Gene silencing in cancer by histone H3 lysine 27 trimethylation independent of promoter DNA methylation

Epigenetic silencing in cancer cells is mediated by at least two distinct histone modifications, polycomb-based histone H3 lysine 27 trimethylation (H3K27triM) and H3K9 dimethylation. The relationship between DNA hypermethylation and these histone modifications is not completely understood. Using chromatin immunoprecipitation microarrays (ChIP-chip) in prostate cancer cells compared to normal prostate, we found that up to 5% of promoters (16% CpG islands and 84% non-CpG islands) were enriched with H3K27triM. These genes were silenced specifically in prostate cancer, and those CpG islands affected showed low levels of DNA methylation. Downregulation of the EZH2 histone methyltransferase restored expression of the H3K27triM target genes alone or in synergy with histone deacetylase inhibition, without affecting promoter DNA methylation, and with no effect on the expression of genes silenced by DNA hypermethylation. These data establish EZH2-mediated H3K27triM as a mechanism of tumor-suppressor gene silencing in cancer that is potentially independent of promoter DNA methylation.     

Management Factors Influencing Location Selection Decisions of Independent Filmmakers: An Exploratory Case Study

This is an exploratory case study of a management issue that confronts filmmakers: how do they select locations for their films? The outcome of those decisions can have vast and immediate effects upon the profitability of a film. This is a sometimes shadowy, but significant industry with a major economic impact. For systems theory advocates, students, and explorers, this study demonstrates that investigative, immersive action research techniques can be used to explore what are otherwise hidden, somewhat cloistered business organizations and industry structures that are ordinarily closed to outsiders. The soft-system of this industry that is rooted upon trust, loyalty, alliances, relationships, and other non-financial bonds influence its decisions, and its institutional performance. While this research is focused upon a fragment of the overall film industry worldwide, and is thus limited in the universality of the propositions that subsequently emerge, this study can serve as a springboard for research into other soft systems, from which new knowledge of managerial perspectives can emerge.     

Cytonuclear genomic dissociation in African elephant species

African forest and savanna elephants are distinct species separated by a hybrid zone. Because hybridization can affect the systematic and conservation status of populations, we examined gene flow between forest and savanna elephants at 21 African locations. We detected cytonuclear dissociation, indicative of different evolutionary histories for nuclear and mitochondrial genomes. Both paternally (n = 205 males) and biparentally (n = 2,123 X-chromosome segments) inherited gene sequences indicated that there was deep genetic separation between forest and savanna elephants. Yet in some savanna locales distant from present-day forest habitats, many individuals with savanna-specific nuclear genotypes carried maternally transmitted forest elephant mitochondrial DNA. This extreme cytonuclear dissociation implies that there were ancient episodes of hybridization between forest females and savanna males, which are larger and reproductively dominant to forest or hybrid males. Recurrent backcrossing of female hybrids to savanna bulls replaced the forest nuclear genome. The persistence of residual forest elephant mitochondria in savanna elephant herds renders evolutionary interpretations based on mitochondrial DNA alone misleading and preserves a genomic record of ancient habitat changes.     

A new family of RhoGEFs activates the Rop molecular switch in plants

In plants, the small GTP-binding proteins called Rops work as signalling switches that control growth, development and plant responses to various environmental stimuli. Rop proteins (Rho of plants, Rac-like and AtRac in Arabidopsis thaliana) belong to the Rho family of Ras-related GTP-binding proteins that turn on signalling pathways by switching from a GDP-bound inactive to a GTP-bound active conformation. Activation depends on guanine nucleotide exchange factors (GEFs) that catalyse the otherwise slow GDP dissociation for subsequent GTP binding. Although numerous RhoGEFs exist in animals and yeasts, no Rop-specific GEFs have yet been identified in plants and so Rop activation has remained elusive. Here we describe a new family of RhoGEF proteins that are exclusive to plants. We define a unique domain within these RopGEFs, termed PRONE (plant-specific Rop nucleotide exchanger), which is exclusively active towards members of the Rop subfamily. It increases nucleotide dissociation from Rop more than a thousand-fold and forms a tight complex with nucleotide-free Rop. RopGEFs may represent the missing link in signal transduction from receptor kinases to Rops and their identification has implications for the evolution of the Rho molecular switch.     

Structure of the cyclic-AMP-responsive exchange factor Epac2 in its auto-inhibited state

Epac proteins (exchange proteins directly activated by cAMP) are guanine-nucleotide-exchange factors (GEFs) for the small GTP-binding proteins Rap1 and Rap2 that are directly regulated by the second messenger cyclic AMP and function in the control of diverse cellular processes, including cell adhesion and insulin secretion. Here we report the three-dimensional structure of full-length Epac2, a 110-kDa protein that contains an amino-terminal regulatory region with two cyclic-nucleotide-binding domains and a carboxy-terminal catalytic region. The structure was solved in the absence of cAMP and shows the auto-inhibited state of Epac. The regulatory region is positioned with respect to the catalytic region by a rigid, tripartite beta-sheet-like structure we refer to as the 'switchboard' and an ionic interaction we call the 'ionic latch'. As a consequence of this arrangement, the access of Rap to the catalytic site is sterically blocked. Mutational analysis suggests a model for cAMP-induced Epac activation with rigid body movement of the regulatory region, the features of which are universally conserved in cAMP-regulated proteins.