States Study Autumn 1974

Political attitudes before the state parliament elections 1974 in the states Schleswig-Holstein, Rhineland-Palatinate, North Rhine-Westphalia, Bavaria and Hesse. Judgement on parties and politicians. The questionnaire consists of a nationally uniform part and a state-specific part. Topics: 1. nationally uniform part: judgement on the economic situation and assumed economic development; extent to which politically informed and political interest; attitude to tax reform; spectrum of parties voted for by respondent in Federal Parliament elections since 1949; behavior at the polls in the Federal Parliament election 1972; personal change in party voted for; party preference and party ties; preferred federal chancellor; judgement on the government Schmidt/Genscher as well as the relationship of the FDP to the SPD; party inclination (split); changes of party sympathies in the course of time; most important political tasks in the FRG; preferred CDU/CSU politicians in leading positions; issue ability of the parties and selected politicians; sympathy scale for the SPD, CDU/CSU and the FDP; judgement on the Young Union as well as the Jusos {Young Socialists}; assessment of personal political location on the question of price stability and securing jobs as well as assumed attitudes of top politicians and parties; judgement on current foreign policy relations and their significance for the future; parties one cannot vote for; assessment of changes within a party since 1969; assumed identity of interests of the parties represented in the Federal Parliament; attitude to rejection of draft laws by the CDU/CSU majority in the Federal Upper House; judgement on Schmidt as federal chancellor (scale). 2a. State-specific part: judgement on the economic situation of the state; degree of familiarity of state politicians; knowledge about the party affiliation of selected politicians; preferred prime minister; profiles of characteristics of the prime minister candidates; most important state political goals; party preference at state level (ballot procedure); behavior at the polls in the last state parliament election; preferred government coalition; sympathy scale for the parties at state level. 2b. In Lower Saxony (as follow-up survey after the state parliament election had taken place): assessment of the economic situation and economic development; personal election participation and behavior at the polls in the last state parliament election; time of voting decision; satisfaction with the prime minister and the election result; election result corresponding to one´s own expectation; preferred government coalition; judgement on the election campaign; assessment of state or national political character of the state parliament election; expected influence of the change of federal chancellor on this election; knowledge about the counting error in the state parliament election; assumed reasons for this error; assumed party influence on this counting error; attitude to a new election and presumed change of personal behavior at the polls in this case; party preference (ballot procedure); sympathy changes regarding selected politicians and parties since the last state parliament election. 3. To all: knowledge of lay-offs in one´s own economic area; probability of loss of job next year regarding respondent as well as a member of the household; possession of a telephone. Demography: age; sex; marital status; religious denomination; frequency of church attendance; school education; occupational position; employment; household income; household composition; respondent is head of household; characteristics of head of household; self-assessment of social class; city size; state; union membership; Interviewer rating: length of interview; day of interview; interest of respondent in interview; number of contact attempts.Politische Einstellungen vor den Landtagswahlen 1974 in den Bundesländern Schleswig-Holstein, Rheinland-Pfalz, Nordrhein-Westfalen, Bayern und Hessen. Beurteilung von Parteien und Politikern. Der Fragebogen besteht aus einem bundeseinheitlichen Teil und einem länderspezifischen Teil. Themen: 1. Bundeseinheitlicher Teil: Beurteilung der wirtschaftlichen Lage und der vermuteten Wirtschaftsentwicklung; politische Informiertheit und politisches Interesse; Einstellung zur Steuerreform; Spektrum der vom Befragten seit 1949 bei Bundestagswahlen gewählten Parteien; Wahlverhalten bei der Bundestagswahl 1972; eigene Wechselwählerschaft; Parteipräferenz und Parteibindung; präferierter Bundeskanzler; Beurteilung der Regierung Schmidt/ Genscher sowie des Verhältnisses der FDP zur SPD; Parteineigung (Split); Änderungen der Parteisympathien im Zeitverlauf; wichtigste politische Aufgaben in der BRD; präferierte CDU/CSU-Politiker in Führungspositionen; Issue-Kompetenz der Parteien und ausgewählter Politiker; Sympathie-Skalometer für die SPD, CDU/CSU und die FDP; Beurteilung der Jungen Union sowie der Jusos; Einschätzung des eigenen politischen Standorts in der Frage von Preisstabilität und Arbeitsplatzsicherung sowie vermutete Einstellungen von Spitzenpolitiker und Parteien; Beurteilung der derzeitigen außenpolitischen Beziehungen und ihrer Bedeutung für die Zukunft; nicht wählbare Parteien; Einschätzung der parteiinternen Veränderungen seit 1969; vermutete Interessenidentität der im Bundestag vertretenen Parteien; Einstellung zur Ablehnung von Gesetzesvorlagen durch die CDU/CSU-Mehrheit im Bundesrat; Beurteilung von Schmidt als Bundeskanzler (Skala). 2a.) Länderspezifischer Teil: Beurteilung der wirtschaftlichen Lage des Bundeslandes; Bekanntheitsgrad von Landespolitikern; Kenntnis der Parteizugehörigkeit ausgewählter Politiker; präferierter Ministerpräsident; Eigenschaftsprofile der Ministerpräsidenten-Kan didaten; wichtigste landespolitische Ziele; Parteipräferenz auf Landesebene (Stimmzettelverfahren); Wahlverhalten bei der letzten Landtagswahl; präferierte Regierungskoalition; Sympathie-Skalometer für die Parteien auf Landesebene. 2b.) In Niedersachsen (als Nachbefragung zur stattgefundenen Landtagswahl): Einschätzung der wirtschaftlichen Lage und der Wirtschaftsentwicklung; eigene Wahlbeteiligung und Wahlverhalten bei der letzten Landtagswahl; Zeitpunkt der Wahlentscheidung; Zufriedenheit mit dem Ministerpräsidenten und dem Wahlausgang; Wahlausgang entsprechend der eigenen Erwartung; präferierte Regierungskoalition; Beurteilung des Wahlkampfes; Einschätzung des landes-oder bundespolitischen Charakters der Landtagswahl; vermuteter Einfluß des Bundeskanzlerwechsels auf diese Wahl; Kenntnis des Auszählungsfehlers bei der Landtagswahl; vermutete Gründe für diese Fehler; vermuteter Parteieneinfluß auf diesen Auszählfehler; Einstellung zu einer Neuwahl und vermutliche Änderung des eigenen Wahlverhaltens in diesem Falle; Parteipräferenz (Stimmzettelverfahren); Sympathieveränderungen gegenüber ausgewählten Politikern und Parteien seit der letzten Landtagswahl. 3.) An alle: Kenntnis von Entlassungen im eigenen Wirtschaftsbereich; Wahrscheinlichkeit eines Arbeitsplatzverlustes im nächsten Jahr, bezogen auf den Befragten sowie ein Haushaltsmitglied; Telefonbesitz. Demographie: Alter; Geschlecht; Familienstand; Konfession; Kirchgangshäufigkeit; Schulbildung; Berufliche Position; Berufstätigkeit; Haushaltseinkommen; Haushaltszusammensetzung; Befragter ist Haushaltsvorstand; Charakteristika des Haushaltsvorstands; Selbsteinschätzung der Schichtzugehörigkeit; Ortsgröße; Bundesland; Gewerkschaftsmitgliedschaft. Interviewerrating: Interviewdauer; Interviewtag; Interesse des Befragten am Interview; Anzahl der Kontaktversuche

States Study Spring 1974

The political opinion climate before the state parliament elections 1974 in the states Schleswig-Holstein, Hesse, Lower Saxony, Rhineland-Palatinate, North Rhine-Westphalia and Bavaria. Judgement on parties, politics and politicians. Topics: The questionnaire for all investigated states consists of a uniform ´federal part´ and a specific state part. 1. Federal part: judgement on current and future economic situation of the FRG; political interest and frequency of political conversation; memory of the most important political news; preferred federal chancellor; party preference (ballot procedure); memory of behavior at the polls in the state parliament election 1972; most important political tasks and party most able to fulfill these tasks; politicians with greatest influence in the Federal Republic; party inclination (three-way split); fear of loss of job; preferred politicians for the CDU leadership; attitude to the Young Union and the Jusos {Young Socialists}; perceived influence of Jusos on the SPD in general and on the local SPD; personal opinion leadership; making personal political preferences public in discussions; sympathy scale for the SPD, CDU/CSU and FDP; judgement on foreign policy and the importance of relations to the USA, the Soviet Union, the EC and the GDR; perceived changes within the parties since 1969. 2. State part: the state parts are largely identical in their survey contents. Merely the respective politician names were changed. Judgement on current and future economic situation of the state; most important political tasks and most able party; judgement on the degree of familiarity of selected state politicians; preferred prime minister; characteristics of the most important top politicians of the state parties; memory of personal behavior at the polls in the last state parliament election and intended voting decision in the next state parliament election; preferred coalition; attitude to a coalition commitment before the election; knowledge of coalition arrangements; sympathy scale for the state parties. Demography: age; sex; marital status; religious denomination; frequency of church attendance; school education; occupational training; employment; number of recipients of income; household income; respondent is head of household; characteristics of head of household; refugee status; memberships. Interviewer rating: interest of respondent in interview; number of contact attempts; length of interview; date of interview.Das politische Meinungsklima vor den Landtagswahlen 1974 in den Bundesländern Schleswig-Holstein, Hessen, Niedersachsen, Rheinland-Pfalz, Nordrhein-Westfalen und Bayern. Beurteilung von Parteien, Politik und Politikern. Themen: Der Fragebogen besteht für alle untersuchten Bundesländer aus einem einheitlichen "Bundesteil" und einem spezifischen Länderteil. 1. Bundesteil: Beurteilung der aktuellen und zukünftigen wirtschaftlichen Lage der BRD; politisches Interesse und Häufigkeit politischer Gespräche; Rückerinnerung an die wichtigsten politischen Nachrichten; präferierter Bundeskanzler; Parteipräferenz (Stimmzettelverfahren); Rückerinnerung an das Wahlverhalten bei der Landtagswahl 1972; wichtigste politische Aufgaben und kompetenteste Partei zur Aufgabenerfüllung; einflußreichster Politiker in der Bundesrepublik; Parteineigung (dreifacher Split); Furcht vor Arbeitsplatzverlust; präferierte Politiker für die CDU-Spitze; Einstellung zur jungen Union und den Jusos; perzipierter Einfluß der Jusos auf die SPD allgemein und auf die lokale SPD; eigene Meinungsführerschaft; Offenlegung eigener politischer Präferenzen in Diskussionen; Sympathie-Skalometer für die SPD, CDU/CSU und die FDP; Beurteilung der Außenpolitik und der Wichtigkeit der Beziehungen zu den USA, zur Sowjetunion, zur EG und zur DDR; wahrgenommene Veränderungen innerhalb der Parteien seit 1969. 2. Länderteil: Die Länderteile sind in ihren Befragungsinhalten weitgehend identisch. Es wurden lediglich die jeweiligen Politikernamen geändert. Beurteilung der aktuellen und zukünftigen wirtschaftlichen Lage des Bundeslandes; wichtigste politische Aufgaben und kompetenteste Partei; Beurteilung des Bekanntheitsgrades ausgewählter Landespolitiker; präferierter Ministerpräsident; Eigenschaften der wichtigsten Spitzenpolitiker der Landesparteien; Rückerinnerung an das eigene Wahlverhalten bei der letzten Landtagswahl und beabsichtigte Wahlentscheidung bei der nächsten Landtagswahl; bevorzugte Koalitionsbildung; Einstellung zu einer Koalitionszusage vor der Wahl; Kenntnis von Koalitionsabsprachen; Sympathie-Skalometer für die Landesparteien. Demographie: Alter; Geschlecht; Familienstand; Konfession; Kirchgangshäufigkeit; Schulbildung; Berufsausbildung; Berufstätigkeit; Anzahl Einkommensempfänger; Haushaltseinkommen; Befragter ist Haushaltsvorstand; Charakteristika des Haushaltsvorstands; Flüchtlingsstatus; Mitgliedschaften. Interviewerrating: Interesse des Befragten am Interview; Anzahl der Kontaktversuche; Interviewdauer; Interviewdatum

CDX2-driven leukemogenesis involves KLF4 repression and deregulated PPARγ signaling

Aberrant expression of the homeodomain transcription factor CDX2 occurs in most cases of acute myeloid leukemia (AML) and promotes leukemogenesis, making CDX2, in principle, an attractive therapeutic target. Conversely, CDX2 acts as a tumor suppressor in colonic epithelium. The effectors mediating the leukemogenic activity of CDX2 and the mechanism underlying its context-dependent properties are poorly characterized, and strategies for interfering with CDX2 function in AML remain elusive. We report data implicating repression of the transcription factor KLF4 as important for the oncogenic activity of CDX2, and demonstrate that CDX2 differentially regulates KLF4 in AML versus colon cancer cells through a mechanism that involves tissue-specific patterns of promoter binding and epigenetic modifications. Furthermore, we identified deregulation of the PPARγ signaling pathway as a feature of CDX2-associated AML and observed that PPARγ agonists derepressed KLF4 and were preferentially toxic to CDX2+ leukemic cells. These data delineate transcriptional programs associated with CDX2 expression in hematopoietic cells, provide insight into the antagonistic duality of CDX2 function in AML versus colon cancer, and suggest reactivation of KLF4 expression, through modulation of PPARγ signaling, as a therapeutic modality in a large proportion of AML patients

Epigenetic Upregulation of lncRNAs at 13q14.3 in Leukemia Is Linked to the <i>In Cis</i> Downregulation of a Gene Cluster That Targets NF-kB

<div><p>Non-coding RNAs are much more common than previously thought. However, for the vast majority of non-coding RNAs, the cellular function remains enigmatic. The two long non-coding RNA (lncRNA) genes <i>DLEU1</i> and <i>DLEU2</i> map to a critical region at chromosomal band 13q14.3 that is recurrently deleted in solid tumors and hematopoietic malignancies like chronic lymphocytic leukemia (CLL). While no point mutations have been found in the protein coding candidate genes at 13q14.3, they are deregulated in malignant cells, suggesting an epigenetic tumor suppressor mechanism. We therefore characterized the epigenetic makeup of 13q14.3 in CLL cells and found histone modifications by chromatin-immunoprecipitation (ChIP) that are associated with activated transcription and significant DNA-demethylation at the transcriptional start sites of <i>DLEU1</i> and <i>DLEU2</i> using 5 different semi-quantitative and quantitative methods (aPRIMES, BioCOBRA, MCIp, MassARRAY, and bisulfite sequencing). These epigenetic aberrations were correlated with transcriptional deregulation of the neighboring candidate tumor suppressor genes, suggesting a coregulation <i>in cis</i> of this gene cluster. We found that the 13q14.3 genes in addition to their previously known functions regulate NF-kB activity, which we could show after overexpression, siRNA–mediated knockdown, and dominant-negative mutant genes by using Western blots with previously undescribed antibodies, by a customized ELISA as well as by reporter assays. In addition, we performed an unbiased screen of 810 human miRNAs and identified the <i>miR-15/16</i> family of genes at 13q14.3 as the strongest inducers of NF-kB activity. In summary, the tumor suppressor mechanism at 13q14.3 is a cluster of genes controlled by two lncRNA genes that are regulated by DNA-methylation and histone modifications and whose members all regulate NF-kB. Therefore, the tumor suppressor mechanism in 13q14.3 underlines the role both of epigenetic aberrations and of lncRNA genes in human tumorigenesis and is an example of colocalization of a functionally related gene cluster.</p> </div

Hypomethylation at the TSS of the large ncRNAs affects the majority of CLL patients independent of 13q gene dosage and IGHV mutation status.

<p>(A) For validation of DNA-hypomethylation at different CpG islands in 13q14.3 (x-axis; for localization see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003373#pgen-1003373-g001" target="_blank">figure 1</a>), quantitative measurement with BioCOBRA and MassARRAY (“Rb”) technology was performed in B-cells of healthy donors (“h”, n = 15–19, white diamonds) and PBMCs of CLL patients (“p”, n = 47–82, black diamonds; “miR” = 308 bp fragment spanning miR-15a; “Rb” = intragenic CpG island). (B, C) CLL patients with different genomic aberrations show different degree of hypomethylation that is distinct from healthy probands younger (“young”) or older (“old”) than 45 years. DNA-methylation was quantified with BioCOBRA (D6) and MassARRAY (E6). Analyzed were patients with normal karyotype (“nk”), deletion of 13q14.3 (“13q”), deletion of 11q22-q23 (“11q”), deletion of 17p (“17p”) or combinations thereof. Statistics were performed using Wilcoxon rank sum test (*** p<0.001, ** p 0.001 to 0.01, * 0.01 to 0.05 and ns not significant p>0.05). (D) DNA-methylation of D6 or E6 is aberrant in 58/61 CLL patients. Black diamonds: Patient samples with two copies of the critical region in 13q, grey diamonds: samples with deletion of one copy of 13q14.3, white diamonds: healthy donor samples. Dashed lines represent lowest levels of methylation of non-malignant B-cells.</p

The critical region at 13q14.3 displays relaxed chromatin at the transcriptional start site (TSS) of the long non-coding RNA genes <i>DLEU1</i> and <i>DLEU2</i> in CLL cells.

<p>The critical region on 13q14.3 was analyzed for DNA-methylation (lanes 1–3), distribution of histone modifications (lanes 4, 5) and gene expression (lanes 6, 7). DNA-hypomethylation was detected in CLL cells at the transcriptional start sites of the lncRNA genes <i>DLEU1</i> and <i>DLEU2</i> variant <i>Alt1</i>. This finding coincides with enrichment of H3K4me2 and depletion of macroH2A, corroborating relaxation of 13q14.3 in CLL. While expression of the protein coding genes and <i>DLEU2</i> is decreased in CLL cells, lncRNA genes <i>DLEU1</i> and <i>DLEU2/Alt1</i> show upregulation, suggesting a direct regulation by DNA-methylation. Base pair positions on top refer to genome GRCh37 build hg18. The conserved CpG islands (A–E) are shown in green, less stringent CpG islands shown in light green. <i>Lane 1</i>: Methyl-CpG Immunoprecipitation (MCIp) allowed semi-quantitative DNA-methylation analysis in non-malignant B-cells sorted from healthy donors (n = 7) and CLL samples (n = 25). Precipitated DNA was hybridized onto a custom tiling microarray covering the 13q14.3 critical region. Depicted is the median log2 fold enrichment of CLL samples from which the median of log2 fold enrichment of healthy donor B-cell samples has been subtracted. <i>Lane 2</i>: Restriction digest-based analysis of DNA-methylation (aPRIMES) was performed in CpG islands C–E at 13q14.3 at 1 kbp resolution. Shown is the median log2 signal intensity of CLL patients from which the median of non-malignant B cell samples has been subtracted. <i>Lane 3</i>: Hypomethylation at D6 and E6 was validated by BioCOBRA and MassARRAY/Sequenome analyses (for details see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003373#pgen-1003373-g002" target="_blank">Figure 2</a>). Shown is the difference of the median percentage of methylation in CLL cells and non-malignant B cells. <i>Lanes 4,5</i>: ChIP was performed for macroH2A (lane 4) and H3K4me2 (lane 5). Precipitated DNA was quantified using qPCR. Enrichment was normalized to non-specific IgG and two control promoters (CDH12, HK2) that displayed similar enrichment for the two histone marks in CLL samples (n = 7) and peripheral blood mononuclear cells (PBMC) from healthy probands (n = 5). Enrichment was calculated as median log2 fold enrichment in precipitate vs. input for CLL samples after subtraction of enrichment from non-malignant B-cells. <i>Lane 6</i>: Gene expression profiling (GEP) was performed using bead chip arrays (Illumina) in CLL patients (n = 25) and sorted B-cells from healthy donors (n = 9). Plotted is the difference of log fold changes of CLL samples and non-malignant B cells. <i>Lane 7</i>: QRT-PCR for gene expression analysis of 13q14.3 candidate genes (for details see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003373#pgen-1003373-g003" target="_blank">Figure 3</a>).</p

Downregulation of 13q14.3 candidate tumor suppressor genes and upregulation of lncRNA genes <i>DLEU1</i> and <i>DLEU2/Alt1</i> correlates with DNA-methylation.

<p>(A–C) Expression of the protein-coding genes <i>KPNA3, C13ORF1</i> and <i>RFP2</i>, the miRNA genes <i>miR-15a</i> and <i>miR-16-1</i> and the lncRNA transcripts from genes <i>DLEU1, DLEU2</i> and <i>alternative transcript Alt1</i> were quantified with qRT-PCR in CLL cells from patients with retention of both copies of 13q14.3 (“p”, n = 34; see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003373#pgen.1003373.s007" target="_blank">Table S1</a>) and compared to B-cells from healthy donors (“h”, n = 20). Mean expression is indicated by a black horizontal line. “HKG” = housekeeping genes, average of <i>PGK, LMNB1</i> and <i>PPIA</i>. (D, E) DNA-methylation levels correlate with transcript levels of genes in 13q14.3: While expression (y-axis) of candidate tumor suppressor genes is directly correlated to DNA-methylation of regions D6 and E6 (x-axis; left panel), lncRNA genes are anti-correlated (right panel). Pearson correlation coefficients are color-coded, blue = negative, yellow = positive correlation. (F) Pearson coefficients of correlation of gene expression (rows) and DNA-methylation at D6 and E6 (columns), colour coded (see legend). DLEU1a = exon1 to exon4; DLEU1b = exon 1 to exon 2; ALT1a/b = exon1, two different amplicons (G) Significance values of Pearson correlation coefficients (t-distribution), values p<0.05 are coded green. (H) The bidirectional promoter of <i>DLEU1</i> and <i>DLEU2</i> and the flanking CpG island was cloned into pCpGL luciferase vector, either including D6 (orange) or excluding D6 (green). (I) Constructs depicted in (H) were either methylated <i>in-vitro</i> using SSsI methylase (m, dark bars) or left unmethylated (u, light bars) and subsequently transfected into Mec1, Granta519 and HeLa cells and luciferase activity quantified. Grey: Luciferase CMV expression vector (V) not containing CpGs was used as negative control, green: excluding D6 region, orange: including D6 region. Error bars signify standard deviation of 2 independent experiments, each performed in duplicate. No changes were detected for E6 (<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003373#pgen.1003373.s004" target="_blank">Figure S4G</a>–<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003373#pgen.1003373.s004" target="_blank">S4J</a>). (J) Binding of the DNA-methylation sensitive chromatin reader CTCF was enriched in CLL samples both in the D6 and E6 regions compared to non-malignant B-cells. CTCF-bound chromatin was immunoprecipitated and quantitated with qPCR. Specific binding was shown by quantification of a sequence not bound by CTCF 2 kb upstream of the DM1 insulator (“ctr”). Statistics were performed using Wilcoxon rank sum test (*** p<0.001, ** p 0.001 to 0.01, * 0.01 to 0.05 and ns not significant p>0.05).</p