histone H3 phosphorylation on a subfraction of nucleosomes--including those at Fos and Jun--concomitant with gene induction. S10 and S28 on the H3 tail have both been shown to be phospho-acceptors in vivo. Both phospho-epitopes appear wi...

The rRNA gene cluster consists of multiple transcription units. Half of these are active, while the other half are transcriptionally inactive. Previously, in vivo studies have demonstrated that silencing of ribosomal DNA (rDNA) is mediated by the chromatin remodeling NoRC (nucleolar remodeling complex). To explore the mechanisms underlying NoRC-dir...

The ATPase ISWI is the catalytic core of several nucleosome remodeling complexes, which are able to alter histone–DNA interactions within nucleosomes such that the sliding of histone octamers on DNA is facilitated. Dynamic nucleosome repositioning may be involved in the assembly of chromatin with regularly spaced nucleosomes and accessible regulato...

The nuclear hormone receptor co-activator CARM1 has the potential to methylate histone H3 at arginine residues in vitro. The methyltransferase activity of CARM1 is necessary for its co-activator functions in transient transfection assays. However, the role of this methyltransferase in vivo is unclear, given that methylation of arginines is not easi...

The human beta-globin genes are regulated by the locus control region (LCR), an element composed of multiple DNase I-hypersensitive sites (HS sites) located 5' to the genes. Various functional studies indicate that the LCR confers high-level, position-independent, and copy number-dependent expression to linked globin genes in transgenic mice. Howev...

The ATPase ISWI can be considered the catalytic core of several multiprotein nucleosome remodeling machines. Alone or in the context of nucleosome remodeling factor, the chromatin accessibility complex (CHRAC), or ACF, ISWI catalyzes a number of ATP-dependent transitions of chromatin structure that are currently best explained by its ability to ind...

In contrast to its behavior as naked DNA, the MMTV promoter assembled in minichromosomes can be activated synergistically by the progesterone receptor and NF1 in a process involving ATP-dependent chromatin remodeling. The DNA-binding domain of NF1 is required and sufficient for stable occupancy of all receptor-binding sites and for functional syner...

We would like to thank all speakers at the workshop for help in preparing this report and apologize to those whose work we could not describe in full detail because of space constraints. We are grateful to the organizers of the workshop, Pierre Chambon, Toshio Fukasawa, and Roger Kornberg, as well as to Lars Thelander for careful reading of the man...

In this review we describe how the extract-mediated chromatin assembly system derived from preblastoderm Drosophila embryos can be modified to assemble chromatin from defined histones. This approach combines the advantages of assembling (i) chromatin templates from homogeneous histones with (ii) an assembly system that generates chromatin with phys...

A number of activators are known to increase transcription by RNA polymerase (pol) II through protein acetylation. While the physiological substrates for those acetylases are poorly defined, possible targets include general transcription factors, activator proteins and histones. Using a cell-free system to reconstitute chromatin with increased hist...

Nucleosomes and the chromatin structures they assemble provide the architectural foundation for the transcription process. Components of the eukaryotic transcriptional machinery share common structural features with the histones and high mobility group (HMG) proteins (Grosschedl et al. 1994; Wolffe and Pruss 1996a). Other transcription factors have...

We examined the structural and functional consequences of incorporating either histone H1, histone B4 or HMG1 into a synthetic dinucleosome containing two 5S rRNA genes. We found that all three proteins bind to linker DNA, stabilizing an additional 20 bp from micrococcal nuclease digestion and restrict nucleosome mobility. Histone H1 has the highes...

We have reconstituted nucleosomes containing the Xenopus borealis 5 S rRNA gene, a single histone octamer, and 1 or 2 molecules of histone H1. We determine that the 1st molecule of histone H1 to associate with the 5 S nucleosome binds with high affinity (K 2 nM), and the 2nd molecule of H1 binds with a reduced affinity (K 10 nM). This latter bindin...

The high mobility group proteins 1 and 2 (HMG1/2) and histone B4 are major components of chromatin within the nuclei assembled during the incubation of Xenopus sperm chromatin in Xenopus egg extract. To investigate their potential structural and functional roles, we have cloned and expressed Xenopus HMG1 and histone B4. Purified histone B4 and HMG1...

We examine the association of transcription factor TFIIIA with RNA-DNA heteroduplexes containing sequences from the Xenopus borealis 5 S rRNA gene. Under conditions where TFIIIA selectively binds to 5 S rRNA or the internal control region of the 5 S rRNA gene, no specific association of TFIIIA with DNA-RNA heteroduplexes containing either strand of...

We demonstrate that methylation of the 12 dinucleotide CpGs within a GC-rich DNA fragment containing a Xenopus borealis 5 S rRNA gene does not influence histone H1 binding to naked or nucleosomal 5 S DNA. Thus a simple mechanism in which histone H1 selectively associates with nucleosomes containing methylated CpG cannot explain the repressive effec...

We have studied the light-activated cleavage of DNA by cobalt-bleomycin using a series of synthetic DNA fragments containing (AT)n and (GC)n. This cleavage reaction requires high concentrations of the antibiotic and appears to be a stoichiometric process rather than a catalytic process. We find that, in common with the iron-complex, cobalt-bleomyci...

The antitumor agent bleomycin (Fig. 1) is a glycopeptide antibiotic which acts by degrading DNA in a reaction requiring the presence of Fe2+ and molecular oxygen. The antibiotic can be considered to contain two functional regions; the DNA binding domain formed by the bithiazole and terminal amine moieties, and the metal binding domain consisting of...

We have examined the cleavage of several synthetic DNA sequences by Iron(II)-bleomycin. We find that, although bleomycin cuts mixed sequence DNAs with a preference for GC = GT>GA> >GG, It efficiently cleaves regions of (AT)n cutting exclusively at ApT, not TpA. isolated ApT steps show very little cleavage while blocks of three or more contiguous AT...

The interaction of bleomycin with a kinetoplast DNA fragment has been examined using various footprinting techniques. This DNA adopts a bent structure and displays an unusually low gel mobility on account of its phased runs of adenines. The bleomycin-cobalt complex increases the mobility of this DNA fragment, in contrast with other DNAs which show...