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Experimental Therapeutics, Preclinical Pharmacology
Adenovirus-mediated p53 Gene Transduction Inhibits Telomerase Activity Independent of Its Effects on Cell Cycle Arrest and Apoptosis in Human Pancreatic Cancer Cells
Masahiro Kusumoto, Takahiro Ogawa, Kazuhiro Mizumoto, Hikaru Ueno, Hideaki Niiyama, Norihiro Sato, Masafumi Nakamura and Masao Tanaka
Masahiro Kusumoto
Takahiro Ogawa
Kazuhiro Mizumoto
Hikaru Ueno
Hideaki Niiyama
Norihiro Sato
Masafumi Nakamura
Masao Tanaka
DOI: Published August 1999
Evidence for a relationship between overexpression of wild-type p53 and telomerase activity remains controversial. We investigated whether p53 gene transduction could cause telomerase inhibition in pancreatic cancer cell lines, focusing on the relation of transduction to growth arrest, cell cycle arrest, and apoptotic cell death. The cells were infected with recombinant adenovirus expressing wild-type p53 or p21WAF1 at a multiplicity of infection of 100 or were continuously exposed to 10 μm VP-16, which is well known to induce apoptosis. Adenovirus-mediated p53 gene transduction caused G1 cell cycle arrest, apoptosis, and resultant growth inhibition in MIA PaCa-2 cells; the cell number 2 days after infection was 50% of preinfection value, and 13% of the cells were dead. Moreover, the transduction resulted in complete depression of telomerase activity through down-regulation of hTERT mRNA expression. In contrast, p21WAF1 gene transduction only arrested cell growth and cell cycle at G1 phase, and VP-16 treatment inhibited cell growth with G2-M arrest and apoptosis; after treatment, the cell number was 73% of pretreatment, and 12% of the cells were dead. Neither p21WAF1 gene transduction nor VP-16 treatment caused telomerase inhibition. Similar results were obtained in two other pancreatic cancer cell lines, SUIT-2 and AsPC-1. Thus, our results demonstrate that the p53 gene transduction directly inhibits telomerase activity, independent of its effects on cell growth arrest, cell cycle arrest, and apoptosis.
Telomerase is a ribonucleoprotein enzyme containing an integral short RNA template that directs the synthesis of telomeric repeats (5′-TTAGGG-3′) at the end of replicating chromosomes called telomeres (1 , 2) . In most human somatic cells, telomerase activity is undetectable, and the telomere length is progressively shortened during cell proliferation (3 , 4) , leading to cellular senescence (5 , 6) . In contrast, telomerase is activated in the vast majority of cancer cells (7 , 8) . We previously detected telomerase activity in 80% of a series of surgically resected pancreatic cancers (9) , and furthermore, we have used telomerase activity in the pancreatic juice as a useful diagnostic tool (10) .
Reportedly, telomerase activity is regulated in a cell cycle-dependent manner (11) . For example, maximal telomerase activity is detected in the S phase, whereas it is barely detectable in the G2-M phase (12) . There are some reports that, although telomerase is active throughout the cell cycle in dividing, immortal cells, its activity is repressed in cells that exit the cell cycle (13 , 14) ; therefore, its activity correlates with growth rate (15) and cellular proliferation (16) . Telomerase activity is also apparently suppressed by apoptosis induced by anticancer chemotherapeutic agents (17) .
p53 gene mutations and/or abnormal p53 protein accumulation are reported in ∼60% of pancreatic cancers (18 , 19) and are associated with its poor prognosis (20 , 21) . Wild-type p53 protein can transcriptionally transactivate the p21WAF1 gene, which mediates G1 cell cycle arrest by causing inhibition of the cyclin-dependent kinase activity required for progression from the G1 to the S phase (22 , 23) . Moreover, wild-type p53 protein up-regulates Bax expression, which, in turn, inhibits Bcl-2 expression, resulting in apoptosis induction as a strictly programmed mechanism for removal of damaged cells (24) .
It was reported that induced expression of wild-type p53 resulted in the down-regulation of telomerase activity in immortalized fibroblasts (25) and that telomerase activity was significantly reduced in lung cancer cell lines transfected with sense p53 cDNA (26) . In contrast, Maxwell et al. (27) proposed that telomerase activity is unaffected by overexpression of p53 in immortalized endothelial cells, suggesting that the evidence for a relationship between overexpression of p53 and telomerase activity remains controversial.
Here, we investigated whether adenovirus-mediated p53 gene transduction causes telomerase inhibition in human pancreatic cancer cells, focusing on the relation of transduction to cell growth arrest, cell cycle arrest, and apoptotic cell death.
Cell Lines.
Three human pancreatic cancer cell lines were used: MIA PaCa-2, generously provided by the Japanese Cancer Resource Bank (Tokyo, Japan), and SUIT-2 and AsPC-1, generously provided by Dr. H. Iguchi (National Kyushu Cancer Center, Fukuoka, Japan). Point mutations of the p53 gene were found in two cell lines: a C→T transversion at the first base of codon 248 in MIA PaCa-2 cells and a G→A transversion at the second base of codon 273 in SUIT-2 cells (28) . A single-base deletion between codons 134 and 135 (TTT TGC→TTTGC) was observed in AsPC-1 cells, which resulted in the appearance of a stop codon (TGA) between codons 169 and 170 (ATG ACG→A TGA CG; Ref. 28 ). The cell lines were cultured in DMEM (Life Technologies, Inc., Rockville, MD) supplemented with 10% fetal bovine serum, 100 units/ml penicillin, and 100 mg/ml streptomycin.
Preparation of Adenoviral Vectors.
Replication-defective E1- and E3- adenoviral vectors expressing either human wild-type p53 (Ad-p53; Ref. 29 ), p21WAF1 (Ad-p21; Ref. 30 ), or bacterial human β-galactosidase (Ad-lacZ; Ref. 31 ) were prepared as described previously (32, 33, 34) . All cDNA were driven by a CA promoter composed of a cytomegalovirus enhancer and chicken β-actin promoter (35) . The recombinant adenoviruses were purified by ultracentrifugation through a CsCl2 gradient, followed by extensive dialysis. Contamination by wild-type adenovirus was excluded by PCR designed for E1 amplification. The virus titer was assessed by a plaque formation assay using 293 cells.
VP-16, a topoisomerase II inhibitor, was obtained in a powder form from Nippon Kayaku Co. (Tokyo, Japan), dissolved in DMSO (0.1% final concentration), and added to aliquots to a final concentration of 10 μm.
Cell Growth Assay.
Exponentially growing cells were plated at an initial density of either 4 × 104 cells (MIA PaCa-2 and SUIT-2 cells) or 6 × 104 cells (AsPC-1 cells) per well in 24-well plates and incubated overnight at 37°C in a humidified atmosphere containing 10% CO2 in DMEM containing 10% FBS. Then, the cells were infected with PBS, Ad-lacZ, Ad-p21, or Ad-p53 at a MOI3 of 100 in 150 μl of PBS or were continuously exposed to 10 μm VP-16 at 37°C in a humidified atmosphere containing 10% CO2. After 1 h, the culture supernatant was replaced with 1 ml of fresh culture medium. For 4 days, cells floating in the culture medium and attached cells collected by trypsinization (200 μl/well) were combined in a 1500-μl Eppendorf tube, and half were counted with a particle distribution analyzer, CDA500 (Sysmex, Kobe, Japan). The remaining cell pellets were stored at −80°C for the telomerase activity assay.
Cell Viability Assay.
The percentage of dead cells was determined by staining with propidium iodide and performing an assay using a fluorescence multiple-well plate reader, Cytofluor II (PerSeptive Biosystems, Framingham, MA), as described previously (36) . Briefly, cells were plated and treated as described above. Propidium iodide was added to aliquots (final concentration, 30 μm) to label nuclei of dead cells. Fluorescence was measured after incubation for 60 min at 530-nm (excitation) and 645-nm (emission) wavelengths. Subsequently, digitonin was added to aliquots (final concentration, 300 μm) to permeabilize all cells and label all nuclei with propidium iodide. Fluorescence was measured again after incubation for 30 min to obtain a value corresponding to 100% cell death. The proportion of the former fluorescence to the latter was defined as the percentage of dead cells.
Measurement of DNA Synthesis.
Cells were plated and treated as described above. After 48 h, the cells were pulsed for 4 h with 1 μCi/ml [3 H]thymidine. The incorporation of [3 H]thymidine into the trichloroacetic acid-insoluble material was measured using a scintillation counter.
Morphological Analysis.
Cells were plated on a coated coverslip in a 24-well plate and treated as described above. The cells attached to the coverslip were stained with 10 μg of propidium iodide and observed under UV fluorescence microscopy.
Western Blot Analysis.
Floating and trypsinized adherent cells were combined and washed with PBS. Cells were then lysed in SDS solubilization buffer [62.5 mm Tris-HCl (pH 6.8), 10% glycerol, 5% β-mercaptoethanol, and 2% SDS]. Fifteen μg of protein from the samples were boiled for 5 min, subjected to 12% SDS-PAGE, and transferred to a Hybond-ECL membrane (Amersham, Tokyo, Japan). The membrane was blocked in PBS containing 5% dried milk and 0.1% Tween 20 and probed with a primary antibody (mouse antihuman p53 monoclonal antibody OP09 or p21 monoclonal antibody OP76; Calbiochem, Cambridge, MA) and a secondary antibody (horseradish peroxidase-conjugated rabbit antimouse IgG1; ICN Pharmaceuticals, Inc., Aurora, OH). An Amersham ECL chemiluminescent Western system (Amersham) was used to detect secondary probes.
Flow Cytometric Analysis.
Floating and trypsinized adherent cells were collected, washed with PBS, subjected to cold 70% ethanol for 4 h at 4°C, washed with PBS, suspended in phosphate-citrate buffer for 30 min at room temperature, and resuspended in PBS containing 1 mg/ml RNase A for 30 min at room temperature. Samples were then stained with propidium iodide at 50 μg/ml and analyzed in a cell sorter (FACScan; Becton Dickinson, Mountain View, CA) for DNA content. G0 /G1, S, and G2-M populations were quantified using the ModFit LT program for the Macintosh (Version. 1.01; Verity Software House, Inc.).
Telomerase Activity Assay.
Telomerase was assayed by the PCR-based telomeric repeat amplification protocol described previously (7) , with minor modifications. The stored cell pellets obtained in the cell growth assay were suspended in 2 μl per 1000 cells of 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid lysis buffer [10 mm Tris (pH 7.5), 1 mm MgCl2, 1 mm EGTA, 0.1 mm 4-(2-aminoethyl)-benzenesulfonyl fluoride hydrochloride, 5 mm β-mercaptoethanol, 0.5% 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid, 10% glycerol, and 1 mg/ml each protease inhibitor (antipain, leupeptin, phosphoramidon, elastatinal, pepstatin A, and chymostatin; Peptide Institute, Osaka, Japan)] and incubated for 30 min on ice. The lysate were centrifuged at 15,000 × g for 20 min at 4°C. Two μl of the extracts, equivalent to 1000 cells, were incubated in a solution of 20 mm Tris (pH 8.3), 1 mm MgCl2, 63 mm KCl, 0.005% Tween 20, 1 mm EGTA, 50 μm dNTP, 0.3 μCi [α-32 P]dCTP, 2 units of Taq DNA polymerase (Promega, Madison, WI), and 0.1 μg of TS primer (5′-AATCCGTCGAGCAGAGAGTT-3′) at 20°C for 30 min and then heated to 90°C for 3 min to inactivate telomerase activity. After the inactivation of telomerase, 0.1 μg of CX primer (5′-CCCTTACCCTTACCCTTACCCTTAA-3′) was added, and the reaction mixture was subjected to 31 PCR cycles at 94°C for 30 s, 50°C for 30 s, and 72°C for 90 s (2 min for the final step). Half of the PCR products were analyzed by electrophoresis in 0.5× Tris-borate EDTA buffer on 12% polyacrylamide nondenaturing gels. Signal intensity was measured by NIH Image (Version 1.60; NTIS, Springfield, VA). The relative densities of the telomeric ladders in each sample were expressed as percentages of pretreated cells after subtraction from background. A 36-bp internal standard (Oncor, Inc., Gaithersburg, MD) was used as internal control.
RT-PCR Analysis.
RNA was isolated from MIA PaCa-2 cells treated as described above. Two μg of total RNA were reverse-transcribed. The resulting cDNA was subjected to PCR with the primers as follows: hTERT, 5′-CGGAAGAGTGTCTCTGGAGCAA-3′ and 5′-GGATGAAGCGGAGTCTGGA-3′; and β-actin (as an internal control), 5′-GTGGGGCGC-CCCAGGCACCA-3′ and 5′-TGTAGCCGCGCTCGGTGAG-GA-3′. The amplification reactions were performed with an initial incubation step at 94°C for 3 min, followed by 33 cycles at 94°C for 45 s, 60°C for 45 s, and 72°C for 90 s. Ten μl of the reaction products were subjected to electrophoresis in 2% agarose gel and visualized by ethidium bromide staining.
Expression of p53 and p21WAF1 Proteins in MIA PaCa-2 Cells 2 Days after Treatment.
The expression of p53 and p21WAF1 proteins was confirmed by Western blotting (Fig. 1)<$REFLINK> . Although MIA PaCa-2 cells infected with PBS or Ad-lacZ expressed p53 slightly, p21WAF1 protein was hardly detected in the cells. The cells transduced with the p21WAF1 gene showed low levels of p53 and high levels of p21WAF1 protein, and those transduced with the p53 gene showed overexpression of both p53 and p21WAF1 proteins. In contrast, the cells treated with VP-16 showed high levels of p53 protein without induction of p21WAF1, which indicates overexpression of mutated p53, as suggested previously (37) .
Western blot analysis of p53 and p21WAF1 levels in MIA PaCa-2 cells 2 days after infection with PBS, Ad-lacZ, Ad-p21, or Ad-p53 or treatment with VP-16. Fifteen μg of protein were loaded into each lane. Blots were probed using p53 or p21WAF1 monoclonal antibodies and visualized using enhanced chemiluminescence.
Effect of p53 and p21WAF1 Gene Transduction and VP-16 Treatment on Telomerase Activity.
In MIA PaCa-2 cells infected with either PBS or Ad-lacZ, the telomerase activity was not altered for 4 days (Fig. 2)<$REFLINK> . In MIA PaCa-2 cells transduced with the p21WAF1 gene, the telomerase activity was only slightly reduced 3 days after infection; however, in those transduced with p53 gene, activity was completely absent 2 days after infection. Telomerase activity was not affected by the mixing of extract from control cells with that from p53-transduced cells (data not shown), suggesting that the lack of telomerase activity is not due to a trans-acting inhibitor of activity. In contrast, telomerase activity increased in the VP-16-treated cells 2 days after treatment and was the same as that in the pretreated cells 3 days after treatment. In both SUIT-2 and AsPC-1 cells, 3 days after treatment, p21WAF1 gene transduction and VP-16 treatment did not decrease telomerase activity, but p53 gene transduction caused a significant depression of this activity (Table 1)<$REFLINK> . The telomerase inhibition by p53 gene transduction was also observed in other cell types such as a breast cancer cell line MCF-7 and a gallbladder cancer cell line GB-d1 after 3 days (data not shown).
Daily changes in telomerase activity in MIA PaCa-2 cells infected with PBS, Ad-lacZ, Ad-p21, or Ad-p53 or treated with VP-16. Telomerase activity assay was measured using a PCR-based telomerase assay. The relative densities of the telomeric ladders in each sample are expressed as percentages of pretreated cells after subtraction from background. Data points, mean relative telomerase activities of three independent wells; bars, SD.
Effects of p53 and p21WAF1 gene transduction on SUIT-2 and AsPC-1 cellsa
Effect of p53 and p21WAF1 Gene Transduction and VP-16 Treatment on hTERT mRNA Expression 2 Days after Treatment.
Down-regulation of hTERT mRNA expression was clearly demonstrated in MIA PaCa-2 cells 2 days after infection with Ad-p53 (Fig. 3)<$REFLINK> . In contrast, the expression levels of hTERT mRNA on RT-PCR were almost unchanged in the cells infected with PBS, Ad-lacZ, and Ad-p21 and were slightly increased in those treated with VP-16.
hTERT mRNA expression in MIA PaCa-2 cells 2 days after infection with PBS, Ad-lacZ, Ad-p21, or Ad-p53 or treatment with VP-16 (top). Control RT-PCR using β-actin primers is also presented (bottom).
Effect of p53 and p21WAF1 Gene Transduction and VP-16 Treatment on Cell Growth in Vitro.
The time course of MIA PaCa-2 cell growth in vitro after treatment is shown in Fig. 4<$REFLINK> . Cells infected with Ad-lacZ had growth rates similar to those of mock-infected cells, which grew exponentially, whereas p21WAF1 gene transduction and VP-16 treatment completely inhibited cell growth. A more severe loss of cell numbers was observed in the cells transduced with p53 gene. Changes in numbers of SUIT-2 and AsPC-1 cells 3 days after treatment are summarized in Table 1<$REFLINK> . Although treatment with VP-16 significantly inhibited cell growth of all cell lines, p53 and p21WAF1 gene transduction had less effect on inhibition of cell growth in SUIT-2 and AsPC-1 cells than it did in MIA PaCa-2 cells.
Daily changes in total cell number of MIA PaCa-2 cells infected with PBS, Ad-lacZ, Ad-p21, or Ad-p53 or treated with VP-16. Cells plated (4 × 104) and incubated overnight were infected with each viral vector at a MOI of 100 for 1 h or were exposed continuously to 10 μm VP-16. Floating and attached cells were counted using a particle distribution analyzer. Data points, mean total cell numbers of three independent wells; bars, SD.
Effect of p53 and p21WAF1 Gene Transduction and VP-16 Treatment on Apoptotic Cell Death.
Cell viability assays were performed based on the binding of propidium iodide to the nuclei of cells whose plasma membranes have become permeable due to cell death (Fig. 5, a–e)<$REFLINK> . After addition of digitonin, the nuclei of all cells were labeled with propidium iodide (Fig. 5f)<$REFLINK> . Fluorescence of propidium iodide, measured with a fluorescence multiple-well plate reader, increased in proportion to the number of permeabilized cells. Fig. 6<$REFLINK> depicts the time course of the percentage of MIA PaCa-2 dead cells after treatment. Degraded cells or cells that had disappeared were not counted in the cell growth assay and are not reflected in the percentage of dead cells. Cells infected with Ad-lacZ showed a percentage of dead cells similar to that of parental cells and had a similar morphology as parental cells (Fig. 5, a and b)<$REFLINK> . In the cells transduced with the p21WAF1 gene, the percentage of dead cells was slightly increased, and the morphology was altered (e.g., an increased cytoplasmic-to-nuclear ratio), but apoptotic changes were not prominent (Fig. 5c)<$REFLINK> . In contrast, both p53 gene transduction and treatment with VP-16 caused a marked time-dependent increase in the percentage of dead cells and induced specific morphological changes in some cells such as shrunken or fragmented nuclei, which are characteristics of apoptotic cell death (Fig. 5, d and e)<$REFLINK> . In both SUIT-2 and AsPC-1 cells, 3 days after treatment, p53 gene transduction and treatment with VP-16 increased the percentage of dead cells, but the effect was less than that on MIA PaCa-2 cells (Table 1)<$REFLINK> .
Propidium iodide staining of MIA PaCa-2 cells 2 days after infection with PBS (a), Ad-lacZ (b), Ad-p21 (c), or Ad-p53 (d) or treatment with VP-16 (e). Nuclei of dead cells were stained with propidium iodide (red). Arrows, apoptotic changes. After addition of digitonin, nuclei of all mock-infected cells were labeled with propidium iodide (f).
Daily changes in percentage of dead cells in MIA PaCa-2 cells infected with PBS, Ad-lacZ, Ad-p21, or Ad-p53 or treated with VP-16. Cells plated (4 × 104) and incubated overnight were infected with each viral vector at a MOI of 100 for 1 h or were exposed continuously to 10 μm VP-16. Percentage of dead cells was determined by staining with propidium iodide and performing an assay pre- and postaddition of digitonin using a fluorescence multiple-well plate reader. Columns, mean percentages of dead cells of three independent wells; bars, SD.
DNA Synthesis and Cell Cycle Transitions in MIA PaCa-2 Cells 2 Days after Treatment.
[3 H]Thymidine incorporation of mock-infected MIA PaCa-2 cells was assigned a value of 100 ± 2 (mean ± SD). The relative incorporations of Ad-lacZ-, Ad-p21-, and Ad-p53-infected cells and that of VP-16-treated cells were 90 ± 3, 40 ± 4, 24 ± 2, and 30 ± 4, respectively. Cell cycle analysis of MIA PaCa-2 cells 2 days after treatment is depicted in Fig. 7<$REFLINK> . Neither mock-infected nor Ad-lacZ-infected cells showed a specific pattern of cell cycle distribution. In contrast, flow cytometry of cells infected with either Ad-p21 or Ad-p53 demonstrated a decrease of S phase and an increase of G0 /G1 phase, thus indicating G1 arrest. Moreover, cells transduced with the p53 gene exhibited a sub-G1 area, indicative of apoptotic cell death. VP-16 caused G2-M arrest with apoptosis.
Cell cycle analysis of MIA PaCa-2 cells 2 days after infection with PBS, Ad-lacZ, Ad-p21, or Ad-p53 or treatment with VP-16. Floating and attached cells were analyzed in a cell sorter. Arrows, population of cells in the sub-G1 group (apoptotic cells). Proportions of G0/G1, S, and G2-M cells were quantified (columns).
Here, we demonstrated that adenovirus-mediated p53 gene transduction caused a marked inhibition of the pancreatic cancer cell growth with apoptotic cell death and G1 arrest and that p21WAF1 gene transduction arrested the cell growth and the cell cycle at the G1 phase without prominent apoptotic changes. These results suggest that recombinant adenoviral vectors containing tumor suppressor genes can be used effectively for gene therapy of pancreatic cancer and that p53 gene therapy is more effective than the p21WAF1 gene therapy, probably due to induction of apoptosis.
Reportedly, telomeres on chromosome 17p, where the p53 gene is encoded, are shortened more rapidly than others during successive cell divisions (38) . The loss of telomeric repeats would cause additional genetic changes as a consequence of chromosome instability (39) . Although a direct relationship between p53 alteration and telomerase activation has not been observed, it is possible that telomerase activation is promoted by alterations in the p53 gene. Some investigators have reported a close relationship between overexpressed p53 and telomerase activity. Induced expression of wild-type p53 in immortalized fibroblasts resulted in the down-regulation of telomerase activity after 48 h (25) , and telomerase activity was significantly reduced in lung cancer cell lines transfected with sense p53 cDNA and increased in those transfected with antisense p53 cDNA (26) . Here, we demonstrated that p53 gene transduction caused complete inhibition of telomerase activity in all three pancreatic cancer cell lines. In addition, we demonstrated that MIA PaCa-2 cells treated with VP-16 showed high levels of mutant p53, and we detected significantly high levels of telomerase activity in this cell line. These findings may indicate that there is a linkage between mutant p53 gene expression and telomerase activation, a supposition that is consistent with a previous report that telomerase is activated many population doublings after introduction with mutant p53 into normal human mammary epithelial cells (40) . These results seems to be supported by the fact that mutant p53 can transactivate c-Myc (41) , which induces telomerase in normal human fibroblasts and mammary epithelial cells (42) . In contrast, Maxwell et al. (27) proposed that telomerase activity is unaffected by overexpression of p53 and apoptosis in immortalized endothelial cells; however, they measured telomerase activity 18 h postinfection. We also demonstrated that telomerase activity was unaffected by p53 gene transduction 24 h postinfection.
The p53 tumor suppressor gene plays important roles in regulating the cell cycle and in apoptosis; however, our results showed that both cell growth inhibition and cell cycle arrest had little influence on telomerase activity of pancreatic cancer cells. We observed just a slight decrease or no change in telomerase activity of the cells infected with recombinant adenovirus harboring the p21WAF1 gene compared to the preinfectious, mock- or Ad-lacZ-infected cells in all three pancreatic cancer cell lines. Kallassy et al. (43) reported that retrovirus-mediated p21WAF1 gene transduction led to the suppression of telomerase activity in p53-mutated immortalized human keratinocytes 14 days after infection. The discrepancy between their findings and ours may be based on differences in conditions such as different viral vectors, cell lines, and/or duration from viral infection to measurement of telomerase activity. In addition, our results showed that there was no relation between induction of apoptosis and inactivation of telomerase in pancreatic cancer cells because the treatment with VP-16, which induced the G2-M cell cycle arrest and apoptotic cell death, did not repress telomerase activity.
Here, we further showed that telomerase inhibition by p53 gene transduction correlates with decline in hTERT mRNA expression, suggesting that p53 regulates telomerase at the transcriptional level in MIA PaCa-2 cells. Currently, we are examining the effect of p53 gene transduction on hTERT expression in various types of cells with different p53 status. This would provide a clue to understand a signaling pathway for telomerase-inhibitory effect by overexpressed p53. Although the precise mechanism remains to be elucidated, our results suggest that the p53 gene transduction directly inhibits telomerase activity in pancreatic cancer cells through down-regulation of hTERT mRNA expression.
We thank Takaaki Kanemaru, Morphology Core, Kyushu University Faculty of Medicine, for expert technical assistance. In addition, we are grateful to Dr. Li Zhang and Shoko Nishio for their technical assistance.
↵1 Supported in part by Grant-in-Aid 10671196 (to T. O.) from the Ministry of Education, Science and Culture, Japan.
↵2 To whom requests for reprints should be addressed, at Department of Surgery 1, Kyushu University Faculty of Medicine, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan. Phone: 81-92-642-5441; Fax: 81-92-642-5457; E-mail: takas1{at}med.kyushu-u.ac.jp
↵3 The abbreviations used are: MOI, multiplicity of infection; RT-PCR, reverse transcriptase-PCR.
Accepted May 12, 1999.
Received March 12, 1999.
Revision received May 11, 1999.
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Maxwell S. A., Capp D., Acosta S. A. Telomerase activity in immortalized endothelial cells undergoing p53-mediated apoptosis. Biochem. Biophys. Res. Commun., 241: 642-645, 1997.
Iguchi H., Morita D., Yasuda D., Takayanagi R., Ikeda Y., Takada Y., Shimazoe T., Nawata H., Kono A. Alterations of the p53 tumor suppressor gene and Ki-ras oncogene in human pancreatic cancer-derived cell lines with different metastatic potential. Oncol. Rep., 1: 1223-1227, 1994.
Takayama K., Ueno H., Pei X. H., Nakanishi Y., Yatsunami J., Hara N. The levels of integrin αV β5 may predict the susceptibility to adenovirus-mediated gene transfer in human lung cancer cells. Gene Ther., 5: 361-368, 1998.
Ueno H., Masuda S., Nishio S., Li J. J., Yamamoto H., Takeshita A. Adenovirus-mediated transfer of cyclin-dependent kinase inhibitor-p21 suppresses neointimal formation in the balloon-injured rat carotid arteries in vivo. Ann. N. Y. Acad. Sci., 811: 401-411, 1997.
Ueno H., Li J. J., Tomita H., Yamamoto H., Pan Y., Kanegae Y., Saito I., Takeshita A. Quantitative analysis of repeat adenovirus-mediated gene transfer into injured canine femoral arteries. Arterioscl. Thromb. Vasc. Biol., 15: 2246-2253, 1995.
Miyake S., Makimura M., Kanegae Y., Harada S., Sato Y., Takamori K., Tokuda C., Saito I. Efficient generation of recombinant adenoviruses using adenovirus DNA-terminal protein complex and a cosmid bearing the full-length virus genome. Proc. Natl. Acad. Sci. USA, 93: 1320-1324, 1996.
Ueno H., Yamamoto H., Ito S., Li J. J., Takeshita A. Adenovirus-mediated transfer of a dominant-negative H-ras suppresses neointimal formation in balloon-injured arteries in vivo. Arterioscl. Thromb. Vasc. Biol., 17: 898-904, 1997.
Ueno H., Li J. J., Masuda S., Qi Z., Yamamoto H., Takeshita A. Adenovirus-mediated expression of the secreted form of basic fibroblast growth factor (FGF-2) induces cellular proliferation and angiogenesis in vivo. Arterioscl. Thromb. Vasc. Biol., 17: 2453-2460, 1997.
Niwa H., Yamamura K., Miyazaki J. Efficient selection for high-expression transfectants with a novel eukaryotic vector. Gene, 108: 193-199, 1991.
Nieminen A. L., Gores G. J., Bond J. M., Imberti R., Herman B., Lemasters J. J. A novel cytotoxicity screening assay using a multiwell fluorescence scanner. Toxicol. Appl. Pharmacol., 115: 147-155, 1992.
Chresta C. M., Masters J. R. W., Hickman J. A. Hypersensitivity of human testicular tumors to etoposide-induced apoptosis is associated with functional p53 and a high Bax:Bcl-2 ratio. Cancer Res., 56: 1834-1841, 1996.
Martens U. M., Zijlmans J. M. J. M., Poon S. S. S., Dragowska W., Yui J., Chavez E. A., Ward R. K., Lansdorp P. M. Short telomeres on human chromosome 17p. Nat. Genet., 18: 76-80, 1998.
Hiyama K., Ishioka S., Shirotani Y., Inai K., Hiyama E., Murakami I., Isobe T., Inamizu T., Yamakido M. Alterations in telomeric repeat length in lung cancer are associated with loss of heterozygosity in p53 and Rb. Oncogene, 10: 937-944, 1995.
Gollahon L. S., Shay J. W. Immortalization of human mammary epithelial cells transfected with mutant p53 (273his). Oncogene, 12: 715-725, 1996.
Frazier M. W., He X., Wang J., Gu Z., Cleveland J. L., Zambetti G. P. Activation of c-myc gene expression by tumor-derived p53 mutants requires a discrete C-terminal domain. Mol. Cell. Biol., 18: 3735-3743, 1998.
Wang J., Xie L. Y., Allan S., Beach D., Hannon G. J. Myc activates telomerase. Genes Dev., 12: 1769-1774, 1998.
Kallassy M., Martel N., Damour O., Yamasaki H., Nakazawa H. Growth arrest of immortalized human keratinocytes and suppression of telomerase activity by p21WAF1 gene expression. Mol. Carcinog., 21: 26-36, 1998.
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GENE HOGLAN On Next DARK ANGEL Album: 'We're Just Trying To Make The Most Aggressive Music That We Can Make'
DARK ANGEL drummer Gene Hoglan spoke to "Poppitt's Corner", the video podcast of MADROST mainman Tanner Poppitt, about the reactivated California thrash metal pioneers' plans for new music. He said (as transcribed by BLABBERMOUTH.NET): "We've been playing a lot of live shows, and we tend to play festivals 'cause that just kind of works out the best. We're writing the next album, and I've been concentrating on writing that, and [DARK ANGEL guitarist] Jim Durkin is also writing. At the moment, Jim, he's had to take a little bit time off from some live shows, but that was always kind of built into our system. 'Cause when we got ourselves back together, he was, like, 'Hey, guys, just so you know, there's gonna be some times somewhere in the future where I might not be able to make certain shows, or there could an entire run that I might not be able to make. As long as you guys are cool moving forward with it. And we were able to get a good eight years with Jim, at least. So there you go."
He added: "We're just writing, and we're just trying to make it the most aggressive DARK ANGEL music that we can make."
This past August, Hoglan told Oxygène Radio's "Metal Zone" that he and his bandmates were hoping to have a new DARK ANGEL album out "by the middle part of 2023."
DARK ANGEL's first CD since 1991's "Time Does Not Heal" will feature the lineup that has performed sporadic live shows since reuniting in 2013: Hoglan, Durkin, Eric Meyer (guitar),Ron Rinehart (vocals) and Michael Gonzalez (bass).
Durkin has been sitting out some of DARK ANGEL's recent gigs. He was replaced at the shows by Hoglan's wife Laura Christine.
This past February, Hoglan said during a Twitch livestream from his home that he and his DARK ANGEL bandmates were "in the process" of making a new album. "That's actually one of the reasons I can't be doing TESTAMENT stuff this year. Because, [with] DARK ANGEL, we are writing a record, we are moving forward," he said. "Just a week and a half ago, two weeks ago, I just had Mr. Eric Meyer from DARK ANGEL in this room. We were jamming it out, we wrote a new DARK ANGEL song.
"I've been moving forward with writing. We have gigs coming up, so this year, it won't be heavily, heavily, heavily DARK ANGEL, but now that my schedule's open, I can commit a little harder to some DARK ANGEL gigs," he explained. "We do intend to be in the studio before the end of this year. So I have a lot of work ahead of me, and so does Jim Durkin. He's been sending over songs, riffs, stuff like that, [which has been] super exciting. We're all of the same mindset of making this just the most absolutely kick-ass DARK ANGEL record that we can."
DARK ANGEL released two albums with Don Doty on vocals — 1984's "We Have Arrived" and 1986's "Darkness Descends" — before he exited the group and was replaced by Rinehart (after a brief stint with Jim Drabos in 1987). The band issued two more studio LPs — 1989's "Leave Scars" and the aforementioned "Time Does Not Heal" — before calling it quits in 1992.
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Shop Artist's Full Portfolio
I work using a combination of techniques developed in Japanese lacquer art, European medieval religious art and 'faux' painting Working on wood or composition board panels, I use various combinations of the following media: Japan paints (finely ground oil pigments that dry in a day and can be sanded), metallic powders, powdered pigments, varnishes, precious and common metal leaf, and acids.
I build up layers of paint, varnish and metal leaf, often sanding through the upper layers to expose the under painting. Some of the more complex pieces have up to fifty or more layers. The surfaces that are created are inherently abstract and free associative. By manipulating these properties the finished image is produced.
I consider myself an icon maker. My main source of inspiration – besides exploring the artistic possibilities of the techniques and media I use – is nature. Whether pictorial or abstract, nature is generally at the heart of the work. Being both an environmentalist and an artist, I hope to convey to others the sense of sacredness that I find in the natural world.
Solo Exhibition:
200/2001 Paintings by Ernest Ely, Gallery at 400 Castro, San Francisco, CA
2014 222 Paul Mahder Gallery, Healdsburg, CA
2012 Paul Mahder Gallery, San Francisco, CA
2005 Blue Room Gallery, San Francisco, CA
Living Proof, LGBT Center, San Francisco, CA
Locos Peligrosos, Mission Cultural Center, San Francisco, CA
Thoreau Center of Sustainability, Presidio San Francisco, CA
Spice, Catherine Clark Gallery, San Francisco, CA
2002 Scapes, Space 743, San Francisco, CA
Visual Aid Annual Exhibition, San Francisco, CA
Toomey Tourell Gallery, San Francisco, CA
Traveling Show, Positive
San Francisco, Main Library, San Francisco, CA
Amfar Convention, Bill Graham Civic Auditorium, San
Francisco, CA
SF LGBT Community Center, San Francisco, CA
2001 Crossing Bridges, CBS Market Watch, San Francisco, CA
2000 Visual Aid Annual Exhibition, San Francisco, CA
1999 New Conservatory Theater, San Francisco, CA
1997 McKesson Building Gallery, San Francisco, CA
1996 de Young Museum, San Francisco, CA
Levi Strauss Plaza, San Francisco, CA
1994 OPTS Art Show, Ansel Adams Center, San Francisco, CA
1993 College of Marin Gallery, Kentfield, CA
1992 Selections 1991, SomArts Gallery, San Francisco, CA
1991 Max Nugus, The Cannery, San Francisco, CA
Open Studios, San Francisco, CA
Artist Equity Show, SomArts Gallery, San Francisco, CA
1990 Open Studios, San Francisco, CA
1986 Gale Laurence Studio-Workshop, San Francisco, CA
1985 Neiman-Marcus Gallery, San Francisco, CA
Columbus College of Art and Design, Columbus, OH
Ohio State University, Columbus, OH
Day Studio – Workshop, San Francisco, CA
International Design Award
The Good Design Award
Artist's Work


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By litofJuly 11, 2018 No Comments
CHRISTINE PELISEK
A Los Angeles woman who once ran a successful PR company catering to clients like Ford and Payless ShoeSource was found murdered at a posh resort in South Africa – and her Guatemalan boyfriend has been arrested and charged with premeditated murder in connection with her death.
The July 29 killing of Gabriela Alban, 39, at the Camps Bay Retreat Boutique Hotel in Cape Town, where rooms cost up to $10,000 a night, has left her friends shocked.
"It is devastating," friend Aida Phillips tells PEOPLE. "I just found out she was dating this guy a few months ago. She was super happy."
"She was the sweetest, most caring woman," says another friend, Monica Prieto. "One of those women that like to nurture others, which led her to some toxic relationships with troubled men."
"It was a really big shock," says another friend, Ivette Serrano. "Nobody really knows what happened."
An official cause of death has not yet been released. Her 41-year-old boyfriend, Diego Dougherty Novella, who was reportedly a member of a powerful Central American family, was arrested after her body was found.
Alban was "very focused on her company and career, her dogs and family," Prieto says. "Smart women sometimes fall for the wrong men."
Prieto adds that Alban was "absolutely loved" in the PR industry. "The media world is brutal, lots of envy, backstabbing, however I never met one person, not one, that did not love her. Not one."
Alban, who grew up in Los Angeles, attended Pepperdine University in Malibu. She started Conexién, a PR company focused on the Hispanic media market with Martén Llorens, a well-known author and a renowned fashion and lifestyle expert, in 2002.
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It won't be just a regular art opening when the Hollywood Academy of Arts & Science (HAAS) holds its "Through the Eyes of Young Artists" exhibition reception at the Young Circle ArtsPark Gallery on Thursday April 5 from 6 to 7:30 pm. It will be more of a production experience rather than a standard art exhibition.
Under the guidance of art teacher Hilary Fine, this is the school art department's 12th Annual Art Exhibition at a professional gallery. It'll be filled with special guests too, like Mayor Josh Levy and Vice Mayor Deborah Case of Hollywood, dignitaries from the Art & Culture Center/Hollywood, Young at Art Museum, Broward Cultural Division – Art in Education Committee, Broward Schools and CSUSA and other educational divisions. They're also hoping to have the Holocaust Memorial of Miami Beach and FIU Museum represented.
But mostly it's special because it was put together by students of HAAS at a highly professional level. Hollywood Academy of Arts & Science is an elementary and middle school which specializes in an "enhanced traditional curriculum with a focus on the arts and science." Students named the exhibition "Through the Eyes of Young Artists" because they read the Holocaust book "Entombed" by Bernard Mayer and from there did a special photo and video shoot as they "saw" human rights. The photos and the video they produced is for human rights based on the Holocaust and survivor Fred Mulbauer's experience. Archival photos were from the Singer/Friedman family albums and current photos and videos were taken at the Holocaust Memorial, Miami Beach. Students titled their video "Never Again", in respect of the Jewish mantra regarding the Holocaust, but tragically their 'Never Again" also became the motto for the recent high school shooting in Parkland. So, the HAAS students dedicated their film to these students and to worldwide human rights. This film will be shown at the ArtsPark Exhibition Reception.
This has been an impressive year for HAAS's students. They were winners both nationally and in all top categories for the entire State of Florida with the EPA's Radon Awareness Poster, and it is expected that representatives of the Florida Department of Health and the American Lung Association will be at the event presenting students with crystal awards for art and writing. Young At Art Museum will also present this year's winners from the Art In The Capitol Awards and the Scholastic Awards as well.
Students will perform at this event, with art masters including visual artist Daniel Pontet who will be painting with his feet! Documentary producer Cathleen Dean, and Grammy musicians Federico Britos and Thomas McWilliams will also be there. Also performing will be guest musicians Jeffrey Lee, Diego Fuica, Jorge Alfano, Fernando Moraes, and guest singer Brenton Williams.


Main Title Age-Dependent Lung Dosimetry of Radon Progeny.
CORP Author Health Effects Research Lab., Research Triangle Park, NC. ;Northrop Services, Inc./Environmental Sciences, Research Triangle Park, NC. ;Duke Univ. Medical Center, Durham, NC.
Abstract Two morphometric models differing in the tracheobronchial region, were compared in the present paper: Model 1 is based on the adult morphology of Weibel, assuming that all bronchial airways grow in equal proportion; while Model 2 adopts the adult structure proposed by Yeh and Schum, using measured airway dimensions in the right upper lobe as a function of age. Tidal volume and respiratory frequency also vary with age: while the breathing frequency decreases with rising age, tidal volume increases. Radiation doses in each bronchial airway generation were computed for the deep lying basal cells as well as for the more uniformly distributed serous (SMGS) cells, which are currently assumed to be the progenitor cells for bronchial carcinomas. Radiation doses to both target cells were significantly higher in the newborn than in the adult, for all simulated breathing patterns, showing the highest relative increase in upper bronchial airways. Comparing both tracheobronchial growth models, Model 1 predicts higher doses at early ages, but produced lower doses in the adult lung.
Supplementary Notes Prepared in cooperation with Northrop Services, Inc./Environmental Sciences, Research Triangle Park, NC., and Duke Univ. Medical Center, Durham, NC.


In new interviews, Conny Plank's collaborators – including Brian Eno and Holger Czukay – remember the radical music producer.
What do we mean when we call someone, or something, pretentious?
It's Not Only Rock 'n' Roll, Baby!


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Het kwartier van Frederik Hendrik of Kwartier van de Prins in Vught was een Staats legerkamp dat gebouwd werd tijdens het Beleg van 's-Hertogenbosch in 1629. Van de vijf kwartieren die rond de stad gebouwd werden tijdens de belegering was deze verreweg het grootste. In dit kwartier waren ongeveer 20.000 militairen gestationeerd.

In het oosten werd dit kwartier begrensd door de huidige Pestdijk. De zuidgrens van het kwartier lag waar nu Afslag Vught is op de A2. De noordgrens is iets ten zuiden van de A65, terwijl de westgrens iets ten westen van de Spoorlijn Utrecht - Boxtel lag.

Frederik Hendrik van Oranje had de leiding over het kwartier en verbleef op het Heymshuis. Dit huis is later omgedoopt tot het Kasteel Maurick. Dit kasteel viel net buiten het kwartier. Het had wel een gracht, zodat het beschermd was tegen eventuele aanvallen. De ingang van het kasteel lag wel in het beschermde kwartier.

Na het beleg van 's-Hertogenbosch heeft de Prins zijn hoge gasten in Vught ontvangen. Een paar dagen na het beleg waren zij ook te gast bij de eerste gereformeerde dienst in de Sint-Janskathedraal. De hoge gasten mochten genieten van de uittocht van de religieuzen uit 's-Hertogenbosch die hun heil in het zuiden zochten. De gasten hadden hun verblijf in kasteel Oud-Herlaar.

Zie ook
 Lijst van Kwartieren bij het Beleg van 's-Hertogenbosch

Beleg van 's-Hertogenbosch
Geschiedenis van Vught

redpajama_set_name: RedPajamaWikipedia


I work from private counselling room at my home. On your first visit, I will start by explaining a little more about what counselling involves, what you can expect from me and what I will expect from you.
Before starting counselling, check out that counselling is really what you want?
Sometimes people come for counselling and find it a rather puzzling experience and not what they were expecting. The aim of counselling is to help you to understand yourself and, if you so choose, make changes in your thoughts and behaviours so that your difficulties become more manageable.
When you go to a medical professional like a doctor or dentist and tell them your symptoms, they generally make a diagnosis and prescribe medication or recommend changes to your lifestyle. Occasionally people come to counselling with the expectation that counselling works in the same way. If you come with this in mind, you may initially feel disappointed.
I work from a fundamental belief that you are the expert on you and my job is to help you to access your own inner wisdom and strength to solve your own difficulties. This is ultimately far more powerful and life-changing than if I gave you advice or told you what to do. Counselling sometimes feels like hard work and part of the process may involve experiencing difficult and painful emotions along the way – you could feel worse before you feel better. It may be worth bearing in mind that, because it feels hard, some people give up, in my experience, around the second or third session and this may be the time when it is more important than ever to keep going.
Listen to you with interest, concern and respect.
Tell you what I notice, both verbally and non-verbally in your body language.
Encourage you to focus on your reactions to other people's behaviour rather than on their behaviour.
Encourage you to express your feelings, including feelings about the counselling.
Point out how you may be contributing to your own difficulties and help you to discover new ways of changing.
Discuss ending the counselling when it seems appropriate.
Give you clear cut solutions to your difficulties.
Have any relationship with you other than a counselling one.
Be a permanent support or friend to you.
Talk about myself unless I believe it is of particular value or relevance to you.
Talk about you to anyone else without your permission ( except in very exceptional circumstances which will be discussed with you).
I will ask you a few questions about yourself and give you the opportunity to discuss any worries or ask me any questions you may have. If you are willing, we will then start to explore the issues you wish to address. Towards the end of the session, we will discuss whether or not you feel you would like to continue our work together.
I am often asked how many sessions you will need. This varies from person to person. Some people can get what they need from two or three sessions while other people may want to go deeper or may require many more sessions to get to the bottom of what is causing them discomfort and to make life enhancing changes. The choice is always yours.
Sessions are one hour long and my recommendation to get the best out of counselling, is to make them weekly, but, where this is not possible,they can be as frequently as you choose.
The fees are £45 – £60 per session for individuals and £67 – £80 for one and a half hour sessions for couples. Please note that missed appointments without 24 hours notice must be paid for at half the normal rate.
Contact me today to discuss how counselling can help you.


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At Values Voter Summit, 2016 hopefuls pitch social conservatives
At the Values Voter Summit, Republican hopefuls will attempt to appease the far-right without getting mired in social issues that could alienate them in 2016.
Who's missing from Value Voters Summit?
Sept. 25, 2014, 6:15 PM UTC / Updated Sept. 26, 2014, 11:15 AM UTC
By Jane Timm
With the midterms weeks away and the 2016 cycle kicking off in earnest shortly thereafter, the GOP is trying to figure out how to win elections while keeping their far-right base happy. The Values Voter Summit in Washington, D.C., this weekend will present an ideal opportunity for Republican presidential hopefuls — including Sens. Rand Paul and Ted Cruz, Gov. Bobby Jindal and former Sen. Rick Santorum — to impress social conservatives who would make up a crucial portion of their base voters.
Republicans have often tripped up in recent elections when talking about social issues — think Todd Akin and Richard Mourdock on rape and dozens of Republican comments on women's issues — so how 2016 hopefuls navigate the Values Voter Summit may signal how the party and candidates will handle explosive issues in the upcoming presidential election.
"What annual event has the Left so worried that they are spending tens of thousands in advertising dollars to stop? You guessed it; the Values Voters Summit."'
The focus of several of the weekend's events make it clear that the GOP is well aware of the hurdles that lay ahead.
More sessions than in the past two years advertise a focus on winning the election. "How Conservatives Can Win With Millennials and Women" and "Stop Losing Elections: It's Time to Reclaim Blue Collar Conservatives" push the party's desire to win women and young folks, as well as build the kind of grassroots muscle President Barack Obama utilized to great success in 2008.
Marriage for same-sex couples promises to be a hot issue in particular. As public opinion has moved quickly to the left on marriage equality over the last few years, Republicans have struggled to appease their far-right members while not deterring moderates and independents.
It's a conundrum that is already playing out: A coalition of civil rights groups wrote a letter to the Republican National Committee on Wednesday, asking them to distance themselves from the Values Voter Summit. They also asked elected members not to attend because many of the participating groups have histories of expressing anti-gay rhetoric.
"Today, Chairman Priebus, we ask that you … call upon Republican officials to disassociate themselves from the groups behind the upcoming Values Voter Summit," the groups wrote in a letter published in The Washington Post. "The reason is simple: These groups engage in repeated, groundless demonization of LGBT people — portraying them as sick, vile, incestuous, violent, perverted, and a danger to the nation."
The letter slams top executives at the Family Research Council (FRC), which hosts the summit, and two of the event's sponsors, American Family Association and Liberty Counsel, for making anti-gay remarks.
The RNC did not return requests for comment on the letter, but FRC President Tony Perkins isn't shying away from the issue. He has been criticized for calling pedophilia a "homosexual problem" and the It Gets Better campaign "disgusting" and an effort to "recruit" kids to the gay "lifestyle." Perkins defended his remarks in a lengthy response.
"What annual event has the Left so worried that they are spending tens of thousands in advertising dollars to stop? You guessed it; the Values Voters Summit," he wrote. "The Left is clearly discouraged by the effectiveness of the Values Voter Summit and is willing to fork over a lot of money to stifle speech they disagree with."
A panel session titled "Marriage in America: The Road Ahead Panel" will take place in front of a full-house during Friday's three-hour afternoon plenary session. A breakout session on Saturday is titled "The Future of Marriage: To the Supreme Court and Beyond."
Birth control and abortion — longstanding topics for the summit — promise to remain a top issue in both this weekend's conference and in the 2016 race, as Democrats and Republicans feud furiously over what conservatives say is a matter of religious freedom and liberals say is an attempt to overrule women's reproductive rights.
Three breakout sessions, "Pro-Life Battleground 2014," "Moral Decline Causes Big Government" and "Sexuality in the 'Hook-Up' Culture," will try to reconcile far-right views with Americans' overwhelming acceptance of birth control and highly polarized views on abortion.
In addition to presidential hopefuls, the usual conservative firebrands are all expected to take part in the event. Former Gov. Sarah Palin, Rep. Michele Bachmann, former Gov. Mike Huckabee, as well as the Duggar Family, Duck Dynasty's Alan Robertson, Erick Erickson, and Glenn Beck, are all ready to take the stage this weekend.


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Always ... Patsy Cline
The true story of country music legend Patsy Cline's remarkable friendship with devoted fan Louise Seger.
Created and originally directed by Ted Swindley – based on a true story.
Licensed by the family and estate of Patsy Cline. All rights reserved.
Musical clearances secured and used by permission.
Original cast album available on MCA records.
The true story of country music legend Patsy Cline's remarkable friendship with devoted fan Louise Seger, filled with 24 unforgettable hits, including "Crazy," "I Fall to Pieces," "Sweet Dreams" and "Walkin' After Midnight."
Tickets are $27 for adults and $20 for students of any age, with a $2 discount for seniors on the Sunday matinees.


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New-games
City of Titans reaches its Kickstarter goal after five days
After launching its Kickstarter crowdfunding campaign on October 2nd, City of Titans has met and ex...
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The Phoenix Project dubs itself City of Titans and launches on Kickstarter
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By E. Lefebvre, 10.02.13
Vin Diesel describes upcoming Chronicles of Riddick game as being an MMO
If you can't get enough of media wherein Vin Diesel is a man who can see in the dark... well, he's...
The Daily Grind: How long do you need to play a game before you get a fair impression?
First impressions are tricky things. Play a game for a little while and you'll get an idea of wha...
Fez creator Phil Fish working on two new games
Phil Fish, the man behind the mutton-chops and this awesome indie game you probably haven't heard o...
By J. Conditt, 07.02.12
The Daily Grind: What game have you waited past launch to play?
I'd be lying if I said I'm not interested in The Secret World. What little time I got with the ga...
Dark Legends goes live for Android and Chrome
So you like vampires? It's kind of a thing these days. Maybe you like vampires so much that you r...
SteelWar Online kicks off alpha testing
Are there enough robots in your life? If your answer is "yes," you're most likely not concerned a...
Goblinworks announces Pathfinder Online
For pen-and-paper RPG players, Pathfinder remains a perennial favorite game setting. Those play...
By M. Daniel, 11.22.11
The Daily Grind: Where would you take an MMO vacation?
There are places in the world that are nice as a bit of a change from your usual living space, so...


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The Taming of the Duke
Book 3 in the Essex Sisters series
Imogen, Lady Maitland, has decided to dance on the wild side. After all, she's in the delicious position of being able to take a lover. A discreet male who knows just when to leave in the morning.
But Lady Maitland is still under the watchful eye of her former guardian, the wildly untamed Rafe, the Duke of Holbrook. He believes she is still in need of a "watchdog." She laughs at the idea that someone so insufferably lazy and devoted to drink can demand that she behave with propriety.
It's Rafe's long-lost brother, a man who looks precisely like the duke but with none of his degenerate edge, who interests Imogen. To Imogen, he's the shadow duke…the man who really should hold the title.
But when Imogen agrees to accompany Gabe to a masquerade… whose masked eyes watch her with that intense look of desire? Who exactly is she dancing with? The duke or the shadow duke?
Rafe…or Gabe?
Bookshelf:
See all ordering options, including international and audio
"James's considerable talents for clever prose and tight, breezy plotting are on full display, promising a perennial delight in each coming adventure of the Essex sisters."
~ Publishers Weekly
Romantic Times BOOKClub nominated The Taming of the Duke for "Historical Romance of the Year"!
#18 on USA Today bestseller list.
#17 on the New York Times extended list.
#10 on Publishers Weekly bestseller list.
RITA (romance's top prize) Finalist for Best Short Historical of 2006.
Jump to: Excerpt • The Inside Take • Mea Culpa • Series Reading Order
Stepback for The Taming of the Duke
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Essex Sisters Series Crossword Puzzle
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Audio Excerpt for The Taming of the Duke
This Audio Excerpt for The Taming of the Duke, narrated by Susan Duerden, begins where the book begins, with Chapter One. Enjoy!
Essex Sisters Collectible Card
Eloisa made up these gorgeous collectible cards for readers to celebrate the Essex Sisters. With Tess and Annabel on one side and Imogen and Josie on the other, this 5x7 can be yours. And this isn't the only gorgeous card to be had!
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Inside The Taming of the Duke
Warning! In describing relations between characters, I may wreck a book for you by making it clear who someone marries, or the outcome of a book. Please do not read about The Inside Take if you're wary of knowing who is paired with whom!
Josie distorts Shakespeare on page 113, as Annabel points out. She's quoting from Twelfth Night. As a Shakespeare professor, I often teach those lines in context, but I actually first encountered them in the delicious novel by Georgette Heyer, Venetia. If you haven't read Venetia, run to the nearest store! It's her sexiest, to my mind.
Check out more like this from Eloisa in the Inside Take →
With the Help of Jane Austen…
A short piece where Eloisa acknowledges a Mansfield Park influence, penned while Eloisa was writing The Taming of the Duke, before she even had a title for the book!
Read this essay in full →
The Essex Sisters Family Tree
Interested in knowing how many children the Essex sisters had? Eloisa has a beautiful Essex Family Tree for you to see.
Dreaming Up Gabriel
For the release of The Taming of the Duke, Eloisa asked readers, who had first met Rafe in Much Ado About You, what they thought Rafe's illegitimate half-brother would be like and why he was in Taming. The responses were amazingly passionate, interesting and fun to read. Eloisa was tremendously honored at the time, imagination and affection invested in her characters. Click through for four of her favorite submissions.
Mea Culpa, The Taming of the Duke
Dawa pointed out that when Imogen and Rafe are at Christobel's performance, before Christobel actually performs, the innkeeper tells a patron to put his sword away. As Dawa noted, swords were not common Regency accoutrements. Dawa thought perhaps the whole question of the sword was a euphemism for the patron's manly parts…I wish I'd thought of that! As it is, it's just a mistake. I must have been having a medieval moment.
For more found errors in this book →
"Deception, hidden identity, and secrets combine in an entertaining romp…"
~ Library Journal
"James continues to provide her readers with smart, passionate, uplifting romances that make her a must-read."
~ The Oakland Press
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From Chapter One
In which the Curiosities of Courtship are Reviewed
Ardmore Castle, Scotland
"I wish I were a queen," Miss Josephine Essex said to two of her elder sisters. "I would simply command an appropriate man to marry me by special license."
"What if he refused?" Imogen, sometimes known as Lady Maitland, asked.
"I'd remove his head from his body," Josie said with dignity.
"Given that men make slim use of their heads," Annabel, the Countess of Ardmore said, "you don't have to threaten decapitation; simply allow the fellow to believe that he made up his own mind about marriage." She was tucked in Imogen's bed and appeared little more than a tousle of curls peeking from under the bedcovers.
"That is precisely the kind of advice I need." Josie snapped open a small book and poised her quill. "I am making a study of the skills required to succeed in the marriage market and since you two are both married, you are my primary sources of information."
"I'm a widow," Imogen said. "I know nothing of the marriage market." She was sorting silk stockings and didn't even look up from the dressing table.
"One should be able to dance," Annabel noted. "You really must practice harder, Josie. You were stomping on Mayne's toes the other night."
"I need better advice than that," Josie said to her. "You are the only one of us to have actually gone on the season, and you married into a title. You do remember that I'm to have a season next year, don't you?"
Annabel opened one eye. "Only because you mention it every other minute. Lord, but I'm sleepy!"
"I've heard that marriage rots the brain," her younger sister told her cheerfully.
"In that case, I wonder that you're so interested in it."
Josie ignored that unhelpful comment. "There's more to gaining a husband than not tripping over his feet while waltzing. I want to understand the challenges beforehand. I can't rely on beauty, the way the two of you did."
"That's ridiculous. You are lovely," Annabel said.
"I was in London for the better part of April," Imogen said, "and I saw plenty of young ladies in your situation, Josie. It seemed to me that the primary requirement for a debutante is a smirk. An innocent simper," she clarified.
"Smirk," Josie noted in her book.
"And listen to everything your suitor says as if God himself is speaking. Of course, sometimes it's difficult to stay awake."
"Men can be very boring," Annabel agreed. "They have such a penchant for discussing themselves. You have to learn to endure, which is not one of your best qualities, Josie."
"To this point, you have shown no ability to suffer fools gladly," Imogen said. "Fools have the deepest pockets. It's a proven fact that lack of brains and a large estate go hand-in-hand."
Josie had been writing busily in her book but she looked up at this. "So I smirk at the fool as he talks about himself? Essentially, toe-curling boredom buys a spouse?"
"I think Imogen is overemphasizing the smirk," Annabel put in. "There are moments in courtship that can be rather interesting. In my view, for example, a prospective groom might prefer engaging in a mildly scandalous activity to a mutual smirk."
"Annabel has a point. I suppose you might occasionally engage in an imprudent act,," Imogen said, "but only you found yourself in the company of a truly engaging young man."
"That's a bit steep coming from you," Josie said. "You devoted yourself to outrageous efforts from the very moment you saw Draven Maitland. Remember how he kissed you, after you arranged to fall out of a tree at his feet?"
Imogen's hands stilled for a moment. "Of course I do. It was spring and the apple tree was in bloom."
"And then you fell off a horse, and finally you fell into marriage. Your example seems to go against the model of the innocent simper," Josie said. "I intend to be practical about this business, and I have no particular disinclination to creating a scandal, if that is the most efficacious route to marriage."
"My foolishness is nothing to emulate," Imogen said, returning to her task and folding two pale blue stockings together. "You would do better to find a husband by a more conventional means."
Josie made a note in her book. "Employ an innocent look, no matter how imprudent one's private conduct may be. It sounds like that gentleman thief who is always getting described in the Times. One moment he appears as a fine gentleman and then with a twist of a dishclout, he's transformed into a beggar."
"In fact, the reverse of Imogen's style," Annabel pointed out, a hint of mischief in her tone. "Since Imogen specializes in appearing debauched, no matter how innocent her private activities may be. According to Griselda, all of London now believes you are carrying on an illicit amour with Mayne, whereas in truth the man has achieved slightly more intimacy than a footman."
"Every woman should have an occupation," Imogen said. "Mine is to provide interest to the old biddies." She tossed a few stockings over her shoulder. They gently drifted to the bed and fell on Annabel's legs.
"Well, as to that," Josie said thoughtfully, "You seem to be slightly behind the times, Annabel."
"She's more than behind the times. She's utterly out of style," Imogen said. "Last night she was flirting with her husband at supper. That kind of behavior is beyond unfashionable; it's practically indecent. No one is supposed to pay attention to their spouse in public. Or," she added, "in private either."
Annabel grinned and said nothing.
"I saw Ardmore kissing you in the breakfast parlor yesterday," Josie remarked. "Your husband has lost his head, which suggests that you should be able to help me. You must have better suggestions than improving my dancing."
"I hardly planned my course of action in a thoughtful manner," Annabel pointed out. "I was desperately unhappy with this marriage, remember? The only reason you two are in Scotland is to save me from my terrible fate."
"A slight miscalculation on our parts," Imogen said. "I could be in London at this very moment, surveying the dubious temptations of men interested only in my estate."
Annabel snorted. Imogen's hair was a glossy black, and smooth as a raven's feather unless she decided to curl it–whereupon it kept a perfect ringlet. Her eyes were wide apart and framed by brows in a flaring arch. Her mouth was just as wide and made for laughing, even though she'd done precious little of that since her husband died the previous year.
"There are more than enough besotted men throughout London to catalogue your features for you," Josie said impatiently. "The really interesting point here is that Annabel doesn't seem to realize that you have been making a concerted effort to woo Mayne into far more intimate activities than are generally enjoyed by footmen."
She ducked as a stocking flew over her head.
"Really, Imogen?" Annabel asked.
"I told you in London that I intended to take a cicisbeo," Imogen said with a snap in her voice.
"But I thought you meant merely a gentleman escort, not a cher ami."
"It has been my distinct impression," Josie said, "that Imogen has demanded that Mayne prove his reputation as a Lothario is not exaggerated."
Imogen's scowl should have silenced Josie on the spot.
"And I regret to report," Josie continued, apparently unruffled by her sister's fiercest glare, "that to all appearances Mayne refused the challenge and kept his virtue intact."
"How surprising," Annabel exclaimed, pushing herself up on the pillows and looking altogether more awake. "I was under the impression that he had no virtue."
"To the contrary," Josie said. "No matter how Imogen batted her eyelashes at him during the trip to Scotland, he kept to his own bedchamber."
"Josie," Annabel said. "You should not speak of bedchambers – no, or even contemplate such behavior. You sound positively hurly-burly. It would be disastrous for your marriage prospects if anyone heard you talking in this fashion."
"Don't be a goose, Annabel," Josie said unrepentantly. "It's not as if I intend to imitate that behavior. I know the difference between what's allowed a widow and an unmarried girl."
The color was rising in Imogen's cheeks under Annabel's interested gaze.
"I suppose the crucial point is not the position for which you considered Mayne," Annabel said to her, "but the position he has agreed to take."
"There's the rub," Josie said. "He managed to get all the way to Scotland with the unblemished virtue of a – a debutante."
Imogen threw a petticoat over her head but Josie just talked right through the frail lace. "There she was, batting her eyelashes, as I said –"
"I never bat my eyelashes!" Imogen put in.
"She batted them," Josie repeated, "and spent a great deal of time trying to convince Mayne that she was besotted with his blue eyes."
Imogen threw a whole heap of corsets on top of her little sister. "Hurly-burly is too good a phrase for you."
Annabel was looking fascinated. "Mayne is very handsome. I can certainly sympathize with the impulse."
"No, I–"
"I never said she was truly struck by his eyes," Josie said from under a heap of linens.
"Yes, you did–"
"No." Josie pulled the cloth off her head. "To call a spade a spade, Imogen, you may have tried to turn the earl into your cher ami. But you never, ever looked at him with that fascinated expression with which you used to watch Draven." She turned to Annabel. "So I would deduce that Imogen was not entranced by his eyes. Perhaps by an arm, a leg, or some other…part of his anatomy?"
Annabel frowned at her. "Josie, you wanted advice about the marriage market; I have a serious piece of counsel. Do not indicate that you have the slightest notion what a cher ami is. And never make a joke about parts of the male anatomy that you do not feel comfortable naming."
"I have no reluctance to name–" Josie began readily, but Annabel cut her off.
"That's enough! I don't wish for any anatomy lessons from you."
"If Imogen wishes to forgo a life of celibacy, am I supposed to pretend her efforts don't exist?" Josie said plaintively. "It's not as if people ignored the fact that Mayne took up with the sister of a woman he jilted. His reputation was ruined the first time he danced with Imogen, given his behavior toward Tess."
"Nonsense," Imogen said, finally breaking into the conversation. "Mayne hasn't had a reputation in years. I had nothing to do with it. Any reputation he had left was shattered by his ill-mannered act of jilting Tess in the first place."
"I suppose your disrespectful tone reflects pique," Josie said. "It must be highly annoying to be refused by a man who has so generously spread his attentions around the ton."
"Mayne is an idler, and I have no wish to engage in any sort of intimacies with him."
"Excellent," Josie said heartily. "I shall follow your lead and thoroughly dislike all gentlemen who don't instantly succumb to my charms. Of course, given my girth, I just ruled out most of the available gentlemen in London."
"You are an extremely annoying person," Imogen said. "That alone may keep you unmarried."
"Could we return to that particular challenge for a moment?" Josie asked. "I am serious: I need to know how to attain a proposal of marriage, preferably within a few weeks of the season opening."
Annabel shook her head. "None of us have married in a conventional fashion, Josie. Tess married Felton only after Mayne jilted her. I married Ewan because I had to after that scandal broke."
"I chose my husband in the normal way," Imogen said, "and Lord knows that didn't work out very well."
"It would have, if Draven had lived," Annabel pointed out. "You can hardly blame his death on your elopement."
"It's very annoying," Josie muttered. "How am I to do this? How am I to find a husband?"
"I'll be there," Imogen said consolingly. "And Griselda has already agreed to be your chaperone. You know that she knows all the ins and outs of the ton."
"She told me that her father arranged her marriage," Josie said, looking uncharacteristically helpless. "We don't have a father."
"We have Rafe," Annabel said.
Imogen shrugged. "When he's sober."
"You're just cross because he didn't like it when you took up with Mayne," Josie said.
"Rafe doesn't seem to understand that my marriage freed him from the need to act as my guardian."
"But you were only married a few weeks," Annabel said gently. "I can see why Rafe still feels responsible for your welfare."
"I have agreed to return to his house, haven't I? I had planned to set up my own establishment but instead I'll be living with Rafe and trundling around with Griselda as my chaperone. I'm a widow. Why do I need a chaperone?"
"You seem to have left me out of that delectable picture," Josie said. "So, on that note, Annabel, would you consider allowing me to stay with you for the winter? Apparently, dancing is the only practical skill I need to polish before next spring, and I'm sure there must be a tutor somewhere in Scotland. It's so lovely to be back in the highlands; I loathe the thought of returning to the south."
"This country is damp, cold, and winter's coming," Imogen pointed out.
"I would love to have you stay with me, Josie," Annabel said.
"Will you be quite all right if I don't join the two of you?" Imogen asked. "And I doubt very much that Griselda would like to winter in the highlands."
Annabel had snuggled back down in the covers. "Of course I will be. I'm married." There was a little smile in her eyes.
"I thought you might be nervous about the babe," Imogen said.
Josie gaped, and Annabel sat upright again. "How did you know?"
Imogen laughed. "For goodness' sake, Annabel, you generally retire to bed for two days when your flux appears. We've been here since the end of May, and now it's August. You've spent no time whatsoever groaning about the unfairness of a female's condition. In fact, you look utterly pleased to be female."
"Oh, I am," Annabel said, the smile in her eyes growing.
"A baby!" Josie said. "When will it be born?"
"Not for ages yet," Annabel said. "Likely in January or February."
"I needn't return to England for the season until the end of March!"
"Your company would make me very happy," Annabel said, grinning at her little sister.
"Are you certain that you wouldn't like me to stay as well?" Imogen asked, feeling a tremendous reluctance to do so. It wasn't that she was bitter.
A surge of honesty corrected the thought. Of course she was bitter. Two of her sisters were happily married, and now Annabel was having a child. The memories of her two-week long marriage with Draven were a cold comfort.
"I would love it if you wished to stay," Annabel said, holding out her hand to Imogen. "But I think you should go to London and drive the gentlemen mad by acting like the light widow you so emphatically are not."
"The season is over," Imogen said. "Griselda and I won't go to London. We'll stay with Rafe in the country."
"And Mayne?" Annabel asked.
Imogen shook her head. "A passing fancy," she said. "Luckily he was shrewd enough to see that before I did."
Annabel squeezed her hand.
"Perhaps over the winter you could occupy yourself by making me a list of appropriate parti," Josie suggested. "I don't want to waste my smirks on a man who is lacking in the necessary prerequisites. So many people float through Rafe's house that you are sure to hear all the gossip."
"And those prerequisites are?" Imogen asked, amused.
"I've made a list, garnered from reading every single romantic novels published by Minerva Press." Josie consulted her book.
"An estate is necessary, and a title would be nice. He should be able to read, but not too passionately. Unless he likes novels. And I don't want him to be overly fashionable."
"Don't you have any physical requirements?" Annabel asked.
Josie shrugged. "I would prefer that my husband be taller than I am. Since I am very short, I foresee no difficulty there." She frowned. "Why are you both laughing? There's nothing ludicrous about my ambitions. My list is likely very close to yours, Imogen."
"My what?"
"Your list," Josie said. "Every woman has a list, even if they don't write it down."
"I don't," Imogen said, her lips tight.
"It's been almost a year since Draven died," Josie said, as usual wading in where any hearty soul would hesitate. "You'll have to think of marriage again at some point. You don't want to wither into nothing more than an aunt to Annabel's children."
She caught Imogen's sharp gaze but missed Annabel's. "Well, for goodness' sake, you certainly found it acceptable to contemplate intimacies with Mayne. From what I understand, marriage is merely a regularizing of that sort of relation."
"Josie!" Annabel moaned.
Imogen started laughing again. "Now there's a cold-eyed look at matrimony."
"Your list and mine are likely the same," Josie said. "You simply haven't clarified your demands and I have."
"Tell me again what qualities I am looking for?"
"An estate. A title, if possible. Intelligence, but not to an uncomfortable degree. The same goes with fashion. One would dislike being married to a man who always looked better than oneself."
"I think you should be a tad more specific," Imogen said. "Our own guardian would fit every category you mentioned: Rafe has an estate, a title, sufficient height, no fashion sense whatsoever, and a reasonable amount of intelligence, if slightly pickled."
"You're right," Josie said. "I shall add an age limitation." She sat down, quill poised. "Shall I cut them off at thirty, or twenty-five?"
"My point was more that Rafe is a drunk," Imogen said. "Your list overlooks every important characteristic that one would want in a husband."
"I suppose you are talking about steadiness of character," Josie said. "Rafe actually has that. He's attractive too, very. He's just too old for me."
Imogen suddenly noticed that both Annabel and Josie were watching her. "He's far too old and too drunk for me," she said quickly.
"You are over twenty-one," Josie said with her customary crushing truthfulness. "And you are a widow. I think it is an entirely inappropriate match, as far as age is concerned."
"Rafe may not be perfect for you, darling," Annabel said, taking her hand. "But someone will be."
A little wry smile turned the corner of Imogen's lips. "In truth," she said, "I'm one of those people who fall in love only once, Annabel."
"If we could all plan the moment when we would fall in love as easily as I am making this list," Josie said, "the world would be an altogether more tolerable place. For one thing, I would make certain to fall in love only after a man had sworn undying love."
"Good luck," Imogen said, hearing the disconsolate ring in her own voice.
Annabel squeezed her hand again.
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The Taming of the Duke is Book 3 in the Essex Sisters series.
The full series reading order is as follows:
Book 1: Much Ado About You
Book 2: Kiss Me, Annabel
Book 3: The Taming of the Duke
Book 4: Pleasure for Pleasure
Companion Guide: Essex Sisters Companion Guide
Novella: A Gentleman Never Tells


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Posts tagged 'Six Nations'
Rogue Warriors Live Up To Stereotype
November 16, 2009, 7:30 am by Bob Goulais
Smoke engulfs a minivan as it is overturned during 2006 land-claim protests near disputed Caledonia, Ont., site. Neil Dring Photo/Grand River Sachem
"Whoa! Gee, those nadways sure get violent, i'nit?" "Oh yeah. Those warriors sure mean business, cuz."
Yes, even their fellow indigenous people think the Haudenosaunee play the occupation and resistance card a little too aggressively.
They came out, guns blazing, during the conflict at Kanesatake, the biggest indigenous flashpoint in the last 30 years. During the Oka Crisis of 1990, the Mohawk Warrior Society took up arms against police and the Canadian Forces. In 2006, they began occupation and control over a tract of residential development land in Caledonia. In 2007, they blocked Canada's biggest freeway and passenger railway corridor during First Nations' national day of action.
"They" are members of the Mohawk warrior society. They represent indigenous militancy in the 21 century. They have gotten far more headlines in recent years than the American Indian Movement, who immortalized indigenous resistance in the 1970s.
Unfortunately, for First Nations advocates and even the non-violent indigenous resistance, the Mohawk warrior society is a lasting example of militant truism and living up to contemporary stereotypes. First Nations activists do wave scary flags, wear masks, fatigues and balaclavas in the nice weather. We set things on fire. We beat up people. We are lawless terrorists.
I strongly object your honour! We shouldn't be painted with the same camouflage brush.
This week, a couple from Caledonia is taking the Ontario Provincial Police to court over damages they have incurred as a result of the alleged inaction of the police during the occupation of the Douglas Creek Estates development in 2006. Through the evidence and testimony in court, we are seeing many examples of First Nations-led violence.
By all accounts it was a war zone. Cars were overturned. Property was burned. Citizens and police were brutally assaulted. A hydro sub-station was destroyed.
It's hard to say that First Nations activists are not terrorists, when quite clearly, that's exactly what the organizers wanted to get across.
Or was it?
As far as I know, violence was never condoned by the traditional Haudenosaunee confederacy. Sure, they are tough negotiators, tacticians and they too, mean business – but the Chiefs and Clan Mothers did not mean to hurt others.
The trouble began when the OPP went in and made arrests. There were some skirmishes but the first wave of protesters retreated. But through the wonders of technology, more than a few protesters made urgent phone calls to friends and family. Within an hour, reinforcements from Six Nations arrived and they weren't too happy. They were ready to kick ass.
Violence erupted and the OPP where overwhelmed. The OPP retreated.
I think the OPP made the right decision. If they pressed the matter, my guess is that an all-out civil war could have taken place. Guns would have been brought in, and we'd have Kanesatake all over again.
I would hasten to argue, that the lawlessness and violence brought about during the Caledonia occupation was instigated by hooligans from the Six Nations community and elsewhere. There was a significant lack of discipline within the ranks of the organizers and the warrior society in not dealing with these rogue warriors. Believe me, the leadership did not have any control over the Caledonia occupation during this time.
It is these rogue warriors that live up to the violent stereotype.
But why would something get so out of hand? Why do the Haudenosaunee resist so aggressively?
We have to walk a mile in their moccasins, or whatever footwear the Haudenosaunee use.
The Haudenasaunee are also know as the six nations of the Iroquois confederacy. The six nations are the Mohawk, Oneida, Seneca, Onondaga, Cayuga and the Tuscarora.
Traditionally and in their history, they are a powerful warrior society. So powerful and strong, that they made war upon their neighbours with great vigour, ruthlessness and lust for victory. They nearly destroyed my people, the Nipissing, during the 1700s.
However, at one time in their history, this strength and vigour was turned into itself and civil war waged amongst each other. Through their Great Peace, they literally "buried the hatchet" under the Great Tree of Peace. This led to a new era of unity and harmony and formed the basis of their Haudenosaunee Confederacy which is still in place today.
Part of their traditional territory in Ontario is known as the Haldimand Tract, which runs 6 miles on each side of the Grand River from its source to Lake Erie. Over the years, this land was systematically appropriated by the Crown, in contravention to the Haldimand Proclamation of 1794. Douglas Creek Estates was the last straw in a dispute running over a hundred years.
The Haudenasaunee were the last holdouts against colonial government. In 1924, the Six Nations hereditary council was removed, by gunpoint, and the community was forced to adhere to the Indian Act. To their credit, they have always opposed assimilation aggressively and have never agreed to be subject to Her Majesty's laws. In fact, the traditional government in Six Nations remains intact.
None of this justifies terrorism and violence against persons and property. The Haudenosaunee and the Mohawk Warrior Society have a responsibility to clean up their act, or more people will be hurt the next time tempers flare. But the Haudenosaunee have the right to stand up aggressively against their oppressor.
Where were the police to protect the Six Nations community in 1924 when the Council was threatened at gunpoint?
They were holding the guns.
Tags: Aboriginal, Caledonia, Cayuga, Chiefs, Confederacy, First Nation, Haudenasaunee, Mohawk, Nipissing, Occupation, Oneida, Onondaga, Ontario Provincial Police, OPP, Seneca, Six Nations, Tuscarora, Warrior
Category: Anishinaabek | 3 Comments
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