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AID/APOBEC cytosine deaminase induces genome-wide kataegis
Artem G Lada1,
Alok Dhar2,
Robert J Boissy3,
Masayuki Hirano4,
Aleksandr A Rubel5,6,
Igor B Rogozin7,8 &
Youri I Pavlov1,5
Clusters of localized hypermutation in human breast cancer genomes, named "kataegis" (from the Greek for thunderstorm), are hypothesized to result from multiple cytosine deaminations catalyzed by AID/APOBEC proteins. However, a direct link between APOBECs and kataegis is still lacking. We have sequenced the genomes of yeast mutants induced in diploids by expression of the gene for PmCDA1, a hypermutagenic deaminase from sea lamprey. Analysis of the distribution of 5,138 induced mutations revealed localized clusters very similar to those found in tumors. Our data provide evidence that unleashed cytosine deaminase activity is an evolutionary conserved, prominent source of genome-wide kataegis events.
This article was reviewed by: Professor Sandor Pongor, Professor Shamil R. Sunyaev, and Dr Vladimir Kuznetsov.
Sequencing of cancer genomes has revealed that tumors contain numerous mutations [1]. It is unclear how such a large number of DNA sequence changes are induced and what factors influence the distributions of the rates of mutations among cells and within regions of the genome during tumor development. Recently, regions of localized hypermutation, called kataegis, have been detected in breast cancer genomes [2]. Based on the prevalence of C:G->T:A transitions in these regions and the sequence context of the mutations, it has been hypothesized that mutation clusters in cancer are induced by AID/APOBEC editing deaminases [2, 3]. This is consistent with the ability of deaminases to produce multiple deaminations in ssDNA in vitro that can be recovered as clustered mutations in bacteria [4, 5]. Using Sanger sequencing, Liu et al. discovered AID-induced mutations in various loci in B-cells [6]. APOBEC3B can introduce base substitutions (detected by 3D-PCR) in a reporter gene integrated into the genome of a human cell line [7]. However, a direct link between APOBECs and kataegistic clustered mutations has not been reported. A yeast model is an efficient approach to study this phenomenon. Regions of ssDNA are recognized as a prerequisite for kataegis-like events induced by an alkylation agent in yeast, and by extrapolation, have been proposed to be a prerequisite for the kataegistic action of deaminases in humans [3]. Double-strand DNA breaks in the vicinity of a reporter gene synergistically stimulate mutagenesis by AID, and in yeast this behavior might be related to the generation of ssDNA during homologous recombination [8].
We examined kataegis induced in diploid yeast by the most mutagenic AID/APOBEC protein, PmCDA1 from sea lamprey [9]. AID/APOBECs belong to a superfamily of proteins with diverse functions, from RNA editing to humoral and innate immunity and DNA demethylation [10]. Intriguingly, the basis of such a plethora of functions is a relatively simple reaction: the deamination of cytosine to uracil in ssDNA or RNA. During replication, uracil pairs with adenine resulting in a C:G->T:A transitions in the next round of replication. We expressed an exogenous PmCDA1 gene in a diploid yeast strain LAN210 defective for uracil-DNA-glycosylase (ung1). The Ung1 protein excises uracil during base excision repair in yeast; therefore, UNG1 inactivation sensitizes yeast to APOBEC effects. At the same time, Ung1 deficiency abrogates PmCDA1's ability to induce mitotic recombination [9]. PmCDA1-induced canavanine-resistant (Canr) mutants were selected and their genomes were resequenced using the Illumina platform, which involved the mapping of reads corresponding to the mutant clones against a reference produced by DNA sequencing and de novo assembly of the basic LAN210 genome. We also sequenced the genomes of Canr mutants induced in an isogenic diploid strain by the powerful base analog mutagen 6-hydroxylaminopurine (HAP), one that also (like PmCDA1 in the ung1 strains) is not excised by base excision repair and does not induce recombination in yeast ([11, 12] and references therein). Therefore, the distributions of mutations obtained in our study represent unbiased snapshots of genome-wide mutagen-induced base substitutions.
To analyze the distribution of mutations in resequenced genomes, we pooled the results from four genomes of PmCDA1-induced mutants and eight genomes of HAP-induced mutants. Each chromosome sequence was divided into 1-Kbp intervals, and the number of mutations per window was calculated. The mutation densities were calculated across the entire genome and plotted as a function of each interval's chromosomal coordinate for all 16 chromosomes (Figure 1A). The distributions of the intervals with a given number of mutations are shown as insets in Figure 1B and C. Mutation randomness analysis was done using C.A.MAN [13, 14] by calculating the threshold values of the mutation densities per window. The details of experimental procedures are in Additional File 1 and in the article by AGL, Elena G. Stepchenkova, Irina S.-R. Waisertreiger, Vladimir N. Noskov, AD, James D. Eudy, RJB, MH, IBR and YIP, which is currently under review. Analysis of the distribution of HAP-induced mutations revealed three classes of windows. The first class includes windows with 5 or less mutations, the second class includes highly mutable regions (6 to 18 mutations). The threshold value of six mutations per window was chosen as for determining highly mutable windows. Analysis of the PmCDA1-induced mutations also revealed three classes of windows. The first class includes windows with 4 or less mutations, the second class includes highly mutable windows with 5 to 11 mutations per window, and the third class comprises obvious hypermutable windows (number of mutations 14, 15, 17, and 22). The threshold value of five mutations per window was chosen for determining highly mutable windows. Thus, for the respective classes of mutagenic agents, the thresholds for highly mutable 1-Kb intervals were defined as those that contained six or more HAP-induced mutations or five or more PmCDA1-induced mutations.
Improbable hotspots of mutagenesis induced by PmCDA1 in yeast genomes. A. Genome-wide distributions of mutation (also called single nucleotide variants, SNVs) frequencies in PmCDA1 (red) and HAP (blue) induced mutants. SNV frequencies across the entire genome are shown. Negative values (i.e., bars below the X axis) indicate regions of the genome excluded from analysis (mostly repetitive regions). B. Distribution of 1 Kb windows with the indicated number of SNVs for PmCDA1 (red bars) and HAP (blue bars), log scale. C. Same as B, but using a linear scale, and where values >5 are shown. D. Multiple alignment of the DNA sequences of the hypermutable region on chromosome X from different mutant clones: LAN210-L1 – LAN210-L2 – clones used for genomic sequencing; PmCDA1-cl1 – PmCDA1-cl4 – independent mutants with this region sequenced by the Sanger method; AID_cl2 and AID_cl8 – AID-induced mutants that were found to contain mutations in this region. Homozygous G->A substitutions are red; heterozygous G->A substitutions are blue Rs; heterozygous C->T substitutions are green Ys.
Overall, we found that mutation densities ≥ 5 and ≥ 6 mutations per Kb (~0.078% for PmCDA1 and 0.046% for HAP, respectively) indicate a non-random mutation induction. We found 108 such regions for PmCDA1 and 24 for HAP (see Figure 1B, C and Table 1 for the distributions of the actual counts of mutations per Kb).
Table 1 Distributions of mutations in 1000 bp windows
These numbers underscore the striking differences between the genome-wide distributions of PmCDA1- and HAP-induced mutations in the sequenced clones (Figure 1). Deaminase-induced mutation clusters are evident at various genomic loci (Figure 1A, red graphs), whereas the density of HAP-induced mutations is much more uniform (Figure 1A, blue graphs; see also Figure 1B and C for mutation distributions). Very dense clusters of deaminase-induced mutations were found on chromosomes IV, V, VII, X, XII, XIV and XVI (Figure 1A). The most striking example of a kataegistic deamination cluster is on chromosome X (at chromosomal coordinate ~174000, between the URA2 and TRK1 genes, i.e., the highest peak in chromosome X in Figure 1A). In this region, we observed 22 mutations in a 1,000 bp interval, which corresponds to ~0.43% of the mutations found in four genomes (yeast haploid genome size is ~12 Mb). All four sequenced clones possess mutations in this region (Figure 1D). These results were confirmed by Sanger sequencing. We PCR-amplified ~2,100 bp between the URA2 and TRK1 genes and sequenced the resulting product using three primers (primer sequences are available upon request). In contrast, mutation clusters were practically not found in the genomes of HAP-induced mutants. Sequencing of the region between URA2 and TRK1 in four independent PmCDA1-induced mutants confirmed that this genomic region is highly prone to dense mutation clusters caused by localized "thunderstorms" of enzymatic deamination. At position 173,122 on chromosome X, a G->A transition was found in 6 out of 8 clones, and this mutation was homozygous in 5 clones (Figure 1D). Given the average mutation load in mutant clones and the GC content of the yeast genome, the probability of observing this by chance is ~9x10-43, assuming independent mutations on homologous chromosomes. We also sequenced this hypermutable region in Canr mutants induced by a different deaminase, human AID, and found mutations in 2 out of 12 clones (Figure 1D). Since this AID is less active than PmCDA1 in yeast, we concluded that hypermutability of this region is independent of the deaminase studied, i.e., it has certain fundamental properties making it prone to mutation induction by these enzymes. In addition, a strong strand bias was found in this region (predominantly G->A mutations), which may be a result of preferential deamination on the non-transcribed strand of nearby genes.
The clusters of cytosine deaminase-induced mutations described here are very similar to those found in cancer cells [2, 3]. They can be explained by the processive activity of AID/APOBEC enzymes [4] in genomic regions where ssDNA is exposed and is highly accessible to the deaminase. The level of adjacent transcription is one of the factors that can influence the accessibility of genomic DNA to deaminases. Our results establish, for the first time, a direct link between cytosine deaminases and kataegis. It should be noted that in our system the induction of homologous recombination was suppressed, either by the nature of the mutagen (HAP) or the use of ung1 strains for PmCDA1. This makes our model more similar to human cells where this type of recombination is relatively rare. The use of diploid strains was also critical for uncovering the full mutagenic potential of the deaminase. In terms of inter-mutation distances, the strongest mutation clusters observed in our study (such as the one shown on Figure 1D) resemble the micro-clusters found in the macro-clusters in the kataegistic regions of breast cancer genomes (See Figure 4A in [2]). We found that PmCDA1 predominantly introduces mutations in ATC motifs (more details in the manuscript under review, see above). Similarly, a group of mutations in breast cancer kataegis is characterized by a TCX motif signature. Further studies will reveal if PmCDA1, a lamprey enzyme, possesses a conserved specificity. Importantly, our study reveals that the AID/APOBEC proteins can induce kataegis in the genome. The nature of the deaminase participating in this process will dictate the sequence motifs where mutations occur.
These findings have broad implications for cancer biology and evolution, especially in the context of the recent discovery of AID/APOBEC-like proteins in various prokaryotes and eukaryotes [15]. Thus, kataegis might be a widespread phenomenon in the evolution of different forms of life. Recently, localized mutation clusters were discovered in yeast mutants obtained by a different mutagen, methyl metanesulfonate (MMS) [3]. At present it is not possible to predict the impact of kataegis on the existing evolutionary models of mutation frequencies, which assume the independence of mutation events. Kataegis may also significantly influence the outcome of other studies of discrete evolutionary events, e.g. the detection of recombination patches.
Reviewer's report Title: AID/APOBEC cytosine deaminase induces genome-wide mutation clusters Version: 1 Date: 1 November 2012 Reviewer number: 1 Professor Sandor Pongor
Report form:
Regions of localized hypermutations – so-called kataegis regions – were recently found to be colocalised with regions of somatic genome rearrangements in cancer genomes. As C:G->T:A transitions were overabundant in these regions, it was hypothesized that AID/APOBEC editing deaminases that are responsible for cytosine to uracil deamination in single-stranded DNA or RNA, may be one of the causative agents generating localized hypermutations. However plausible in the chemical sense, this hypothesis is difficult to prove by experiment. In this Discovery Note, Lada et al. describe an experiment designed to provide a very interesting piece of supporting evidence to this hypothesis. The authors used a diploid yeast sensitized to deamination effects by the removal of the uracil DNA glycosylase gene (ung1) as the model organism. Then they expressed a hyperactive AID/APOBEC protein from sea lamprey in this organism and explored the distribution of mutations along the chromosomes. It was found that distribution of these mutations is highly uneven, and the differeces are especially striking in comparison with those induced by the base analog mutagen 6-hydroxlaminopurine (HAP) which was used as a control. The findings are straightforward and provide strong support to the hypothesis that unleashed AID/APOBEC may be the causative agents of hypermutations found in cancer genomes. The link between the two phenomena is that clusters of deaminase-induced mutations in yeast are very similar to those found in cancer cells. It would be very interesting to see a more detailed description of this similarity. Are there similarities in the sequence contexts? It is a convincing argument that HAP-induced mutations in ung1- mutants are entirely random, but perhaps there are other examples of or analogies with more uneven mutations in the literature where, for instance, the context of the mutations are different.
Author's response: We are glad that Dr. Pongor considered our study interesting and convincing. We would like to thank Dr. Pongor for the constructive suggestions on manuscript improvements. We have discussed the sequence context of the mutations found in our study and in breast cancer samples, as well as the recent paper (Roberts et al.) where clustered mutations were induced in yeast by a different mutagen-MMS.
The experiments are complex, and the details are described in an experiment under review. I would suggest the authors add a brief description of the experiment as an appendix to this note.
Author's response: As suggested, we have added a short overview of experiments undertaken. We also provide more details in the responses to Reviewer #3. In addition, since the format of the Discovery Notes is adapted to the short communications, we refer the interested readers to the paper (Lada et al.) that is currently under review. This paper contains the details of experiments undertaken.
In summary, I find the experiment well-designed and the conclusions convincing.
Reviewer's response: I accept the revisions.
Quality of written English: Acceptable
Reviewer's report Title: AID/APOBEC cytosine deaminase induces genome-wide mutation clusters Version: 1 Date: 15 November 2012 Reviewer number: 2 Professor Shamil R. Sunyaev
I find this manuscript to be of great interest. Two recent publications reported presence of mutation clusters induced by APOBEC proteins in cancer genomes, shedding new light on the nature of spontaneous somatic mutagenesis. This manuscript provides experimental evidence supporting the hypothesis of recent observational studies. The authors report that genomes of yeast mutants carrying the hypermutagenic deaminase contained mutation clusters highly similar to clusters (putatively caused by APOBECs) observed in tumor genomes. This is an important result and I have no suggestions for improvements.
Author's response: We are excited that Professor Sunyaev found our work to be of great interest and that it provides new information on spontaneous mutagenesis.
Reviewer's response: I did not have any concerns with the manuscript.
Reviewer's reportTitle: AID/APOBEC cytosine deaminase induces genome-wide mutation clustersVersion: 1Date: 15 November 2012Reviewer number: 3Dr Vladimir Kuznetsov
Report form: Comments
Recent papers (2,3) have provided detail descriptions of clustered mutation sites in the genomes of four human cancers and in yeast cells. In (3) functional and structural association of APOBEC proteins with clustered mutation have been suggested. However, more direct functional associations of APOBEC family member(s) proteins with clustered mutations have to be carrying out. In this study, the authors used sequencing technique and their yeast model to study genome clusters of hypermutation activity of deaminases PmCDA1 and AID.
Major concerns and my recommendations:
1. Analysis of the literature is essentially incomplete
The authors claimed: "… a direct link between APOBEC deaminase activity and genome-wide hypermutagenesis is still lacking." However, this claim has to be debated. Atomic force microscopy studies provided direct evidence of the structural details of direct interaction of APOBEC3G with ssDNA on a specific site at a sing molecular level and at nanometer resolution (Shlyakhtenko et al., 2011). Yamane et al. (2011) reported about deep sequencing analysis of mutations and identification of the genomic targets of AID in mouse B-cells and provided the evidences of association of ssDNA hypermutation sites with APOBEC binding motif. At least two papers reported functional and structural connection between AID/APOBECs and genome-wide hypermutation (Klein et al. 2011; Yamane et al. 2011). Both papers studied the impact of AID in mouse B-cells at the genome scale.
Author's response to 1: We thank Dr. Kuznetsov for extensive review of our paper that took significant effort and almost two months. In response to the critique we added a more balanced discussion of the papers that we deemed ultimately related to our study (references 2–8). As to papers mentioned by the reviewer, a very interesting article by Yamane et al. (2011) is devoted to the construction of ChIP-based whole-genome maps of AID and RPA occupancies and is neither analyzing genome-wide mutation distributions nor report the discovery of clustered mutations. Moreover, in our opinion, interpretation of the very solid experimental results of this study should be re-considered, because they are in direct disagreement to data obtained in our lab (Lada et al., 2011) and by Dr. Myron Goodman's group (Pham et al., 2008; Chelico et al., 2009). The paper by Klein et al. (2011) presents a very thorough genome-wide study where the authors used a powerful translocation-capture sequencing method to map chromosomal rearrangements in B lymphocytes. Although the authors do report that translocation hotspots were accompanied by the base substitutions, we would like to point out that, similar to the paper by Yamane et al. (2011), the genome-wide mutagenesis study is not performed in this study and mutational clusters are not detected. Moreover, the paper by Nik-Zainal (2012) reporting the discovery of kataegis and discussing the potential involvement of the APOBEC protein in the formation of clusters of mutations was published later than all of the mentioned papers. In addition, studies of activated B-cells, which provide the natural environment for the AID activity, do not explain how the genomes of breast cells become edited by the APOBEC proteins.
2. There is no description of the sequencing methods. Even the number of reads was not reported.
3. Raw and processed data are not available.
4. Sequencing generation, sequence data analysis, genome assembly and mapping procedures and results of these steps omitted.
Author's response to 2–4: The format of the Discovery Notes does not allow us to include all the Materials and Methods related to our data. We refer to our parallel paper (Lada et al., currently under review) where all the details of experimental procedures and data analysis are described in detail. However, we have added a short description of materials and methods used in this manuscript, including the numbers of reads, coverage and the NCBI accession number for the raw data. This text is available as an Additional File 1 .
5. Authors did not provide systematic evidences of accuracy of their finding.
Statistical model(s) of background noise, testing methods, and analysis of experimental results are not reported. There are no any estimates of specificity and sensitivity of the proposed experimentally detected mutation sites and clustered mutations associated with PmCDA1 and AID activity.
Author's response to 5: All draft reference genome assemblies performed in this study were manually edited and assembly errors were excluded from analysis. The remaining questionable few regions were sequenced using the Sanger method to confirm or reject the SNVs detected. The detailed description of these procedures is beyond the scope of the Discovery Notes, see response to comments 2–4.
6. A work needs to develop an analysis of the boundaries of clustered mutations; result should include the frequency tables of all observed mutation transitions occurred in clustered mutation as well as in the regions out of the clusters.
Author's response to 6: There are methods to analyze the clustering of mutations that attempt to locate the boundaries of the regions with an elevated frequency of mutations (P.J. Gearhart, D.F. Bogenhagen, 1983. Clusters of point mutations are found exclusively around rearranged antibody variable genes. Proc. Natl. Acad. Sci. U.S.A. 80, 3439–3443; H. Tang, R.C. Lewontin 1999. Locating regions of differential variability in DNA and protein sequences. Genetics 153, 485–495). These methods, however, require a much higher frequency of mutations per nucleotide and were tested for relatively short sequences.
We have used a classification approach to analyze the distribution of mutations across yeast chromosomes using non-overlapping windows. This method is not capable of finding the exact boundaries of hypermutable regions, however it allows for the detection of the general trends in a robust way. It is described in more details in the revised draft and in new Additional File 1 .
7. There are no final lists of clustered mutations and their genome coordinates and biological interpretation.
Author's response to 7: We have added Table 1, which contains distribution of mutations in 1 Kb windows.
8–9. The number of C to T substitution mutation is reported only for two genes on chrX; There are no quantitative data and numerical/statistical characteristics for clustered mutation sites, any other genes, regions and chromosomes.9. Statistical distributions of all base transitions (e.g. % of C to T, G to A etc.) should be presented and discussed. The work should provide mutation' classification and include description of the substitution mutation in the clusters on positive and negative strands and supporting by APOBEX/AID motif(s) co-localization.
Author's response to 8–9: See response to comments 2–4.
10. There is no comparison of the results of this genome-wide finding with alternative studies.
Author's response to 10: We have included a more extensive discussion of the results by Roberts et al. (2012).
11. A reason of using 6-hydroxlaminopurine (HAP) treated cells as a negative control should be explained.
Author's response to 11: We are especially grateful for this comment. One of the major emphases of the paper is to study mutagenesis in diploid yeast independent of recombination, which is uniquely frequent in this organism. We have chosen conditions and mutagens when induced recombination is suppressed and the situation is closer to processes in human cells (both HAP and PmCDA1 in ung1- strains does not induce recombination in yeast). We have updated the text accordingly to make this more transparent.
This work is essentially incomplete and poorly performed; there is no way to reproduce its methods, results and evaluate their actual value.
Author's response: See answers to comments 2–4 and 8–9. We also think that even without the knowledge of fine experimental details there is a straightforward way to reproduce the results of this work by expression of PmCDA1 gene in diploid ung1- yeast strain or treatment by HAP, selection of mutants and genome sequencing.
Quality of written English: Not suitable for publication unless extensively edited
Author's response: Please see evaluation by reviewers 1 and 2. Nevertheless, we have put forth additional effort and we have carefully edited the manuscript.
Reviewer's response:
1. "….clusters very similar to those found in tumors".
What kind of parameter(s) is similar? What kind of similarity/dissimilarity measure(s) between mutation clusters in yeast and human cancer genomes was used? Is there some statistical estimation? If it is statistical-based analysis, the test and confidence values should be reported.
2. "We also think that even without the knowledge of fine experimental details there is a straightforward way to reproduce the results of this work by expression of PmCDA1 gene in diploid ung1- yeast strain or treatment by HAP, selection of mutants and genome sequencing."
Unfortunately, NGS is not well matured and standardized technic, specifically, in context of 'The details' of experimental procedures, data analysis and interpretation. Perhaps many readers of BD whose have an experience to use NGS technics and corresponding analytical method, should disagree with the authors point. Specifically -processing, alignment, mapping results and analysis of data are not trivial steps and are usually reported in publications as regular (not referred to unpublished data). As usual publication practices, it should be presented in suppl. file.
3. "primer sequences are available upon request".
Why? This information should be present in the work, if no commercial interest.
4. "Importantly, our study reveals that the AID/APOBEC proteins can induce kataegis in the genome"
This conclusion should be too strong. The inducer(s) of "kataegis" were not defined; it might be identified in future works.
5. "Our data provide evidence that unleashed cytosine deaminase activity is an evolutionary conserved, prominent source of genome-wide kataegis events."
It might be too strong conclusion. The evidences of the evolution conservation of cytosine deaminase activity in "genome-wide kataegis" loci across species were not reported and they should be done for specific kataegis loci if any.
6. Minor: NGS Instrument model should be indicated in the manuscripts.
Quality of written English: Acceptable.
UNG:
Uracil DNA glycosylase (uracil-N-glycosylase)
HAP:
6-hydroxylaminopurine
SNV:
Single nucleotide variant
ssDNA:
Single-stranded DNA.
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We are grateful to Dr. James D. Eudy, Director of UNMC Sequencing Core Facility, for help with next-generation sequencing and Kristi Berger for expert editing of the manuscript. This work was supported by UNMC Eppley Cancer Center seed grants to YIP in 2010–2011; Smoking Disease Research Program DHHS grant 2013–21 to YIP; NCI grant CA129925 to YIP; by NIH grants R01AI072435 and R01GM100151 to MDC; and by the Russian federal program "Innovative scientific personnel," State contract #8654 to YIP, Federal Grant-in-Aid Program «Human Capital for Science and Education in Innovative Russia 2009-2013». IBR is supported by the Intramural Research Program of the National Library of Medicine at the National Institutes of Health/DHHS. AGL is supported by a Graduate Research Assistantship from the University of Nebraska Medical Center (UNMC). AAR is supported by a research grant from St. Petersburg University. We appreciate the partial support that the University of Nebraska Medical Center's DNA Sequencing Core receives from the NCRR (1S10RR027754-01, 5P20RR016469, RR018788-08) and the National Institute for General Medical Science (NIGMS) (8P20GM103427, GM103471-09). The work was partially carried out at the "Development of Molecular and Cellular Technologies Resource Center" at St. Petersburg University.
Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, NE, USA
Artem G Lada & Youri I Pavlov
Department of Genetics, Cell Biology and Anatomy and Munroe-Meyer Institute, University of Nebraska Medical Center, Omaha, NE, USA
Alok Dhar
Department of Internal Medicine, University of Nebraska Medical Center, Omaha, NE, USA
Robert J Boissy
Emory Vaccine Center, Department of Pathology and Laboratory Medicine, Emory University, Atlanta, GA, USA
Masayuki Hirano
Department of Genetics, Saint Petersburg University, Universitetskaya emb. 7/9, St. Petersburg, 199034, Russia
Aleksandr A Rubel & Youri I Pavlov
St. Petersburg Branch of Vavilov Institute of General Genetics, St. Petersburg, Universitetskaya emb. 7/9, St Petersburg, 199034, Russia
Aleksandr A Rubel
National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD, USA
Igor B Rogozin
Institute of Cytology and Genetics, Novosibirsk, Russia
Artem G Lada
Youri I Pavlov
Correspondence to Youri I Pavlov.
The authors declare no competing interests.
AGL performed the experiments, analyzed the data and wrote the manuscript. AD performed the NGS sequencing. RJB participated in data analysis and in the writing of the manuscript. MH performed the cloning and characterization of PmCDA1. AAR participated in confirmatory Sanger sequencing. IBR participated in the data analysis and writing. YIP conceived the work, discussed its experimental aspects and the outline of the work and finished writing the manuscript. All authors edited and approved the final version.
Additional file 1: Supplementary Experimental Procedures.(DOCX 17 KB)
Lada, A.G., Dhar, A., Boissy, R.J. et al. AID/APOBEC cytosine deaminase induces genome-wide kataegis. Biol Direct 7, 47 (2012). https://doi.org/10.1186/1745-6150-7-47
APOBEC
Kataegis
Diploid yeast
Hypermutation


54 years ago today an assassin's bullet shook up America and ended the supposed hopeful period of Camelot. Of course, the Whiteman's Camelot did not truly exist in Black America because Jim Crow was still surging and most of Black America questioned if justice and equality for any man or woman of color in the world or specifically existed in this nation. Our fraternity of choice was embryonic just 2 months old with 12 black brothers hoping for an organization that dreamt so bold. That is my Wednesday afternoon story to be told. How we in 63 came to be.


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Who will do the technical jobs needed to fuel a commercial spacecraft industry?
With NSF funding, SpaceTEC is preparing the next generation of spacecraft technicians
A Francis Tuttle Technology Center instructor certifies a student in CertTEC Aviation Structures.
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For more than 100 years, humans have been buying airline tickets to the next best destination. Soon, that destination could be space, and space technology companies are working hard to have the first commercial spacecraft in the air.
To be space ready, they need aerospace technicians with the right credentials, and one center is situated to provide them.
SpaceTEC National Resource Center for Aerospace Technical Education is at the front end of preparing a technician workforce for this new commercial space travel.
SpaceTEC, funded by the National Science Foundation (NSF) since 2002 through the Advanced Technological Education (ATE) Program, provides resources and nationally recognized certifications for aerospace technology. It offers assessment tools and performance-based certifications to its 18 college partners, according to SpaceTEC managing director Steven Kane.
"The commercial space companies now are like the aviation industry was back in the 1920's, opening doors for entrepreneurs like Bill Boeing to take an aircraft, stretch it, put some seats in it and start carrying people," says Kane.
According to Kane, SpaceTEC saw a drop in enrollment when NASA's space shuttle program ended in 2011, but the industry underwent some retuning.
"Companies are starting to recognize that the curriculum that was created through SpaceTEC fits their needs for those technicians to start doing that work again," says Kane. And, he adds, SpaceTEC has certified about 425 technicians for this industry.
Educating a qualified workforce is part of keeping the U.S. globally competitive in this industry, according to Celeste Carter, lead program director for the ATE Program.
Creating SpaceTEC
SpaceTEC began as a way to create a national certification program that all companies would recognize. Earlier, companies conducted their own training and certification process. This meant that a technician starting a contract at a new company would have to go through training again and start at zero, no matter his or her experience.
Brian Madgett, a SpaceTEC certified examiner, has worked in the aerospace industry for over 30 years and says he knows how essential certifications are.
"It was difficult to get into the industry because there were no standardized certifications for this type of work," says Madgett.
After being certified Madgett says the technical certification gave him an edge over other applicants.
"Employers need to see the proof of your training and experience," he says.
Partnering with educators and employers
The center works with colleges, companies, government agencies and the military to ensure that the curriculum SpaceTEC created is accepted at the educator and employer levels, according to Kane.
This way, he says, educators and industry representatives have a seat at the table when the decisions are about the curriculum.
Some colleges like Eastern Shore Community College (ESCC) look to SpaceTEC to guide their program and curriculum, according to assistant professor of electronics and computer technology John Floyd.
"As a national resource center, with decades of experience relating to the shuttle program and other high profile Aerospace ventures, the partnership gives us access to information and standards that we do not have locally," says Floyd.
He also says that ESCC partnered with SpaceTEC in 2011 to prepare students for the aerospace industry and for job opportunities at the close-by NASA Wallops Flight Facility.
Government agencies like NASA make the SpaceTEC certificate a requirement in their employee contracts.
"The industry collaborations are essential to ensure that educators are producing the graduates that industry values," says Carter.
Mechanical engineering technician Kevin McLain took the SpaceTEC exam at Thomas Nelson Community College as a requirement for his NASA contract.
"I think they made it a requirement to ensure that I had sufficient knowledge of the aerospace industry and could prove to them that [I'm] serious about [my] career field," McLain says.
SpaceTEC expands to other industries
Before SpaceTEC experienced a resurgence in certifications, the center saw an opportunity to apply the same skill sets required in their certification program in other industries, such as the automobile, airline and energy industries.
So NSF granted SpaceTEC a supplement in 2009 to create CertTEC, which offers performance-based, national certifications for technical training beyond aerospace technology. It includes certifications for Basic Electricity & Electronics, Aviation Structures, Aviation Mechanical Assembly and Basic Composites. CertTEC has certified almost 1200 technicians, according to Kane.
And like SpaceTEC, the program helps students enter and move up in their industry, according to CertTEC examiner and senior instructor Kenny Payton of the Francis Tuttle Technology Center.
"Being in the aviation field for over 40 years, the training and testing baseline insures the employee has the skills that the employer is seeking, making for an easier transition into productive employment," says Payton.
A new project for veterans
SpaceTEC's newest project is VetTEC, a one-stop-shop to help veterans apply the skills they learned in the military to industry jobs.
"When they come out of the military, there's nothing really that provides industry a guide to what this person knows," says Kane.
According to Kane, the military uses skill codes, according to Kane, but they don't translate well to job postings' qualifications. VetTEC would be a tool that would let veterans input their skill code, find the certifications that fit their skills and see the jobs available for that skill set.
The project is a work in progress, but could give veterans a way into the commercial space industry, among others.
Beyond national certification, an ongoing dialogue with industry is key to SpaceTEC's success.
"The benefit of a national certification is that no matter what college teaches the information, everybody has to meet this national standard," says Kane. "In order to ensure that there's a home for these certified technicians that are created, the industry has to be involved."
-- Kierstyn Schneck
-- Maria C. Zacharias, (703) 292-8454 mzachari@nsf.gov
A Wichita Area Technical College instructor reviews CertTEC Basic Composites examiner training.
A SpaceTEC examiner gives a student the SpaceTEC Aerospace Core certification.
Steven Kane
Albert Koller
Tad Montgomery
Michael Reynolds
William Fletcher
Related Institutions/Organizations
Cocoa , Florida
Advanced Technological Education
Related Awards
#1303935 SpaceTEC Renewal - National Resource Center

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Kaplan Fox & Kilsheimer LLP serves on the plaintiffs' executive committee for the In re Rail Freight Fuel Surcharge Antitrust Litigation (MDL Docket No. 1869), pending in the United States District Court for the District of Columbia. This is a class action brought on behalf of purchasers of rail freight transportation services during the period July 1, 2003 to December 31, 2008. Plaintiffs allege that the defendants conspired to coordinate their fuel surcharge programs as a means to fix, raise, maintain, and/or stabilize prices of rail freight transportation services sold in the United States.
The four defendants – BNSF Railway Company, Union Pacific Railroad Company, CSX Transportation, Inc. and Norfolk Southern Railway Company – are the dominant freight railroads in the United States. Together, the defendants possess a 90% market share based on track miles and a 94% market share based on rail freight revenue.
On November 6, 2007, the Multidistrict Litigation Panel centralized eighteen related antitrust class actions and transferred the actions to the District of Columbia. Plaintiffs filed a consolidated amended class action complaint on April 15, 2008 and the defendants filed motions to dismiss on May 30, 2008. On November 7, 2008, the Honorable Paul L. Friedman issued an order and opinion denying defendants' motion to dismiss plaintiffs' direct purchaser claims. In re Rail Freight Fuel Surcharge Antitrust Litig., 587 F. Supp. 2d 27 (D.D.C. 2008).
On March 18, 2010, the direct purchaser plaintiffs filed a motion for certification of a class of all entities or persons that at any time from July 1, 2003 until December 31, 2008 purchased rate-unregulated rail freight transportation services directly from one or more of the defendants, as to which defendants assessed a stand-alone rail freight fuel surcharge applied as a percentage of the base rate for the freight transport (or where some or all of the fuel surcharge was included in the base rate through a method referred to as "rebasing"). Defendants filed an opposition to class certification on July 1, 2010.


text:
<?php

/*
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/*
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Njia β

    Njia /n̥ˈʤɪːə/
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Ceterum Censeo

    Ego enim de meo sensu iudico. Nam ita mihi salva re publica
    vobiscum perfrui liceat, ut ego, quod in hac causa vehementior
    sum, non atrocitate animi moveor—quis est enim me mitior?
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redpajama_set_name: RedPajamaGithub


text:
The Revaluation Support Scheme was introduced in April 2017 and will continue until 31st March 2021 to provide discounts to ratepayers on a discretionary basis. Under the scheme, relief will only be provided where a qualifying ratepayer's bill has increased due to the 2017 revaluation. .
Ratepayers who may benefit from the Scheme will be in occupation of a property with a rateable value of less than £200,000 and whose rates bill has increased by £300 or more.
In 2019/2020, the Council will be distributing £1.13m to eligible businesses.
General rules regarding qualification:
businesses must have an increase in their bill of £300 or more, after transitional relief and other discounts and exemptions
the 2017 list rateable value must be less than £200,000
properties must be in use
the revaluation must be the cause of any increase in bill
ratepayers must have been in occupation on or before 31 March 2017
State Aid rules must be complied with (we will send you a form about this with your new bill)
Awards will be automatically allocated to accounts and revised bills issued – businesses would not need to apply for the relief.


text:
Social Sciences and Law
Country Background Information
a guide to resources for researching Latin American politics
Interdisciplinary Search Tools
Find Articles with Subject Specific Resources
Statistics and Data
News and Other Research Databases
These sources will give you a general background about your country and its broad social and political environment to help you frame the reset of your research. Many will also give you general statistics and directory information about government officials. Print resources can give you the best and most concise picture of your country. Be sure to check the publishing dates to be sure you are getting the information you need.
Political Risk Yearbook
Lengthy analysis of a country's political and economic climate and risks, the Political Risk Yearbook is unique in that it provides forecasts on likely political and economic developments. Each Country Report focuses specifically on political and business information: finding developing markets, determining currency movements, preparing for capital investments, or making judgments about corporate security.
Country Reports on 106 countries. Gives political and economic risk analysis available. Each Country Report focuses specifically on political and business information: finding developing markets, determining currency movements, preparing for capital investments, or making judgments about corporate security. Note: the tabs International Country Risk Guide (ICRG) and Political Risk Services (PRS) provide access to information regarding methodology, but do not provide access to the actual products. Access to the Country Data and the Political Risk Yearbook are available through the University Libraries A-Z list.
Political Handbook of the World Online Edition
Annual survey of government, regional issues, and international organizations. Good source for background information about countries and organizations and for an overview of the year's political events. Includes some statistics.
Political Database of the Americas
"The Political Database of the Americas (PDBA) is a non-governmental project of the Center for Latin American Studies (CLAS) at Georgetown University in collaboration with the Secretariat for Political Affairs of the Organization of American States, and with the support of other institutions in the region. Through its website, the PDBA offers centralized and systematized information about institutions and political processes, national constitutions, branches of government, elections, political constitutional studies and other subjects related to the strengthening of democracy in the region."
Country Studies [Library of Congress]
Book-length works covering the history, politics, geography, and economy of individual countries. Some are dated and therefore most useful for getting an historical perspective.
Short (4-5 page) overviews of the government and politics of various countries. Prepared for U.S. Department of State personnel on diplomatic missions.
The Europa World Year Book
Comprehensive listing of international organizations and countries of the world. Includes summary information about the history, economic, and government affairs of each country and statistics on economy, demographics, and industry.
Armed Conflict Database
overs conflicts in Mexico, Colombia, and the Northern Triangle (Honduras, El Salvador, and Guatemala). Provides an interactive source of information on a wide range of conflicts including terrorism, refugees, and returnees, internally displaced persons (IDPs), weapons flows, fatalities, costs ($ U.S.), historical backgrounds, annual updates and timelines. Users can generate reports and download data as well as browse through year-by-year analyses and fact sheets online.
Covers international and internal conflicts, as well as terrorism. It provides an interactive source of information on 70 armed conflicts including terrorism, refugees, and returnees, Internally Displaced Persons (IDPs), weapons used and their flows, fatalities, costs ($US), historical backgrounds, annual updates and timelines. Users can generate reports and download data as well as browse through year-by-year analyses and fact sheets online.
Constitutions of the World Online
presents the results and progress of the international research project The Rise of Modern Constitutionalism, 1776-1849 by making available to the public all constitutional documents from that era.
<< Previous: Find Articles with Subject Specific Resources
Next: Statistics and Data >>
Last Updated: May 9, 2019 12:45 PM
URL: https://guides.libraries.psu.edu/latin-american-politics
Subjects: Social Sciences
Tags: Latin American politics


Caprolactam is a six-carbon one-nitrogen ring that is used to produce nylon 6. The caprolactam ring is opened through hydrolysis and the linear chains are polymerized in a continuous process to produce nylon 6.
Chlorine is a member of the halogen grouping of elements, noted for their high reactivity. Chlorine is also highly toxic and is therefore rarely transported any distance, being produced and consumed at the same site. Chlorine is produced by the electro-chemical separation from brine (concentrated salt-water), which is a highly power-intensive process that also co-produces caustic soda.
Chlorobenzene is produced by the chlorination of benzene in the liquid phase. The major use of chlorobenzene is as an intermediate for the production of nitrochlorobenzenes and diphenyl oxide, which are used to produce herbicides, dyes and rubber.
Crude oil is a naturally occurring liquid that consists of a wide mixture of hydrocarbons created by the high pressure and temperature decomposition of organic materials. Commercially viable crude oil reserves are found in distinct geographic regions, with the Middle East, the North Sea and Venezuela featuring amongst the key sources of supply. The vast majority of transportation fuels and petrochemical feedstocks are derived from the processing of crude oil.
Cyclohexane is a six-carbon saturated ring and is an intermediate in the production of nylon. In nylon production, it is used to produce KA oil, which is oxidized with nitric acid to produce adipic acid, which is then reacted with HMDA to produce nylon 6,6. It is also used as a solvent in many chemical processes.


If you've ever seen me on the road, you know how I am about email. I'm committed to finding a connection and responding within 24 hours. Some people would say "obsessed". But for me, it is an obligation to not let my travel interrupt customer service.
I was asked recently if there were any particular memories or stories about staying online internationally. .
Well, fortunately connections are getting easier than they used to be. I've found cables in Vietnamese hotel rooms, WiFi in Mexico, computers in the business lounge in Hong Kong, and free passwords at the information desk at three in the morning in the Singapore Airport. And all of that has been in the past three months! But it wasn't always that easy.
I'm not really all that smart about this stuff and how it works. But I've tried to be prepared and practical. In 1995, we signed up for a dial-up account and we still maintain it as a back up. So if there are not terminals or a wireless signal, I can just make a quick call. I download into my machine, disconnect, answer emails, then connect again and send responses.
So if you get one of my emails from the festival in Cape Town, know that I'm in kitchen of the B&B, plugged into their phone, and paying for an international call while the mail transfers.
In 1998, I was sharing a hotel room in Tasmania with Peter Lynn. There was a phone, but it was hard-wired into the wall. I used my leatherman to unscrew the wallplate and get access to the raw wires. Back then, I even carried a kit with a modular pug, wires and alligator clips. But despite my efforts, I couldn't get a dial tone.
So I walked down into town with my laptop until I found a book/computer store open. I gave them a credit card and asked if I could buy a call to the States. Downloading mail should take less then three minutes. But in this case, someone sent me a photo attachment the size of Rhode Island. Twenty minutes later, it finally cleared -- and fortunately, so did my credit card.
The B&B in Weymouth England doesn't have telephones at all! For a while, I'd walk into town and use the Internet cafe -- at 1 pound (nearly $2 back then) for every 15 minutes online. And of course, I had to use their machines and check mail live online. That would explain my rushed typing.
Since the cafe was not open in the morning before I was due on the beach, I tried walking around the neighborhood with an open laptop, "dousing" for an unsecured connection.
In Korea, there were no phones in the room, so they let me call from the front desk. Problem was that people kept walking up and asking me for their keys or something. In Germany, the phone was analog and the computer digital (or something like that). So the manager had a conversion kit that he would loan out one-guest-at-a-time.
WiFi has made life much easier.
I was nervous in Istanbul. For a week, I was in a hotel where phone calls had to be placed through the front desk. You'd call the desk, give them a number, and 5-10 minutes later, a call would be connected back to the room. And of course that didn't work with a computer so I was walking to the festival office ten minutes away each morning. Then I discovered that a nearby office was generating a signal. It was one bar - weak - but enough to get though if I was patient. And by now, I was screening out large attachments.
In Malaysia this year, the hotel offered in-room connections in one-hour increments. Log on, and stay connected for up to an hour. If you disconnected sooner, you were done. But in the back of the hotel, there was a free signal. So we'd just take our machines back there.
I now carry a phone that works anywhere in the world. But that means sometimes it rings in the middle of the night and I get expensive calls from friends who think I'm still in the States. Still, it is good to be accessible. My newest acquisition is a gizmo called MagicJack that allows me to make phone calls through the internet for free.
As technology has evolved, I've thought about getting an IPhone. But my sub-notebook is easier to type on, easier for me to read, and contains copies of the web page and all my business records. I'm portable! And often, people I've writing to don't even know I'm out of the country.
Susan has tried emailing me from the cruise she takes with her mom each year. The cost was outrageous, but calls were even more expensive. But she once emailed me to check a specific web-cam site at a specific time, and I got to see her on deck waving as she passed through the Panama Canal. And now she takes my phone with her -- and gets my friends calling in the middle of the night.
My best experience? Lufthansa tested in-flight wireless web access a few years back. That made ten hour flights much more tolerable. Which they had continued the experiment. But I'm sure the time will come.
In spite of the occasional challenge -- it's pretty amazing when you think about it!
Folks in the Forum suggested that we invite readers to send in photos, comments, and anecdotes to combine for our 500th Update. We've received a few, but not all that many.


text:
in-cyprus Thousands rally against Hungary's overtime work law, PM Orban
Thousands rally against Hungary's overtime work law, PM Orban
Thousands marched through Budapest's city centre on Saturday to protest against a new law that allows employers to ask staff to work up to 400 hours per year of overtime.
Opposition groups have staged several rallies in the past weeks in the Hungarian capital and other cities against what they said was an authoritarian rule of conservative nationalist Viktor Orban.
Saturday's rally, organised by opposition parties, trade unions and civic groups, mainly targeted the new labour law dubbed by critics as "slave law".
The protesters marched in snowfall from the historic Heroes Square to the parliament building on the bank of the Danube river, carrying banners such as "Sweep away the regime".
"We disagree with almost everything that is going on since this government got into power (in 2010), from corruption to pseudo-democracy," said 50-year old housewife Eva Demeter.
She said more Hungarians were pouring onto the streets because the slave law "affects a bigger crowd".
Some of the protesters posted on social media or carried banners calling for a "national strike".
The modification to the labour code passed by parliament last month has faced intense criticism and sparked the biggest street protest in over a year. Potentially, it could add two extra hours to an average work day, or the equivalent of an extra workday per week.
Zoltan Mucsi, the head of steelmaker Dunaferr's Vasas trade union, said it was undemocratic that the government did not discuss the labour code changes with the unions.
Some of the main trade union groups may resort to a strike if the government does not sit down with them to negotiate, he told Reuters.
Trade union membership in Hungary is estimated at below a tenth of the workforce, about half the level in Germany, according to data from the Organisation for Economic Co-operation and Development.
The past weeks' anti-government rallies were also against a law that sets up new courts which critics say could be politically manipulated, and against bias in state-controlled media.
The earlier protests have been mostly peaceful, though there were some clashes with police who used tear gas.
The ruling Fidesz party reiterated in a statement on Thursday that the protests were part of a campaign for the European Parliament elections in May to help those who support mass migration into the European Union.
Fidesz won Hungarian elections with a landslide last year, with the ticket of resisting mass immigration into the EU.
Previous articleFrench 'yellow vests' rail against unrepentant Macron, tear gas in Paris
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in-cyprus
Covid-19 variant likely in Cyprus, epidemiologist says
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text:
Q: VS Code ssh; server install path I am trying to change the VS Code server folder install location. The functionality was recently implemented and I've found the following references [1], [2].
However, I have not been able to implement it myself and in trying I get the following error message:
C:\\Users\\username/.ssh/config: line 1: Bad configuration option: remote.ssh.serverinstallpath
C:\\Users\\username/.ssh/config: terminating, 1 bad configuration options

The config file is constructed in the following way
"remote.SSH.serverInstallPath": {"hostname": "/test/location"}

Host hostname
  HostName hostname
  User username

NOTE: I have obscured the connection as 'hostname' and the user as 'username'

A: This config is for vscode's settings.json file (not .ssh/config):
e.g on Windows, do Ctrl+Shift+P > "Preferences: Open User Settings (JSON)". Then, you can add the following lines at the end.
"remote.SSH.serverInstallPath": {
    "<host>": "/test/location"
}

Alternative, if you want do it via settings UI:

*

*File > Preferences > Settings

*Filter: @ext:ms-vscode-remote.remote-ssh install

*Under "Server Install Path" > Add Item

*

*Item = <host>

*Value = /test/location

redpajama_set_name: RedPajamaStackExchange


text:
Tag: etrgf
NCV Details the 2022 Performance of Its Family of Cannabis Indices
As we described in a previous article, the Global Cannabis Stock Index melted down in December after a two-month rally. The index fell 29.0% and ended 2022 down a record decline...
- January 2nd, 2023 at 7:52 am
Earnings and Elections Showdown
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Cannabis Stocks Rally Substantially in October
As we described in an earlier article, the Global Cannabis Stock Index recovered strongly from the challenging September in October, gaining 17.1% The index is still down 59.9% in 2022. In...
- November 1st, 2022 at 9:53 am
LiUNA Pension Fund Commits Additional C$30 Million to Entourage Health Through Credit Facility
Entourage Health and LiUNA Pension Fund Upsize Credit Facility with Additional $30 Million in Non-Dilutive Funding Availability Company also confirms it has signed an updated amended and restated credit...
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Cannabis Indexes Fall Far in September
As we described in an earlier article, the Global Cannabis Stock Index fell an astounding 25.9% during September, which was its worse month in more than three years. This followed a...
- October 3rd, 2022 at 7:55 am
Cannabis Stocks Get Interesting After This Market Participant Bows Out
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NCV Stock Indices Provide a Performance Update for May
In this exclusive article we will summarize the performance of the managed indices that New Cannabis Ventures offers to its readers. We will discuss the performance of the Global Cannabis...
- June 1st, 2022 at 1:28 pm
Entourage Health Q1 Revenue Increases 17% Sequentially to C$12.4 Million
Reports Record First Quarter 2022 Financial Results Gross revenue* of $17.4 million, up 28% sequentially and 37% year over year Total revenue of $15.8 million in Q1, 2022 represents...
- May 30th, 2022 at 7:08 pm
Pineapple Express Enters Agreement to Offer Same-Day Medical Cannabis Delivery to Entourage Health Patients
Fire & Flower continues to expand delivery services through CannDeliv technology platform and industry-leading logistics TORONTO, May 4, 2022 /CNW/ – Fire & Flower Holdings Corp. ("Fire & Flower"...
- May 4th, 2022 at 7:53 am
Entourage Health Receives Additional $15 Million Funding by Upsizing Existing Credit Facility
Entourage Health and LiUNA Pension Fund Upsize Credit Facility with Additional $15 Million in Funding Availability TORONTO, April 29, 2022 (GLOBE NEWSWIRE) — Entourage Health Corp. (formerly WeedMD Inc.)...
- April 29th, 2022 at 4:39 pm
Canadian Cannabis Stocks Fall 1.4% in March
The Canadian Cannabis LP Index fell for the ninth consecutive month, declining 1.4% to 169.46 in March: The index, which fell 29.2% in 2021 to end at 194.95, is down 13.1%...
- March 31st, 2022 at 5:20 pm
Canadian Cannabis Stocks Slide Almost 3% in February
After retreating 26% in Q4, the Canadian Cannabis LP Index extended its losing streak to seven months in January, declining 9.5% to 176.42. In February, the pace of the decline slowed,...
- February 28th, 2022 at 5:05 pm
Large Companies Push Canadian Cannabis Stocks 9.5% Lower in January
After retreating 26% in Q4, the Canadian Cannabis LP Index extended its losing streak to seven months as it fell again in January, declining 9.5% to 176.42: The index, which fell...
- January 31st, 2022 at 5:16 pm
Canadian Cannabis Stocks Cap Challenging Year With 13% Decline in December
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LiUNA Pension Fund to Lend Entourage Health Additional $20 Million
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- December 24th, 2021 at 1:31 pm
Canadian Cannabis Stocks Drop 5.5% in November
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- December 1st, 2021 at 8:18 am
Canadian Cannabis Stocks Drop Over 9% in October
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The Canadian Licensed Producer Index Is Now Down 4% in 2021
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- September 30th, 2021 at 5:19 pm


text:
Derbyshire February 2020
Ballidon All Saints - Open
Bonsall St James - Open Carsington St Margaret - Open
Boylestone St John The Baptist - Open
It was to be a full day out in Derbyshire, with half an eye kept out for the weather. There was a severe weather warning for snow for late in the afternoon and it was a case of getting shot what was possible and then heading home once the weather turned. It didn't quite work out as planned though!
We had reached Ballidon, a hamlet of less than 100 people, to the west of Wirksworth by mid morning. It was bitterly cold and the wind was screaming across the hills. The tiny church of All Saints here is redundant, and cared for the by Friends of Friendless Churches, a charity that I hold dear!
Structurally, this is a simple two cell building of nave and chancel, the two almost seamlessly running in to each other. There is a bellcote to the west, a south porch and a north vestry. The church is very isolated, in the middle of a field, a few houses visible through the trees to the north. Apart from that the only real neighbour is the local quarry.
The church here dates from 1205, built as a chapel of ease to Bradbourne a few miles away to the south. It is thought though that a church may have existed here before that time. In those days the size of the village is liable to have been greater than it is today. The church was added to during the 14th century, and there was a period of restoration in the 1820's and rebuilding in the 1880's.
The website had stated that the church was liable to be open, and so it proved to be. Stepping inside, the noise of the wind was incredible. Memories of taking a communion at Scott Willoughby in Lincolnshire a few years previously; a remote church where the noise of the wind was deafening throughout!
Plain and simple inside, as you would expect, with benches leading up to the restored chancel arch; the Creed and the Lord's Prayer mounted to the wall on either side. There are four steps up to the chancel, which has an oak reredos and the Ten Commandments mounted on to the east wall. The east window has Victorian stained glass.
During a six week period, I saw around 100 churches in Derbyshire; including some of the biggest and the most impressive. Churches like this though hold a special place in my heart. Small, redundant, isolated but still loved and nor forgotten. These churches are part of our heritage and our church preservation charities do a fabulous job in helping to ensure that churches such as this are cared for so that future generations can enjoy them. Each generation are temporary custodians of these buildings and each generation needs to care for them; to pass on to the next.
The church of All Saints, Ballidon.
The severe weather hit just as we were leaving Ballidon. The snow was very hard but the roads were okay at that time so we pressed on. We took a trip back to Crich, which had been closed the previous day, when I had got my opening hours mixed up. This church will be covered on another page. By the time that we got to Bonsall, the snow was settling and all was looking white and lovely.
I found the exterior of St James the Apostle difficult to photograph; the snow was coming down so hard at that time that I was reduced to shooting whatever I could from the cover of whatever trees were around.
The church consists of west tower with spire, nave with north and south aisles, chancel and south porch. The church dates from the 13th and 14th centuries and was restored in the 1860's.From the cover of the trees I could establish that there were gargoyles around the tower. I wasn't prepared to investigate these; nor the church grounds to be honest!
A quick dash to the south porch and it was lovely to see that the church was open. I was pleased for two reasons. Firstly, it was good to see it open! Secondly, it delayed me having to go back to the van!
The lack of daylight, combined with the large amount of stained glass meant that it was very dark inside so lights on here was a must; as much as I don't like using them.
The nave is separated from chancel by a screen, with a wooden carving of the crucifixion above; text below reading 'Not one of them shall be forgotten before God'. Brightly coloured kneelers rest on top of the pews and there was stained glass in the east windows of the chancel and aisles.
One monument caught the eye. A putti, a naked child, is in mourning, wiping away a tear, with hand pressing down on a human skull. We often see occasions where the skull, as a symbol of death is pressed down upon, leaned against or sat on. This appears to have been used as a symbol to point out that death has been beaten. We have unlearned some of the symbols that were used in centuries past. This monument also has a broken column, a symbol of a life that has been cut short.
There is a lot of stained glass here, and it was enjoyable to spend some time looking around it. The east window of the chancel shows the last supper, which in fairness is not the greatest quality. The stained glass in the east end of the south aisle depicts the crucifixion and the ascension at the centre pieces, with four smaller panels around them showing four scenes from Matthew Chapter 25 verses 35 and 36 'For I was hungry and you gave me something to eat, I was thirsty and you gave me something to drink, I was a stranger and you invited me in, I needed clothes and you clothed me, I was sick and you looked after me, I was in prison and you came to visit me.'
Elsewhere, the parable of the Good Samaritan shows the Samaritan tending the wounds of his mortal enemy, a Jew; the priest and a Pharisee walking on ignoring the scene. This window, to my mind, fits in very nicely with the window in the south aisle. What to do and what not to do! How to behave!
The church of St Margaret, Carsington.
We moved onwards, a little slowly to be fair and arrived at Carsington; the church of St Margaret. The scene was just beautiful; but the quantity of snow that had come down; but had now eased was such that we were going to start making our way home after this church had been photographed. The school bus sideways across the road a little way down from the church helping us to make a decision!
There was a church here back as far as the 12th century, but the church that we see today dates from 1648. It stands on the bottom slopes of Carsington Pastures, tree lined behind, with the whole scene looking glorious covered in snow. A scene to remember with great fondness!
The building itself is really just a single cell, with nave and chancel flowing in to one another. There is a bellcote to the west end, a south porch and a north vestry at the western end.
It was good to find the church open. Inside, nave flows in to chancel with no chancel arch. Inside, things are plain and simple. Whitewashed walls, simple alter with no reredos. The nave is panelled at the sides, there is a painted coat of arms on the north wall dated 1706. The font certainly looks to predate the 17th century and has carvings of human heads around the base. The east window is of interest; with Jesus at the centre of three panels, looking directly at those approaching the communion rails, holding a light out in front of Him 'I am the light of the world' reads the text below.
There are two windows to north and south, directly opposite each other. Both are of interest. One is concerned with communion, with wording from the creed under the window reading 'I believe in the communion of Saints'
To the left is St John, who holds a goblet of wine. This has a dragon coming out of it. John was given a poisoned goblet of wine and the poison, symbolised here by the dragon, left the wine after he prayed over it. To the right, holding the bread is St Philip. Philip is present here due to John Chapter 6 verse 5 – 7 " When Jesus looked up and saw a great crowd coming toward him, he said to Philip, "Where shall we buy bread for these people to eat?" He asked this only to test him, for he already had in mind what he was going to do; Philip answered him, "It would take more than half a year's wage to buy enough bread for each one to have a bite!"
The window opposite depicts St Helen, St Giles and Margaret of Scotland. St Helen holds a cross; in her final years it is said that she found the true cross whilst on pilgrimage. St Giles is pictured with a deer and an arrow. Legend states that Giles had a friendship with a deer while he lived as a hermit. He was hunted by the Kings men who shot an arrow at the deer; missing the deer but hitting St Giles. He became the patron saint for those with physical disability. Margaret is crowned and holds a Bible.
A beautiful church, photographed in the most glorious of surroundings! Although the snow had made the Derbyshire countryside just fabulous, it was hard going on the roads and Gary's sat nav kept updating as roads were being closed. At one point there was a brief discussion about whether we would have to find a travel lodge or similar to hole up in for the night.
Eventually though, we got through the worst and daylight was fading by the time that we went approached the turning for Boylestone. There had not been so much snow here, so we were safe to stay and take a quick look at the church of St John The Baptist.
There is some history to this small village, which can be found eight miles east of Uttoxeter. There was a village mentioned here at the time of the Domesday Survey in 1086. The village is famous for the English Civil War 'Bloodless Battle of Boylestone.' One evening during August 1644, Royalist troops on their way to Wingfield Manor took refuge in St John the Baptist church. During the night they were surrounded by Cromwell's men and forced surrender.
The church of St John The Baptist is in a delightfully rural setting, set apart from the rest of the village a little. The church dates back to the 14th century, with the tower added in 1844. The chancel was restored in the late 1880's.
Approaching the church from the west, the visitor immediately sees that the tower is offset to the south west. The church consists of nave with tower, south aisle and chancel. The three stage tower has a clock on the south face and a stair turret to the south also. On top of the tower is a very small spire.
There was a thin coating of snow, but they hadn't had the amount that they had had a few miles away. It was late in the day, on a freezing cold winter afternoon, it would be dark in a little while; but still the church was open! It is really easy to forget the army of volunteers that keep these churches open to visitors throughout the year; rain or shine and in the depths of winter. They make our hobby possible.
The main entrance is through a door in the west face of the tower. Script over the inner door reads 'This is none other than the house of God'.
Moving inside, the pews are Victorian; the chancel is separated from the nave by an iron gate; standing at the chancel and looking west, we can see that the west end of the church is screened off. Unusually, the plain octagonal 14th century font is to be found towards the east end of the south aisle rather than traditionally the west end of the nave. Boards are mounted to the east wall of the south aisle, detailing the Creed and the Lord's Prayer.
The stained glass in the east window is on two levels. The upper level depicts the crucifixion in three panels. The bottom level, partly hidden by the Victorian oak reredos show the annunciation, the nativity and Jesus as a baby presented to Simeon in the temple.
Another stained glass panel shows John the Baptist, with tunic made out of hair 'Ecce Agnus Dei' reads the text alongside 'Behold the lamb of God'.
It would have been the easiest thing in the world to stay inside; and in fairness doubtless more sensible! As a photographer I believe that you get out what you put in. If you are prepared to go out in the cold and the snow and the wind you will see some lovely sights; and this was the case here! Some pleasant memories; both of the churches and the conditions that some of them were photographed in,


The Edge Setup Quick Reference Guides are included in the box when we ship Edge's to the customer. These guides provide straightforward visual instruction on how to get an Edge up and running, but are intended to be used alongside the standard, and more detailed, documentation found here.
Information and procedures on other telephony components.


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Thought for the week: What is your most vivid memory of 2015 and why?
My most vivid memory of 2015 is when I went to Chesington because it was my first time ever.
My most vivid memory in 2015 is when I went to the House of Parliament with my class and we had the best time we saw the many MPs that was the most vivid moment in 2015.


If you are planning to go on vacation, one of the things you need to do is plan well. This is a process, which can be stressful and hectic. You need to be able to take it easy and know that when time comes for your trip things will go as planned. It is important for you to select a good travel company if you want things to go well. It takes even more keenness to pick a luxury travel company. You have to choose the company that will be worth the money you are asked to pay. Highlighted here are some of the things you ought to know when selecting a luxury travel company.
First, it is important that you get to know what your alternatives are. It helps to have a variety of alternatives to choose from. Do take your time to note down the various alternatives you come across while searching for a luxury travel service on the web. Create a list of at least three or so companies you feel would offer this service to you the best. After you finish your list, you can continue with narrowing down your alternatives to one that best suits you.
You should also take time to read online reviews. You can get a lot of quality information that can help you make an informed decision online. There is so much that you can learn by reading reviews because of how resourceful they are. Through reviews you can learn about the cost of services, the reputation of the company and quality of services that you can expect. All these aspects enable you to work towards making the right decision.
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In conclusion, it is important to ask for advice before you settle on a specific luxury travel company. Looking for advice allows you to get the kind of luxury service provider that you need within a short time. There are so many places that you can ask for recommendations. Thanks to the internet, there is no type of information you won't be able to find online. Look for relevant forums or even ask any of your colleagues to recommend a good luxury travel agency. However, you need to be careful only to take advice from people who are trustworthy.


The hack-n-slash RPG challenges players to defeat evil forces and save the world from darkness.
Nexon and NAT Games announced the launch of Heroes of Incredible Tales (HIT) on iOS and Android. The hack-n-slash RPG challenges players to defeat evil forces and save the world from darkness.
In each of HIT's 200 campaign levels, gamers can use a virtual joystick to move their characters around environments, and tap buttons to attack, activate skills or defend against oncoming attacks. Users can also toggle auto play, which allows the computer to control their characters.
As gamers progress, their heroes will level up, which improves their stats. Users can also upgrade their heroes' skills, as well as unlock new skills for their heroes over time. Finally, gamers can collect new upgradable weapons and armor items for their heroes as they play.
Outside of the game's single-player campaign, users can complete daily dungeons, as well as participate in a single-player Tower of Trials mode. The game also includes cooperative and competitive real-time multiplayer modes, including an Arena and Guild War, among others.
With HIT, we set out to raise the bar of core gaming standards and redefine player expectations on these powerful mobile devices. With the game's visceral HD graphics, living 3-D environments and fast and fun combat, we're confident that console and PC players around the world will feel right at home when they get their hands on HIT.
Heroes of Incredible Tales is available to download for free on the iTunes App Store and Google Play.


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HomeLatest News & PoliticsINEC Chairman should resign now – PDP
INEC Chairman should resign now – PDP
February 16, 2019 Yomie Latest News & Politics, Today's Trending.... 0
Uche Secondus, national chairman of the Peoples Democratic Party (PDP), has asked Mahmood Yakubu to resign as the chairman of the Independent National Electoral Commission (INEC).
Speaking through Ike Abonyi, his media aide, shortly after the elections were postponed by INEC, Secondus said his party was rejecting the new dates.
He said it was a "deliberate, pre-determined agenda of President Muhammadu Buhari to cling on to power even when it's obvious to him that Nigerians want him out".
Secondus said the postponement was part of a "grand design" by the All Progressives Congress (APC) to thwart the will of Nigerians "at all cost".
He said the APC "in connivance with INEC have been trying all options including but not limited to burning down INEC offices in some states and destroying of electoral materials to create artificial problems upon which to stand for their dubious act".
"With several of their rigging options failing, they have to force INEC to agree to a shift in the election or a staggered election with flimsy excuses pre-manufactured for the purpose," he said in the statement sent to TheCable.
"For the avoidance of doubt the PDP sees this action as wicked and we are also aware of other dubious designs like the deployment of hooded security operatives who would be ruthless on the people ostensibly to scare them away."
Secondus said the "wicked killing" of over 60 persons mostly women and children in southern Kaduna on the eve of election "is a copious ploy by the APC to frighten the people away from voting knowing too well that they were not going to record any vote from the area".
"Recall that the Governor of Kaduna state, Mallam Nasir el-Rufai had earlier threatened international election observers of going to their country in body bags and with the fatal violence in the state on the eve of election, it's clear what the motives are, to frighten the observers from the state so that he can carry out his nefarious acts," he said.
The PDP chairman also drew the attention to the statement of President Muhammadu Buhari that nobody can unseat him from office as an indication of what he wanted to do.
Don't be discouraged – Saraki tells Nigerians
Election postponement a national embarrassment – Oshiomhole
I will contest in 2018 to complete my truncated first term, says Fayose
May 22, 2017 AdeLove Latest News & Politics, Today's Trending.... 2
Governor Ayodele Fayose of Ekiti State, on Monday evening said he would contest the 2018 gubernatorial election so that he could serve his term which was truncated in 2006. Fayose was impeached in 2006. However, […]
PDP needs Southwest to succeed – Bayelsa Governor
December 15, 2017 Yomie Latest News & Politics, Today's Trending.... 0
Chairman of the Peoples Democratic Party (PDP) National Committee on Reconciliation and Bayelsa State Governor Henry Seriake Dickson has said the party needs the support of the Southwest to regain power in 2019. A statement […]
Miracle!!! Nigerian Woman Gives Birth After 21 Years of Barrenness … See more photos
November 5, 2015 AdeLove Latest News & Politics, Today's Trending...., Uncategorized 8
Wow! God is so wonderful!!! There is no limit to what God can do as confirming in the life of this woman that has suffered barrenness for the past 21 years. She just gave birth […]


Melissa Data, a provider of global contact data quality solutions, has introduced active data quality tools and services for Microsoft Dynamics CRM.
Trifacta is releasing an enhanced version of its namesake platform, further boosting its data wrangling capabilities and bolstering the solution for more data sources and diverse users. The v4 release extends the ability of users to directly connect to a variety of enterprise data sources, including Oracle, MySQL, Microsoft SQL Server, PostgreSQL, and Teradata, and expands support for customers deploying Trifacta in cloud environments such as Amazon Web Services, Google Cloud Platform, and Microsoft Azure.
At BMC Engage 2016, BMC's annual user conference, the company expanded its Digital Enterprise Management (DEM) strategy with initiatives designed to help customers fast track digital business.
Enterprise-IT-Security.com announced a new release of its system dump and log anonymization solution, SF-SafeDump for z/OS mainframes. Version 4.1 includes new capabilities including zIIP support of up to 95%, which will cut the cost of contractually and legally mandated anonymization procedures to a minimum. Version 4.1 also supports additional dump and log types.
IBM revealed a series of servers designed to help propel cognitive workloads and to drive greater data center efficiency. Featuring a new chip, the Linux-based lineup incorporates innovations from the OpenPOWER community intended to deliver higher levels of performance and greater computing efficiency than available on any x86-based server.
The DevOps approach to software development and IT operations is examined in a new study produced by Unisphere Research and sponsored by Dell Software. At its core, the study points out, DevOp is more than a development process or a set of tools, and reflects innovative thinking and collaboration, and holds the potential for better deployment outcomes.
Hewlett Packard Enterprise (HPE) revealed several new solutions designed to strengthen the approach to intelligent security operations, DevOps, and data protection. "We're providing visibility without boundaries," said Mark Alba, senior director of product marketing for HP Security. "In an intelligent security operations environment, customers need to have access to all types of data in order to establish their security posture, understand what's happening within their environment, and be able to make real-time, fast decisions on how to remediate potential breaches and protect against them."
erwin Inc. has acquired UK-based Corso Ltd, a provider of enterprise architecture solutions. erwin has also announced general availability of erwin CloudCore, an integrated cloud bundle consisting of erwin Data Modeler and Corso Agile EA.
OpenText Corporation, an enterprise information management software provider, has entered into a definitive agreement to acquire Dell EMC's Enterprise Content Division (ECD), including Documentum. ECD offers a suite of enterprise content management solutions with deep industry focus, including the Documentum, InfoArchive, and LEAP product families. The purchase price is $1.62 billion, which is 2.7 times ECD FY15 revenue of $599 million.
At its partner conference this week, Teradata is announcing three key new offerings to support customers choosing hybrid environments spanning cloud and on-premise, and relational and big data technologies.
Oracle is acquiring LogFire, a provider of cloud-based warehouse management applications for integrated warehouse, inventory and workforce management. The pending addition of LogFire Cloud is expected to complement the logistics functionality of the Oracle Supply Chain Management product suite by adding cloud-based warehouse management capabilities. Oracle also said it is currently reviewing the existing LogFire product roadmap and will be providing guidance to customers.
SVA Software, a provider of storage performance management software, announced its company launch in North America along with a new partner program to capitalize on the demand for its BVQ storage optimization solutions, intended to simplify the management of virtualized data center operations.
Luminex Software, Inc., a provider of mainframe virtual tape solutions, has introduced the latest release of its Mainframe Data Integration (MDI) platform. MDI enables tape data to be securely shared between mainframes and distributed systems applications, transformed and made available for offloading mainframe processing or general business operations across the enterprise.
Catalogic Software, a provider of software-defined copy data management, announced support for IBM FlashSystem A9000 and FlashSystem A9000R.
GigaSpaces, a provider of in-memory computing (IMC) technologies, has launched InsightEdge, a data grid-enabled real-time analytics platform that incorporates Apache Spark to enhance data analytics. InsightEdge is the first product to be incubated and launched as a result of GigaSpaces' Open Source initiatives for its IMC business.
Wipro Limited, an information technology, consulting, and business process services company, and Stibo Systems, a provider of master data management solutions (MDM), have formed a partnership through which the two companies will collaborate to offer MDM solutions.
Teradata has introduced four accelerators to help companies derive value from Internet of Things (IoT) data more quickly. Teradata's "Analytics of Things Accelerators" (AoTAs) are comprised of technology-agnostic intellectual property (IP) and professional services, derived from field engagements in manufacturing, transportation, mining, energy and utilities.
Logz.io, a company that offers ELK (Elasticsearch Logstash and Kibana) as a cloud service, is introducing a new analytics solution that combines machine learning technology with human experience. The new solution, called Cognitive Insights, aims to help companies give their customers a better experience by uncovering data issues before they become problems.
As the cloud continues to change the way people interact with data, one MultiValue company is helping its users get accustomed to all the new capabilities this new deployment model brings to long-established systems. "The cloud makes a level playing field for all companies regardless of size," said Mark Pick, president and CEO of Pick Cloud.
Revelation's OpenInsight solution continues to be enhanced, this time with new reporting tool upgrades and other updates. In conjunction with the Linear Hash Reporting and Resizing Tool released in March 2016, Revelation is publishing a sizelock administration tool called RTI_SET_SIZELOCK. This tool will enable OpenInsight system administrators to view linear hash statistics within an application and identify and modify the file sizelock parameter.
Each year, Database Trends and Applications magazine presents the Readers' Choice Awards, recognizing leading vendors across a variety of segments in the IT industry. Unlike any other awards programs conducted by DBTA, this one is unique because the nominees are submitted and the winners are chosen by the experts—you, the readers. MultiValue is one of the most established and trusted data technologies.