ABSTRACT
Protection of euchromatin from invasion by gene-repressive heterochromatin is critical for cellular health and viability. In addition to constitutive loci such as pericentromeres and subtelomeres, heterochromatin can be found interspersed in gene-rich euchromatin, where it regulates gene expression pertinent to cell fate. While heterochromatin and euchromatin are globally poised for mutual antagonism, the mechanisms underlying precise spatial encoding of heterochromatin containment within euchromatic sites remain opaque. We investigated ectopic heterochromatin invasion by manipulating the fission yeast mating type locus boundary using a single-cell spreading reporter system. We found that heterochromatin repulsion is locally encoded by Set1/COMPASS on certain actively transcribed genes and that this protective role is most prominent at heterochromatin islands, small domains interspersed in euchromatin that regulate cell fate specifiers. Sensitivity to invasion by heterochromatin, surprisingly, is not dependent on Set1 altering overall gene expression levels. Rather, the gene-protective effect is strictly dependent on Set1's catalytic activity. H3K4 methylation, the Set1 product, antagonizes spreading in two ways: directly inhibiting catalysis by Suv39/Clr4 and locally disrupting nucleosome stability. Taken together, these results describe a mechanism for spatial encoding of euchromatic signals that repel heterochromatin invasion.
Subject(s)
Cell Cycle Proteins/metabolism , Heterochromatin/metabolism , Histone-Lysine N-Methyltransferase/metabolism , Nucleosomes/metabolism , Schizosaccharomyces pombe Proteins/metabolism , Schizosaccharomyces/enzymology , Schizosaccharomyces/genetics , Transcription Factors/metabolism , Acetylation , Catalysis , Enzyme Activation , Gene Expression Regulation, Fungal , Gene Silencing , Histones/metabolismABSTRACT
Heterochromatin spreading, the expansion of repressive chromatin structure from sequence-specific nucleation sites, is critical for stable gene silencing. Spreading re-establishes gene-poor constitutive heterochromatin across cell cycles but can also invade gene-rich euchromatin de novo to steer cell fate decisions. How chromatin context (i.e. euchromatic, heterochromatic) or different nucleation pathways influence heterochromatin spreading remains poorly understood. Previously, we developed a single-cell sensor in fission yeast that can separately record heterochromatic gene silencing at nucleation sequences and distal sites. Here we couple our quantitative assay to a genetic screen to identify genes encoding nuclear factors linked to the regulation of heterochromatin nucleation and the distal spreading of gene silencing. We find that mechanisms underlying gene silencing distal to a nucleation site differ by chromatin context. For example, Clr6 histone deacetylase complexes containing the Fkh2 transcription factor are specifically required for heterochromatin spreading at constitutive sites. Fkh2 recruits Clr6 to nucleation-distal chromatin sites in such contexts. In addition, we find that a number of chromatin remodeling complexes antagonize nucleation-distal gene silencing. Our results separate the regulation of heterochromatic gene silencing at nucleation versus distal sites and show that it is controlled by context-dependent mechanisms. The results of our genetic analysis constitute a broad community resource that will support further analysis of the mechanisms underlying the spread of epigenetic silencing along chromatin.
Subject(s)
Schizosaccharomyces pombe Proteins , Schizosaccharomyces , Chromatin/genetics , Chromatin/metabolism , Gene Silencing , Heterochromatin/genetics , Heterochromatin/metabolism , Schizosaccharomyces/genetics , Schizosaccharomyces/metabolism , Schizosaccharomyces pombe Proteins/genetics , Schizosaccharomyces pombe Proteins/metabolism , Transcription Factors/genetics , Transcription Factors/metabolismABSTRACT
Heterochromatin spreading, the propagation of repressive chromatin along the chromosome, is a reaction critical to genome stability and defense, as well as maintenance of unique cell fates. Here, we discuss the intrinsic properties of the spreading reaction and circumstances under which its products, formed distal to DNA-encoded nucleation sites, can be epigenetically maintained. Finally, we speculate that the epigenetic properties of heterochromatin evolved together with the need to stabilize cellular identity.
Subject(s)
Epigenesis, Genetic , Gene Silencing , Genomic Instability , Heterochromatin/genetics , Animals , Chromatin Assembly and Disassembly , Fungi/genetics , Fungi/metabolism , Genomics/methods , Heterochromatin/metabolism , Humans , Inheritance PatternsABSTRACT
BACKGROUND: Thymic epithelial tumors (TETs) are rare neoplasms arising in the mediastinum, including thymic carcinomas and thymomas. Due to their rarity, little is known about the genomic profiles of TETs. Herein, we investigated the genomic characteristics of TETs evaluated in a large comprehensive genomic profiling database in a real-world setting. METHODS: We included data from two different cohorts: Foundation Medicine Inc. (FMI) in the United States and the Center for Cancer Genomics and Advanced Therapeutics (C-CAT) in Japan. Samples profiled were examined for all classes of alterations in 253 genes targeted across all assays. Tumor mutational burden (TMB) and microsatellite instability (MSI) were also evaluated. RESULTS: A total of 794 patients were collected in our study, including 722 cases from FMI and 72 cases from C-CAT. In the FMI data, CDKN2A (39.9%), TP53 (30.2%) and CDKN2B (24.6%) were frequently altered in thymic carcinoma, versus TP53 (7.8%), DNMT3A (6.8%), and CDKN2A (5.8%) in thymoma. TMB-high (≥10 mutations/Mb) and MSI were present in 7.0% and 2.3% of thymic carcinomas, and 1.6% and 0.3% of thymomas, respectively. Within C-CAT data, CDKN2A (38.5%), TP53 (36.5%) and CDKN2B (30.8%) were also frequently altered in thymic carcinoma, while alterations of TSC1, SETD2 and LTK (20.0% each) were found in thymoma. CONCLUSIONS: To the best of our knowledge, this is the largest cohort in which genomic alterations, TMB and MSI status of TETs were investigated. Potential targets for treatment previously unbeknownst in TETs are identified in this study, entailing newfound opportunities to advance therapeutic development.
Subject(s)
Neoplasms, Glandular and Epithelial , Thymoma , Thymus Neoplasms , Humans , Thymoma/genetics , Thymoma/pathology , Thymus Neoplasms/genetics , Thymus Neoplasms/pathology , Neoplasms, Glandular and Epithelial/genetics , GenomicsABSTRACT
Heterochromatin formation requires three distinct steps: nucleation, self-propagation (spreading) along the chromosome, and faithful maintenance after each replication cycle. Impeding any of those steps induces heterochromatin defects and improper gene expression. The essential histone chaperone FACT (facilitates chromatin transcription) has been implicated in heterochromatin silencing, but the mechanisms by which FACT engages in this process remain opaque. Here, we pinpoint its function to the heterochromatin spreading process in fission yeast. FACT impairment reduces nucleation-distal H3K9me3 and HP1/Swi6 accumulation at subtelomeres and derepresses genes in the vicinity of heterochromatin boundaries. FACT promotes spreading by repressing heterochromatic histone turnover, which is crucial for the H3K9me2 to me3 transition that enables spreading. FACT mutant spreading defects are suppressed by removal of the H3K9 methylation antagonist Epe1. Together, our study identifies FACT as a histone chaperone that promotes heterochromatin spreading and lends support to the model that regulated histone turnover controls the propagation of repressive methylation marks.
Subject(s)
Aminopeptidases/metabolism , Chromatin Assembly and Disassembly , Heterochromatin/metabolism , Histone Chaperones/metabolism , Histones/metabolism , Protein Processing, Post-Translational , Schizosaccharomyces pombe Proteins/metabolism , Schizosaccharomyces/metabolism , Aminopeptidases/genetics , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Gene Expression Regulation, Fungal , Gene Silencing , Heterochromatin/genetics , Histone Chaperones/genetics , Histone-Lysine N-Methyltransferase/genetics , Histone-Lysine N-Methyltransferase/metabolism , Methylation , Mutation , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Schizosaccharomyces/genetics , Schizosaccharomyces pombe Proteins/genetics , Transcription, GeneticABSTRACT
Gene expression is thought to be affected not only by the concentration of transcription factors (TFs) but also the dynamics of their nuclear translocation. Testing this hypothesis requires direct control of TF dynamics. Here, we engineer CLASP, an optogenetic tool for rapid and tunable translocation of a TF of interest. Using CLASP fused to Crz1, we observe that, for the same integrated concentration of nuclear TF over time, changing input dynamics changes target gene expression: pulsatile inputs yield higher expression than continuous inputs, or vice versa, depending on the target gene. Computational modeling reveals that a dose-response saturating at low TF input can yield higher gene expression for pulsatile versus continuous input, and that multi-state promoter activation can yield the opposite behavior. Our integrated tool development and modeling approach characterize promoter responses to Crz1 nuclear translocation dynamics, extracting quantitative features that may help explain the differential expression of target genes.
Subject(s)
DNA-Binding Proteins/metabolism , Saccharomyces cerevisiae Proteins/metabolism , Transcription Factors/metabolism , Active Transport, Cell Nucleus , Cell Nucleus/metabolism , DNA-Binding Proteins/genetics , Gene Expression , Optogenetics/methods , Promoter Regions, Genetic/genetics , Protein Transport , Saccharomyces cerevisiae , Saccharomyces cerevisiae Proteins/genetics , Transcription Factors/geneticsABSTRACT
The heterochromatin spreading reaction is a central contributor to the formation of gene-repressive structures, which are re-established with high positional precision, or fidelity, following replication. How the spreading reaction contributes to this fidelity is not clear. To resolve the origins of stable inheritance of repression, we probed the intrinsic character of spreading events in fission yeast using a system that quantitatively describes the spreading reaction in live single cells. We show that spreading triggered by noncoding RNA-nucleated elements is stochastic, multimodal, and fluctuates dynamically across time. This lack of stability correlates with high histone turnover. At the mating type locus, this unstable behavior is restrained by an accessory cis-acting element REIII, which represses histone turnover. Further, REIII safeguards epigenetic memory against environmental perturbations. Our results suggest that the most prevalent type of spreading, driven by noncoding RNA-nucleators, is epigenetically unstable and requires collaboration with accessory elements to achieve high fidelity.
Subject(s)
Epigenesis, Genetic , Heterochromatin/metabolism , RNA, Untranslated/genetics , Schizosaccharomyces/genetics , Cell Cycle Checkpoints/drug effects , Epigenesis, Genetic/drug effects , Genes, Mating Type, Fungal , Histones/metabolism , Hydroxamic Acids/pharmacology , Hydroxyurea/pharmacology , Inheritance Patterns/genetics , Mutation/genetics , Reproducibility of Results , Schizosaccharomyces/cytology , Schizosaccharomyces/drug effects , Single-Cell Analysis , Stochastic Processes , Stress, Physiological/drug effects , TemperatureABSTRACT
The narcotic antagonist naltrexone was studied in over 300 opiate addicts. Patient selection was a major factor in determining retention and treatment outcome. Treatment time ranged from 1 week to over a year (mean = 2 months). A quarter of the study patients had multiple treatment episodes. Stabilized patients had few side effects, except for occasional nausea and abdominal cramps. Almost half the subjects tested naltrexone by using opiates at least once; all reported satisfactory narcotic blockade. Very few subjects switched to nonopiates to get high, although several did increase their alcohol consumption during the first weeks of therapy. One-third of subjects contacted in a follow-up study were opiate-free 6 months after stopping naltrexone, indicating a successful short-term treatment modality.
Subject(s)
Naloxone/analogs & derivatives , Naltrexone/therapeutic use , Opioid-Related Disorders/rehabilitation , Adult , Alcohol Drinking , Ambulatory Care , Follow-Up Studies , Heroin Dependence/drug therapy , Heroin Dependence/rehabilitation , Hospitalization , Humans , Middle Aged , Opioid-Related Disorders/drug therapy , Patient Dropouts , Social Support , Substance Withdrawal Syndrome/prevention & controlABSTRACT
Over the past 20 years, methadone maintenance has been shown to be a safe, effective treatment for large numbers of heroin addicts. The majority of patients derive major benefits while in treatment, most measurably in the areas of decreased use of illicit opiates, diminished criminality, increased levels of employment and more stable interpersonal relationships. An advantage of methadone maintenance over other treatments is that it attracts and retains a relatively large segment of the addict population and is reasonably cost-effective. Naltrexone is well suited as a transitional treatment for individuals who have progressed using methadone maintenance. Patients completing a course of methadone maintenance should be encouraged to use naltrexone during the postmethadone period, when symptoms of protracted abstinence often lead them to reinitiate use of heroin. Those with stable family relationships, good jobs, minimal antisocial behavior, and low drug-craving before beginning a course of naltrexone appear to benefit most from the treatment. Rates of retention improve when naltrexone is used within a comprehensive rehabilitation program. Although addicted individuals are often stereotyped, they are, in fact, a heterogeneous group representing a range of psychopathologies and life situations. Thus, within any one facility, a variety of modalities should be available to allow treatment to be tailored to the individual. No single treatment is best for all patients, and, moreover, the preferred modality for any one individual may change over time as a result of progress in treatment or varying life circumstances. Multimodality programs that include methadone and naltrexone enable the maximal number of individuals to benefit from treatment.
Subject(s)
Heroin Dependence/rehabilitation , Methadone/therapeutic use , Naloxone/analogs & derivatives , Naltrexone/therapeutic use , Heroin Dependence/psychology , Heroin Dependence/therapy , Humans , Methadone/administration & dosage , Methadone/pharmacology , Naltrexone/pharmacology , Psychotherapy , Substance Withdrawal Syndrome/drug therapy , Time FactorsABSTRACT
In order to determine the type of patient most likely to benefit from opiate antagonist therapy, a series of multivariate regression analyses were performed on a sample of male veterans who completed induction on naltrexone. Patient background characteristics, demographic factors and during-treatment variables were used to predict outcome measured at 1-month follow-up. Employment at the start of naltrexone and length of naltrexone therapy were significantly related to better outcome at 1-month follow-up. Treatment duration was clearly the best outcome predictor. The finding that at least 30 days of naltrexone therapy was necessary for significant improvement at 1-month follow-up but that longer periods of treatment were not necessarily associated with greater gains suggests that treatment can be limited and still be relatively successful. A second set of analyses attempted to discover patient characteristics predictive of longer treatment duration. Results showed that patients who were employed and/or married at the start of naltrexone therapy were more likely to stay in treatment longer. Similar results have been obtained by others suggesting that these patients have the best family and social supports to sustain a positive treatment outcome. In addition, they may also be more 'motivated' since they have the most to lose personally and financially by readdiction.
Subject(s)
Naloxone/analogs & derivatives , Naltrexone/therapeutic use , Opioid-Related Disorders/rehabilitation , Employment , Follow-Up Studies , Humans , Male , Marriage , Patient Acceptance of Health Care , Regression Analysis , Social Support , Time FactorsABSTRACT
Many types of external trauma have been linked to the genesis of posttraumatic stress disorder (PTSD) and yet recent reports have focused almost exclusively on PTSD occurring in the Vietnam veteran (PTSD/veteran). The extent to which treatment experiences with PTSD/veteran can be generalized to other traumatized patients, for example, acute civilian populations, has not been investigated. Clinical observations comparing PTSD precipitated by a motor vehicle accident with PTSD/veteran suggested there were major differences between these two groups on the following variables: source of referral, age, sex, socioeconomic level, nature of stressor, timing of the stressor, character of the intrusive and avoidance symptoms, and treatment noncompliance behavior. These differences were of sufficient magnitude to call into question the feasibility, at this time, of constructing generalizations regarding PTSD utilizing only the PTSD/veteran population.
Subject(s)
Combat Disorders/psychology , Stress Disorders, Post-Traumatic/psychology , Veterans/psychology , Adaptation, Psychological , Adult , Combat Disorders/diagnosis , Humans , Male , Manuals as Topic , Referral and Consultation , Stress Disorders, Post-Traumatic/diagnosis , VietnamABSTRACT
Psychiatric disorders are best managed by interdisciplinary teams of psychiatrists, psychiatric nurses, social workers, and psychologists. Some Emergency Departments have access to such psychiatric services either in the hospital or in associated clinics or community mental health centers. Unfortunately, many are not staffed with mental health professionals or such expertise is available only on a limited basis, particularly on evenings and weekends. Therefore, one or more members of the Emergency Department staff should have specialized psychiatric training and experience. We have reviewed the format for conducting a psychiatric history and mental status examination and have discussed the most common emergency psychiatric disorders of the elderly and their management in the Emergency Department. In order to assess and treat elderly psychiatric patients properly, emergency personnel must be aware of the medical disorders associated with psychiatric illness and must be prepared to initiate treatment quickly and appropriately. For most patients, emergency intervention is the first step in ensuring that a correct diagnosis is made and that ongoing psychiatric treatment is arranged through timely consultation and referral.
Subject(s)
Emergency Services, Psychiatric , Mental Disorders/diagnosis , Aged , Diagnosis, Differential , Humans , Mental Disorders/drug therapy , Mental Disorders/therapy , Mental Status Schedule , Psychotropic Drugs/therapeutic useSubject(s)
Aged/psychology , Mental Disorders/therapy , Psychotherapy, Group/methods , Veterans/psychology , Family , Humans , Middle Aged , Morale , PennsylvaniaSubject(s)
Dyskinesia, Drug-Induced/nursing , Fluphenazine/therapeutic use , Adult , Aged , Antipsychotic Agents/adverse effects , Dyskinesia, Drug-Induced/diagnosis , Dyskinesia, Drug-Induced/drug therapy , Female , Fluphenazine/administration & dosage , Humans , Injections, Intramuscular , Male , Middle AgedSubject(s)
Dyskinesia, Drug-Induced/nursing , Outpatient Clinics, Hospital , Antipsychotic Agents/adverse effects , Dyskinesia, Drug-Induced/etiology , Hospitals, Veterans , Humans , Interprofessional Relations , Outpatient Clinics, Hospital/organization & administration , Patient Education as Topic , PennsylvaniaABSTRACT
A model for psychiatric consultation in the general hospital is described. The authors stress that extrapsychic and societal factors as well as intrapsychic processes influence patients' reactions to illness and hospitalization. An analogy between dreams and their manifest and latent content and the consultation request itself is made.