If you have not read Harry Potter 1-6, stop reading this right now. There are some spoilers in here.
Unless you've been living under a rock inside a cave at the bottom of an ocean, you know that the sixth book in the Harry Potter series was released a week ago. It sold more copies in the first 24 hours than any other book. You couldn't turn on the TV without seeing scores of 6 to 12 year olds besieging Barnes & Noble stores, threatening death if the book sold out before they got their hands on this 600+ page saga.
And despite the association with children who can barely read Avada Kedavra never mind saying it, I'm proud to say that I managed to snare a copy less than 12 hours after it was released and had the book finished within 24 hours.
My love of the underage wizard began by accident. My mother used to be a librarian for a Catholic elementary school. Because I got out of school before she did in June, she would put me to work in the library as her slave, doing all the work she despised. This meant that while she chatted with other teachers and helped them clean out their rooms, I'd stay in the library and process donated books. The parents in her school were very well-off and liked to outdo each other with donations. My job was to cover each book with clear plastic, glue in the little card slots in the back cover, stamp the name of the school on the title page, and make up card catalog entries with a synopsis of the story. Very boring work. Most books averaged 12 pages and contained nothing deeper than a lost kitty-cat. Even the books for "young adults" were pretty dull: teenager doesn't fit in, takes drugs, learns lesson after not-so-tragic accident. I had just finished high school--I had outgrown that tired cliche. The only plus-side was that I could watch TV while I did all this.
So one day I was processing these tedious wastes of trees when I got to a hardcover copy of Harry Potter and the Sorcerer's Stone. At first I thought that was a weird title: what kind of name is Harry Potter and what is a sorcerer's stone? All the same, I wasn't about to read it; I would just flip around and see what this stone was and off into the finished pile this book would go. Like most children's books, I figured this one must start off with what the goal was, so I turned to the first chapter. No mention of a sorcerer's stone here.... just a grisly double homicide. Interesting... I read a little farther... a scar? Why was this kid special? Why do these people have such strange names? What the hell is a Muggle? By this point, I was reading the book instead of skimming through, so I put it aside so I could plow through my work.
As luck would have it, a morning talk show was on. I almost changed the channel because no one interesting was a guest. But then a woman named JK Rowling was introduced. She started talking, but I was only half listening. Suddenly, I heard the name Harry Potter. She had my full attention. JK Rowling talked about how she wrote The Sorcerer's Stone... and The Chamber of Secrets... and The Prisoner of Azkaban. These characters grow up, she explained. They experience profound loss. People will die.
I was stunned. These are children's books, and people are going to die? There are three of these books? I was curious; I had to know.
By the end of that day, HP:SS was finished. I wanted more. That weekend, I got the first three books. By the end of the third book, I was hooked. Here was progression! Here was fantasy that made my Lord of the Rings look overdone! I had to know if this lovable kid with the odd name triumphed! And so I became a Harryaphile by accident.
When The Goblet of Fire came out, I had it by the end of the week. The threatened deaths had arrived. That's when I stopped seeing Harry Potter as a children's book. Those last few pages describing the rebirth of Voldemort and the death of Cedric... those pages scared me, the adult. And yet, the book did not affect me emotionally--I could care less about Cedric. The Order of the Phoenix took the darkness, the adult nature of the series even farther. JK Rowling killed Sirius Black horribly, in front of Harry, scarring both him and her readers. And yet, the series tightened it's grip on me even more.
And then there was The Half-Blood Prince. First off, I think parents who continue to let their 6 year olds read Harry Potter beyond the first 3 books are crazy. This has ceased to be for children. If I nearly lost it when JK Rowling struck down Dumbledore... And then there was the snogging! How I now love that word.
What I'm trying to convey about Harry Potter is that these books are rare gems in any literature. With the exception of the Little House and Anne of Green Gables books, I have never come across children's books that develop characters so fully to the point of making them grow up. I was 18 when I succumbed to Harry Potter, and yet this 11 year old orphan hooked me, made me want to watch him become the 16 year old teenager that watches everything he loves get destroyed. That darkness is also a lure that taps into a voyeuristic part of me that wants to know how much more this kid can take.
And so, until the 7th and final book is released, I'll continue my rereading of the books and watching of the movies. In every interview she gives, JK Rowling states more deaths are coming. So I'll wait and pray for the paper soul of Harry Potter, that he makes it out alive.
For those of you who love Harry Potter, here are a few nice sites with some interesting theories on where the series is going.


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A nucleolar localizing Rev binding element inhibits HIV replication
Alessandro Michienzi1,3,
Fernanda G De Angelis2,
Irene Bozzoni2 &
John J Rossi1,4
The Rev protein of the human immunodeficiency virus (HIV) facilitates the nuclear export of intron containing viral mRNAs allowing formation of infectious virions. Rev traffics through the nucleolus and shuttles between the nucleus and cytoplasm. Rev multimerization and interaction with the export protein CRM1 takes place in the nucleolus. To test the importance of Rev nucleolar trafficking in the HIV-1 replication cycle, we created a nucleolar localizing Rev Response Element (RRE) decoy and tested this for its anti-HIV activity. The RRE decoy provided marked inhibition of HIV-1 replication in both the CEM T-cell line and in primary CD34+ derived monocytes. These results demonstrate that titration of Rev in the nucleolus impairs HIV-1 replication and supports a functional role for Rev trafficking in this sub-cellular compartment.
HIV-1 gene expression is finely regulated [1]. Transcription from the HIV-1 long terminal repeat (LTR) produces a full length RNA of 9 Kb from which several mRNAs are then generated by splicing (the 4 Kb singly spliced and 2 Kb fully spliced RNAs) [1]. While the fully spliced RNAs are exported to the cytoplasm, where they are translated into the regulatory and accessory proteins, the late structural proteins and reverse transcriptase (RT) are derived from unspliced or singly spliced RNAs which are transported to the cytoplasm only upon binding of Rev to the Rev Responsive Element (RRE), which is contained in the env coding region [1]. Rev is an 18kDa protein that localizes to both the nucleus and nucleolus [2–4]. It contains a nuclear export signal (NES) as well as a nuclear import signal (NLS) that allow nuclear/nucleolus-cytoplasmic shuttling properties [5, 6]. The NES signal of Rev is recognized by the cellular export factor CRM1, which mediates the nuclear-cytoplasmic export of Rev bound RNAs [7, 8]. Expression of Rev in human cells induces re-localization of CRM1 and some nucleoporins (Nup98 and Nup214) into the nucleolus [9]. Recently, by the use of in vivo fluorescence resonance energy transfer (FRET), multimerization of Rev-GFP and BFP fusion proteins has been shown to occur in the nucleoli of HeLa cells [10]. These observations suggest that the nucleolar trafficking of Rev may be critical for Rev mediated export. To test this hypothesis we used a nucleolar localized decoy that contains the Rev binding element (RBE) [11–14] to sequester Rev within this sub-cellular compartment and test its ability to inhibit HIV-1 replication. The well-characterized U16 small nucleolar RNA (snoRNA) [15] was used to direct nucleolar delivery of the RBE.
In the present study we demonstrate that stable expression of the U16-RBE chimeric RNA in cultured T-cells and primary monocytes confers strong inhibition of viral replication. These data provide strong evidence that the nucleolar localization of Rev is critical for its functional role in HIV-1 replication and identifies a novel mechanism for inhibition of HIV replication.
Plasmid constructs
The U16RBE DNA was prepared synthetically as previously described [14]. The Rev Binding Element (RBE) was inserted in the U16 snoRNA sequence by replacing the apical loop [14]. The U16RBE sequence was then subcloned within the Sal I and Xba I sites of the pTZU6+1 expression cassette [16] generating the pTZU16-RBE clone. The Bam HI and Xba I cleaved fragment from the pTZU16-RBE construct was first filled in with the DNA polymerase Klenow fragment and then inserted in the Nhe I site of the pBabe puro retroviral vector (in the U3 region of the 3'LTR, Fig. 1A) giving rise to the pBabe/U16RBE clone (Fig. 1B). The U6 promoter-U16RBE was also cloned into a Stu I site in the U3 region of the MND/eGFP Banshee retroviral vector 3'LTR (Fig. 1C), generating the MND/eGFP U16RBE construct (Fig. 1D).
Retroviral constructs. (A) Schematic representation of the pBabe puro retroviral vector [22]. (B) Schematic representation of the pBabe/U16RBE construct. The PTZU16-RBE cassette was inserted in the U3 region of the 3'LTR, with the U6 promoter. Transcription is oriented in the same direction as the RNA pol II LTR and SV40 promoters. (C) Schematic representation of the MND/eGFP retroviral vector. (D) Schematic representation of the MND/eGFP U16RBE construct. The PTZU16-RBE cassette was inserted in the U3 region of the 3'LTR, with the U6 promoter transcribing in the same orientation as the RNA pol II LTR and SV40 promoters.
Stably transduced CEM clones expressing the U16RBEdecoy. The CEM cells were first transduced with the pBabe constructs and then selected for Puromycin resistance.
From the resulting pooled population single clones were selected by limiting dilution. Total RNA isolated from the single clones was electrophoresed either in a 6% polyacrylamide-7M Urea (A) or 1.2 % Agarose formaldehyde (B) gels. The electrophoresed RNAs were blotted onto a nylon filter followed by hybridization with specific probes (see Material and Methods).
HEK 293, CEM and the PG13 packaging cells were maintained as previously described [17]. Transient transfections were carried out using a Calcium Phosphate DNA precipitation kit in accordance with the manufacturer's protocol (GIBCO/BRL, Invitrogen). Briefly, 1 × 106 HEK 293 cells were plated one day prior to transfection. The transfection was carried out using 2–10 μg of plasmid DNA.
Packaging cell line
PG13 was used for packaging of the pBabe/U16RBE and MND/EGFP U16RBE constructs [17].
CD34+ cell isolation and transduction procedures
CD34+ cells were isolated from fresh umbilical cord blood using anti-CD34+ antibody-coupled magnetic beads (Milteny Biotech, Auburn, CA). Sorted CD34+ cells (purity was above 90% as determined by FACS analysis) were cryopreserved until use for transductions by the viral vectors.
One day prior to transduction, CD34+ cells were thawed rapidly at 37°C, washed and plated at a concentration of 2 × 105 onto Retronectin (Takara, Japan) coated 25 cm2 flasks. The Retronectin concentration for coating was 20 μg/cm2. The CD34+ cells were then pre-stimulated in 5 ml of cytokine media: Iscove's Modified Dulbecco's Media (IMDM – Bio Whittaker, Walkersville, MD), with 20% FBS, 100 U/ml Penicillin, 0.1 mg/ml Streptomycin (Sigma, St. Louis), 2 mM Glutamine (Sigma, St. Louis), IL-3 (10 ng/ml), IL-6 (10 ng/ml), and SCF (10 ng/ml) (R&D Systems, Minneapolis, Minnesota). After 24 hours of pre-stimulation, the media was changed and the CD34+ cells were exposed to the retroviral vector supernatants (MND/eGFP, MND/eGFP U16RBE and Mock). 50% percent retroviral vector supernatant and 50% 2 × cytokine media were used in a total volume of 10 ml per 25 cm2 flask. The 2 × cytokine media has the same composition as the cytokine media but the growth factor concentrations are doubled (20 ng/ml for all cytokines). After 24 hours of infection, the cells were centrifuged, the media removed, and the cells exposed to fresh retroviral supernatant and cytokine media in a total volume of 10 ml.
This process was repeated one more time, leading to 3 rounds of transduction.
After 3 rounds of transduction, CD34+ cells were immediately plated into a Colony Forming Units assay (CFU-assay) or Long Term Bone Marrow Cultures (LTBMCs).
Colony Forming Unit assays
1 × 103 and 2 × 103 transduced CD34+ cells from each vector arm were plated into 1 ml semi-solid methylcellulose media (Methocult, Stem Cell Technologies, Vancouver, Canada). After 14 days, colonies were counted and discriminated into Burst Forming Units Erythroid (BFU-E), Granulocyte-Macrophage Colony Forming Units (CFU-GM) and Granulocyte- Erythroid- Macrophage- Megakaryocyte (CFU-GEMM) Colony Forming Units. The colonies for each vector arm were added and the data expressed as a total colony count per group.
Long term bone marrow cultures
Transduced CD34+ cells were plated into 25 cm2 flasks onto a layer of irradiated normal human bone marrow stroma in cytokine media. The cytokine concentrations for SCF, IL-3 and IL-6 were 10 ng/ml each.
Normal human bone marrow was purchased from Bio Whittaker (Cambrex Bio Science Inc. Rockland, MD), 5 ml were plated directly into Dulbecco's Modified Eagle Media (Cambrex Bio Science) with 10% Fetal Bovine Serum (GIBCO) in a 75 cm2 flask. Bone marrow cells were allowed to adhere overnight. Media and floating cells were removed, and fresh media were added. Stromal cells were allowed to expand for about 4 weeks. After expansion, stromal cells were trypsinised and cryopreserved for later use, or irradiated (2000 rads) and plated into 25 cm2 flasks for LTBMCs at a density of 1 × 105 cells per flask.
CD34+ cells were allowed to expand in the stromal culture flasks. Cell samples were taken for eGFP FACS analysis 2 days after plating to assess the transduction frequency.
10 days after plating, cultures were depleted of the floating cell population, which was used for eGFP cell sorting and subsequent HIV-challenge. After depletion, the remaining cell fraction in the stroma flasks regenerated a high number of floating cells within 14 days. After 6 weeks of LTBMC propagation, a final FACS analysis was performed to assess eGFP expression.
eGFP sorting and propagation of sorted CD34+ progenitor derived cell populations
CD34+ derived cells were sorted for eGFP expression in a MoFLO (Cytomation) cell sorter into 6 well plates containing a layer of irradiated normal human stroma and 2 × cytokine media (20 ng/ml all cytokines). Cells were expanded in these 6 well plates for approximately 5 days. After expansion, cells were removed from the wells and divided into 2 fractions: 1 × 105 cells for eGFP FACS analysis (purity check), and 1 × 106 cells for HIV-challenge.
HIV-challenge
CEM cells (2.5 × 105) derived from stably transduced clones were infected with HIV-1NL4-3 as previously described [18]. For CD34+ derived monocytes, 1 × 106 sorted CD34+ cells were transferred into 12 × 75 tubes, centrifuged, and resuspended in 100 μl of 2 × cytokine media. 100 μl of HIV-1 JRFL with a titer of 1 × 105 /ml was added to each challenge tube, resulting in a multiplicity of infection (m.o.i.) of 0.01. Cells were incubated at 37°C and 5% CO2 for 3 hours and then 200 μl of 2 × cytokine media were added to each tube and the cells were incubated overnight.
On the following day, cells were washed 4 times and plated into 25 cm2 flasks onto a layer of irradiated S17 cells (kindly provided by Kenneth Dorshkind, UCLA), in 1 × cytokine media in a total volume 10 ml. The cultures were propagated for 28 days and samples were taken weekly. Infections were all performed in duplicate or triplicate.
HIV-1 p24 evaluation
The p24 analyses were performed using the HIV-1 p24 antigen capture assay kit (Science Applications International Corp. Frederick) for the experiment in Fig. 3 and a Beckman/Coulter p24 assay kit (Beckman, FL) for the experiments in Fig. 4B.
RNA extraction and Northern Blot analysis
Total RNA was isolated as previously described [18]. Northern Blot analyses were performed by electrophoresing the isolated total RNA either in a 6% polyacrylamide-7M Urea gel or in a 1. % Agarose formaldehyde gel followed by blotting onto a nylon filter. To simultaneously detect the endogenous U16 snoRNA and the U16RBE RNA, a probe complementary to the 3' end of the U16 was used (Fig. 2A). To detect only the U16RBE a probe complementary to the RBE sequence was used (Fig. 2B). As loading controls probes specific for tRNA3 Lys or the β-Actin were used (Fig. 2).
U16RBE in vivo activity. Some of the CEM clones expressing the U16RBE RNA decoy along with the parental CEM cells were challenged with HIV-1 NL4-3 at an m.o.i of 0.001.CEM clones expressing either a nucleolar anti-HIV-1 hammerhead ribozyme wt (U16Rz wt 11) or the mutant disabled mutant version (U16Rz mutant 4) were used respectively as positive and negative controls for inhibition of HIV-1 infection [18]. On days 1, 10 and 17 day post-infection the supernatants from these cell cultures were collected and assayed for HIV-1 p24Gag accumulation. The data presented represent an average of two independent challenge experiments, and the standard errors for each point are around 5%
CD34+ progenitor cell derived monocytes stably expressing U16RBE are resistant to HIV-1 infection. CD34+ cells isolated from human cord blood were first transduced with either the parental MND/eGFP vector or the MND/eGFP U16RBE and then sorted for eGFP expression. (A) total RNA isolated from the eGFP sorted, transduced CD34+ cells was electrophoresed in a 6% polyacrylamide-7M Urea gel and blotted onto a nylon filter. Hybridizations were performed using a probe complementary to the 3' end of the U16 snoRNA and a probe specific for the tRNA3Lys used as a loading control. (B) Parental CD34+ cells along with the transduced and eGFP sorted CD34+ cells were challenged with HIV-1 JFRL using an m.o.i. of 0.01. On days 1, 7, 14, 21 and 28 days post-infection the accumulation of the viral p24gag antigen was assayed. The data presented represent an average of two independent challenge experiments, and the standard errors for each point are approximately 5%
Nucleolar expression of the Rev Binding Element (RBE) was achieved by substitution of the apical loop of the U16 small nucleolar RNA (snoRNA) with the RBE sequence as previously described [14]. The RBE domain is located within the Rev responsive element (RRE) sequence (within stem-loop IIB, which contains a purine rich "bubble"). Rev has a high binding affinity for this element [11–13]. U16 is a member of the C/D box class of snoRNAs that are primarily involved in post-transcriptional modifications (2' O-methylation) of rRNA molecules [19]. In order to achieve expression of U16RBE in human cells, the chimeric gene was cloned downstream the U6 small nuclear RNA Pol III promoter (clone PTZU16-RBE; [14]). Intracellular expression and nucleolar compartmentalization of the U16RBE RNA in 293 cells has been previously demonstrated [14].
To test the intracellular efficacy of the U16RBE against HIV-1 replication, the PTZU16-RBE expression cassette was inserted into the U3 region of the pBabe puro retroviral vector (clone pBabe/U16RBE, Fig. 1). The resulting vector pBabe/U16RBE was then used to transduce the human CEM T cell line. The U6-U16RBE gene inserted within the U3 region of the retroviral vector should be duplicated within the 5' LTR following reverse transcription and integration. Single clones stably expressing U16RBE were isolated following Puromycin selection. Northern blot analyses performed using total RNA extracted from the selected clones showed somewhat varying levels of expression of the RNA decoy (Fig. 2A).
Transcription of the U16RBE could originate from the U6 promoter as well as from the 5' LTR and internal SV40 polII promoters (Fig. 1B). In order to discern the origins of the major transcripts, a Northern blot analysis was performed on RNA separated in a 1.2% Agarose-Formaldeheyde gel to resolve both long and short transcripts. As shown in Fig. 2B, more than 95% of the U16RBE RNA was of a size corresponding to a U6 promoter-driven transcript. Upon longer exposure (data not presented) transcripts of a size originating from the LTR and SV40 promoters can be seen. These results suggest that the majority of the U16RBE RNAs were derived from the Pol III promoters, with minor amounts originating from the LTR or SV40 internal promoters. In addition, transcripts originating from the Pol II promoters that read through the U16RBE structure may be subject to processing by the snoRNA processing machinery. It is not possible to determine how much, if any processing such as this takes place.
We next tested the anti-HIV-1 activity of the U16RBE decoy by challenging several of the stably transduced CEM clones expressing the U16RBE RNA with the HIV-1 NL4-3 strain using a multiplicity of infection of 0.001. As positive and negative controls for this experiment we also challenged CEM clones expressing either a nucleolar localizing anti-HIV-1 wild type ribozyme (wt) or its disabled mutant version (clones U16Rz wt and mutant; [18]).
At days 1, 10 and 17 post-infection the cell culture supernatants were collected and assayed for the release of the viral p24Gag protein. As shown in Fig. 3, the controls (parental CEM cells and a CEM clonal line expressing the mutant U16Rz) were highly permissive to replication during the 17 days of analysis. In contrast, all the clonal lines expressing U16RBE and the one expressing U16Rz wt were highly resistant to the HIV-1 challenge.
To test the anti-HIV-1 activity of U16RBE in more clinically relevant cells, the PTZU16-RBE cassette was inserted within the U3 region of the 3'LTR of the MND/eGFP retroviral vector (Fig. 1C). This vector allows use of eGFP expression to monitor transduction. The recombinant vector was transduced into human CD34+ progenitor cells purified from umbilical cord blood. Cells were propagated under cell culture conditions that allow differentiation into CD4+ monocytes. Cell counts after pre-stimulation and transduction were similar for both mock and recombinant vectors, with cell viability greater than 90% and cell expansion about two-fold in all cases (data not presented). To assess potential toxicities of the vectors, colony assays were carried out using the transduced CD34+ cells. Although some differences in colony forming numbers were observed between the three samples (Table 1A), in our hands these small differences are not significant, suggesting no overt toxicity of the U16RBE expression.
Table 1 CD34+ cell transduction with MND constructs. (A). 1 × 103 and 2 × 10 3 transduced CD34+ cells transduced with the control or U16RBE expression vectors were plated into 1 ml semi-solid methylcelluose media. After 14 days, colonies were counted and analyzed for Erythroid-Burst Forming Units (BFU-E), Granulocyte Macrophage -Colony Forming Units (CFU-GM) and Granulocyte Erythroid Macrophage Megakaryocyte Colony Forming Units (CFU-GEMM). (B) After transduction CD34+ cells were allowed to expand in stroma flasks, and cell samples were taken for eGFP FACS analysis 1 and 45 days after plating to assess transduction frequency. The loss of eGFP expression with time is a consequence of epigenetic silencing of the MND LTR (Castanotto, Li and Rossi, unpublished results).
The transduction efficiency (TD) was assessed by FACS analyses (monitoring eGFP expression) of the cells at days 1 and 45 post transduction (Table 1B). The TD was between 3 and 6 % of the gated population on day 1 post-transduction as determined by measuring eGFP expression. After 45 days, expression of eGFP dropped to less than 1%. The relatively low level of transduction efficiency is a consequence of the vector preparation and can vary significantly from experiment to experiment. The drop in eGFP expression is due to epigenetic silencing of the LTR driving EGFP in this vector. The Pol III promoter expression does not diminish during this time (H.Li, D. Castanotto and J. Rossi-unpublished observations).
Ten days after transduction the cells were sorted for eGFP expression. The sort purity was checked 5 days later with no significant differences among the various vector transduced cells (data not shown). Northern Blot analyses, carried out on total RNAs isolated from the eGFP sorted CD34+ derived cells, showed readily detectable levels of the U16RBE RNA, which were comparable to the levels of the endogenous U16 snoRNA (Fig. 4A).
The sorted cells were next challenged on the day of FACS analysis with the M-tropic HIV-1 JRFL isolate at a multiplicity of infection of 0.01. On days 1, 7, 14, 21 and 28 post-infection, the supernatants were collected and tested for p24Gag protein accumulation.
As shown in figure 4B, while the mock and the eGFP sorted CD34+ cells transduced with the parental MND/EGFP vector were highly permissive to viral replication, cells expressing the MND/eGFP U16RBE resulted in over 4 logs of inhibition out to 28 days post infection.
We previously exploited the use of the U16 snoRNA as a vector for the nucleolar delivery of therapeutic anti-HIV ribozyme and TAR decoy RNAs [17, 18]. It was previously demonstrated that substitution of the apical loop of U16 with the RBE sequence gave rise to an RNA decoy that localized in the nucleoli of 293 cells and was able to interact with Rev in X. laevis oocytes [14]. In these studies the interaction of the U16RBE decoy with Rev induced the re-localization of the decoy from the nucleus to the cytoplasm. Although we were not able to follow this trafficking in the present experiments, we assume this re-localization also took place in the T- cells and monocytes used in the present studies.
Our primary interest in the present studies was whether or not the U16RBE would sequester Rev sufficiently to inhibit HIV-1 replication in human CEM T-cells and in primary monocytes derived from hematopoietic progenitor CD34+ cells.
To test this possibility we inserted the U6 expression cassette containing the U16RBE sequence within the U3 region of the 3'LTR of two different retroviral vectors, pBabe puro and MND/eGFP (giving rise to pBabe/U16RBE and MND/eGFP U16RBE constructs, Fig. 1). Using the pBabe/U16RBE plasmid we first transduced the CEM T cell line and selected Puromycin resistant clones stably expressing the U16RBE (Fig. 2). We challenged some of these clones with HIV-1 NL4-3 and demonstrated that over a period of 17 days the decoy conferred near complete inhibition of viral replication (Fig. 3). To further test the U16RBE RNA as a possible candidate for hematopoietic stem cell gene therapy for HIV-1, the anti-HIV-1 activity of the U16RBE decoy was tested in human umbilical cord blood CD34+ derived monocytes. The CD34+ cells were transduced with either the MND/eGFP or MND/eGFP U16RBE constructs and allowed to differentiate into monocytes in culture. FACS sorted, transduced monocytes were challenged with HIV-1 JRFL at a multiplicity of infection of 0.01 and the infection was monitored for a period of 28 days (Fig. 4B). While the controls (mock and MND/eGFP transduced CD34+ cells) readily supported viral replication resulting in secretion of μg quantities of p24Gag, the CD34+ derived cells expressing the U16RBE showed four logs of p24 inhibition even after 28 days post infectious challenge (Fig. 4B).
In conclusion, we have demonstrated that the nucleolar localizing U16RBE decoy is a potent inhibitor of the HIV-1 replication. These results suggest that this inhibitor could be a candidate for anti-HIV gene therapy. The decoy could be used alone or in combination with other potent therapeutic RNAs, such as anti-HIV short hairpin RNAs [20]. Rev is essential for HIV-1 structural gene expression and full length viral RNA packaging and consequently sequestering of this protein via the decoy resulted in strong anti-HIV activity. It is highly likely that the decoy-Rev complex traffics to the cytoplasm as was observed in the Xenopus oocytes [14] since Crm1 is the transport carrier in both organisms. Targeting Rev in the nucleolus allows capturing of this protein prior to its interaction with the viral RRE. Furthermore, this strategy is unlikely to result in emergence of viral Rev mutants that can no longer bind the decoy since these would also be defective in binding to the viral RBE. In such a dual component system, mutants in Rev would have to co-evolve with altered forms of the RBE to escape the inhibitory activity of the decoy. Since the U16RBE is encoded in a small gene, it should be easy to combine this with other antiviral RNAs such as shRNAs. A combinatorial approach, which includes the U16RBE, should increase the overall antiviral potency [21] and further minimize the occurrence of viral escape mutants. Although the nucleolar localization of some HIV proteins [23], and perhaps of some classes of HIV RNAs [18], has been demonstrated, the functional role of this cellular localization is poorly understood. Concerning Rev, it has been suggested that the nucleoli could be a "storage site" for the protein [24] and in some cases it has been reported that Rev can function without nucleolar localization [25, 26]. At the same time, it is noteworthy to mention that when Rev is expressed ectopically it localizes primarily within the nucleoli of human cells [2–4] where it multimerizes [10] and relocates CRM1 and some nucleoporins [9]. Rev multimerization is a critical event for the nucleocytoplasmic transport of incompletely spliced and unspliced HIV RNAs, and the association with CRM1 and nucleoporins is probably critical for its own nuclear export. These latter observations underline a crucial role of the nucleolus in Rev function and therefore in the HIV-1 replicative cycle. One possibility is that use of the Crm1 export pathway provides a mechanism for unspliced HIV RNAs to escape the nonsense mediated decay mechanism, although this is only a hypothetical explanation at this time. The importance of nucleolar trafficking of Rev and HIV is strengthened by the data reported here and by previously published data wherein we demonstrated that a nucleolar localizing TAR decoy [17] as well as a nucleolar localizing anti-HIV-1 hammerhead ribozyme [18] are both potent inhibitors of HIV-1 replication.
Kingsman SM, Kingsman AJ: The regulation of human immunodeficiency virus type-1 gene expression. Eur J Biochem. 1996, 240: 491-507. 10.1111/j.1432-1033.1996.0491h.x
Cullen BR, Hauber J, Campbell K, Sodroski JG, Haseltine WA, Rosen CA: Subcellular localization of the human immunodeficiency virus trans-acting art gene product. J Virol. 1988, 62: 2498-2501.
Dundr M, Leno GH, Hammarskjold ML, Rekosh D, Helga-Maria C, Olson M: The roles of nucleolar structure and function in the subcellular location of the HIV-1 Rev protein. J Cell Sci. 1995, 108: 2811-2823.
Stauber R, Gaitanaris GA, Pavlakis GN: Analysis of trafficking of Rev and transdominant Rev proteins in living cells using green fluorescent protein fusions: transdominant Rev blocks the export of Rev from the nucleus to the cytoplasm. Virology. 1995, 213: 439-449. 10.1006/viro.1995.0016
Meyer BE, Malim MH: The HIV-1 Rev trans-activator shuttles between the nucleus and the cytoplasm. Genes Dev. 1994, 8: 1538-1547.
Hope TJ: The ins and outs of HIV Rev. Arch Biochem Biophys. 1999, 365: 186-191. 10.1006/abbi.1999.1207
Fukuda M, Asano S, Nakamura T, Adachi M, Yoshida J, Yanagida M, Nishida E: CRM1 is responsible for intracellular transport mediated by the nuclear export signal. Nature. 1997, 390: 308-311. 10.1038/36894
Fornerod M, Ohno M, Yoshida M, Mattaj IW: CRM1 is an export receptor for leucine-rich nuclear export signals. Cell. 1997, 90: 1051-1060. 10.1016/S0092-8674(00)80371-2
Zolotukhin AS, anf Felber BK: Nucleoporins nup98 and nup214 participate in nuclear export of human immunodeficiency virus type 1 Rev. J Virol. 1999, 73: 120-127.
Daelemans D, Costes SV, Cho EH, Erwin-Cohen RA, Lockett S, Pavlakis GN: In vivo HIV-1 Rev multimerization in the nucleolus and cytoplasm identified by fluorescence resonance energy transfer. J Biol Chem. 2004, 279: 50167-50175. 10.1074/jbc.M407713200
Bartel DP, Zapp ML, Green MR, Szostak JW: HIV-1 Rev regulation involves recognition of non-Watson-Crick base pairs in viral RNA. Cell. 1991, 67: 529-536. 10.1016/0092-8674(91)90527-6
Heaphy S, Finch JT, Gait MJ, Karn J, Singh M: Human immunodeficiency virus type 1 regulator of virion expression, rev, forms nucleoprotein filaments after binding to a purine-rich "bubble" located within the rev-responsive region of viral mRNAs. Proc Natl Acad Sci U S A. 1991, 88: 7366-7370. 10.1073/pnas.88.16.7366
Tiley LS, Malim MH, Tewary HK, Stockley PG, Cullen BR: Identification of a high-affinity RNA-binding site for the human immunodeficiency virus type 1 Rev protein. Proc Natl Acad Sci U S A. 1992, 89: 758-762. 10.1073/pnas.89.2.758
Buonuomo SB, Michienzi A, De Angelis FG, Bozzoni I: The Rev protein is able to transport to the cytoplasm small nucleolar RNAs containing a Rev binding element. RNA. 1999, 5: 993-1002. 10.1017/S1355838299990064
Fragapane P, Prislei S, Michienzi A, Caffarelli E, Bozzoni I: A novel small nucleolar RNA (U16) is encoded inside a ribosomal protein intron and originates by processing of the pre-mRNA. EMBO J. 1993, 12: 2921-2928.
Good PD, Krikos AJ, Li SXL, Bertrand E, Lee NS, Giver L, Ellington A, Zaia JA, Rossi JJ, Engleke DR: Expression of small, therapeutic RNAs in human cell nuclei. Gene Ther. 1997, 4: 45-54. 10.1038/sj.gt.3300354
Michienzi A, Li S, Zaia JA, Rossi JJ: A nucleolar TAR decoy inhibitor of HIV-1 replication. Proc Natl Acad Sci U S A. 2002, 99: 14047-14052. 10.1073/pnas.212229599
Michienzi A, Cagnon L, Bahner I, Rossi JJ: Ribozyme-mediated inhibition of HIV 1 suggests nucleolar trafficking of HIV-1 RNA. Proc Natl Acad Sci U S A. 2000, 97: 8955-8960. 10.1073/pnas.97.16.8955
Kiss T: Small nucleolar RNAs: an abundant group of noncoding RNAs with diverse cellular functions. Cell. 2002, 109: 145-148. 10.1016/S0092-8674(02)00718-3
Hannon GJ, Rossi JJ: Unlocking the potential of the human genome with RNA interference. Nature. 2004, 431: 371-378. 10.1038/nature02870
Li MJ, Bauer G, Michienzi A, Yee JK, Lee NS, Kim J, Li A, Castanotto D, Zaia JA, Rossi JJ: Inhibition of HIV-1 infection by lentiviral vectors expressing Pol III-promoted anti-HIV RNAs. Mol Ther. 2003, 8: 196-206. 10.1016/S1525-0016(03)00165-5
Morgestern JP, Land H: Advanced mammalian gene transfer: high titre retroviral vectors with multiple drug selection markers and a complementary helper-free packaging cell line. Nucleic Acids Res. 1990, 18: 3587-3596.
Olson MOJ, Dundr M: The moving parts of the nucleolus. Histochem Cell Biol. 2005, 123: 203-216. 10.1007/s00418-005-0754-9
Kjems J, Askjaer P: Rev protein and its cellular partners. Adv Pharmacol. 2000, 48: 251-298.
McDonald D, Hope TJ, Parslow TG: Posttranscriptional regulation by the human immunodeficiency virus type 1 Rev and human T-cell leukemia virus type I Rex proteins through a heterologous RNA binding site. J Virol. 1992, 66: 7232-72.
Wolff H, Hadian K, Ziegler M, Weierich G, Kramer-Hammerle S, Kleinschmidt A, Erfle V, Brack-Werner R: Analysis of the influence of subcellular localization of the HIV Rev protein on the Rev-dependent gene expression by multi-fluorescence live-cell imaging. Experimental Cell Research. 2006, 312: 443-456.
This work was supported by a MURST (PRIN and "Centro di eccellenza BEMM") and ISS (Programma Nazionale di ricerca sull'AIDS- progetto n. 40D.17) grants to IB and NIH NIAID 42552 and 29329 to JJR. The authors would like to acknowledge the contributions of L. Cagnon and J. Kim for assisting with the HIV assays.
Division of Molecular Biology, Beckman Research Institute of the City of Hope, 1450 East Duarte Rd., Duarte, California, 91010, USA
Alessandro Michienzi
& John J Rossi
Istituto Pasteur, Fondazione Cenci-Bolognetti, Department of Genetics and Molecular Biology, University of Rome La Sapienza and IBPM of CNR, Rome, Italy
Fernanda G De Angelis
& Irene Bozzoni
Present address: Istituto Superiore di Sanita', Department of Cell Biology and Neuroscience, Viale Regina Elena 299, Rome, 00161, Italy
Division of Molecular Biology, Graduate School of Biological Sciences, Beckman Research Institute of the City of Hope, 1450 E. Duarte Rd., Duarte, CA, 91010, USA
John J Rossi
Search for Alessandro Michienzi in:
Search for Fernanda G De Angelis in:
Search for Irene Bozzoni in:
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Correspondence to John J Rossi.
Michienzi, A., De Angelis, F.G., Bozzoni, I. et al. A nucleolar localizing Rev binding element inhibits HIV replication. AIDS Res Ther 3, 13 (2006) doi:10.1186/1742-6405-3-13
Long Terminal Repeat
eGFP Expression
Human Immunodeficiency Virus Replication
Colony Form Unit Assay
Normal Human Bone Marrow

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This entry was posted in Humor, Love, Marriage, Men, Romance and tagged humor, love, marriage, relationships, wife, women by Belle of the Library. Bookmark the permalink.
Ugh, MEN!!!! Mine is like that too, except he still doesn't know how to reach the apology or accepting responsibility for anything stage. Actually it's caused me to consider if we are really going to make it long-term. He's generally a good guy but in any disagreement it's all gaslighting and projecting, never anything else. And he doesn't see that that's even happening, though I've explained it to him. Seems like an important thing to need to be able to get past.
We've both improved our communication skills a great deal, in the four years we've been together. The first time I ever got frustrated with him, he genuinely apologized and made a sincere effort not to do it again.
It's different, now that we're married, but if I give him some time to let his ego deflate, he's usually pretty apologetic. I agree that this is sort of a make or break in a relationship. If he were incapable of humility, I doubt we'd have made it to marriage.


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Review | Star Wars: The Rise Of Skywalker
December 18, 2019 Carl Roberts	Leave a comment
Star Wars: The Rise Of Skywalker is a thrilling rollercoaster ride that will impact on the emotions of the fans without really giving them the ending they want.
Ok, here we go.
After 42 years, eight-episode films, two standalone movies, several animated shows, a cringe-worthy holiday special and a fantastic new live-action TV show, we finally come to the ninth and final instalment in the Skywalker saga, 'The Rise Of Skywalker'. As I stated before in my pre-review article, I saw the film for the first time at a press screening in London on Tuesday night and didn't quite know what to make of it. I said it would take a few viewings to get my head around the film as a whole and it has. It will divide the fanbase again but not to the level of 'The Last Jedi'. It does have some problems with it as a whole which the hardcore fans will take issue with. It is, however, like 'Revenge Of The Sith' before it, the best of the new trilogy of films. Will it be proclaimed as the best Star Wars film of them all? Not a chance, you'll never be able to top 'The Empire Strikes Back' in that department. Will it be in the top five Star Wars films of all time? Debatable. It is though, a thrilling rollercoaster ride that will impact on the emotions of the fans without really giving them the ending they want. It gives us an ending but as it's been said before, 'This won't go the way you think'.
Star Wars: The Rise Of Skywalker | Lucasfilm/Walt Disney Pictures
We open as always with the familiar logo and the blast of the fanfare that has encompassed the saga ever since it blasted onto screens back in 1977. John Williams gives the opening the energy and pomp that it deserves and what we have come to expect. After the familiar crawl, we are thrust straight into the action as Kylo Ren is on the hunt for the source of The Emperor's transmission, only found by using a Sith Wayfinder, a kind of map embedded in a Holocron. This is a nice touch and a nod to the stories seen in the animated series, 'Star Wars: Rebels'. Ren tracks the signal to the planet Exogol, a world not listed on any conventional star chart. Landing his ship and hunting through the temple he finds, Ren is confronted with an astonishing truth. It is here we hear *SPOILER ALERT* the voices of both Snoke AND Vader. This has been revealed before in a clip released before the film was released but is still a joy to be heard. And now we meet and see The Emperor once more. My problem with Palpatine's return is quite simple. We are never told HOW he survived his fate in 'Return Of The Jedi'. We can form our own conclusions with what we hear from him during the film's events but it is never clear. Palpatine gives Kylo Ren a new mission as Supreme Leader and offscreen, reveals everything about Rey, something we will find out for ourselves later in the film.
Next, we are with Poe, Finn, Chewbacca and R2-D2 aboard the Millenium Falcon as they speed through hyperspace in their mission to acquire data transmitted by a First Order spy. This goes sideways and we are treated to a thrilling chase sequence between the Falcon and pursuing TIE Fighters. All this is punctuated by Williams' score. We now move on to find Rey in the middle of her Jedi training, attempting to commune with the spirits of the Jedi from the past and failing. Rey then decides to run the assault course she is required to do as part of her training, chased by a remote droid in a nice throwback to Luke's first training aboard the Falcon in 'A New Hope'. However, Rey is distracted by the feeling of Kylo Ren and fails the course miserably. The Falcon returns and the data contents are revealed, giving the Resistance a countdown before their final destruction is commenced.
To go any further into details about the film and its story will tread heavily into spoiler territory and is something I refuse to do. The film DEMANDS to be seen on the big screen and the audience deserves to go into it CLEAN. If you are planning to see the film, I suggest staying as far away from the internet and in particular, social media sites as, sadly, there are many people out there determined to spoil the joy and surprises that the film contains. The shocks and surprises (and there are quite a few of each) deserve to be discovered during the film's 142-minute runtime.
Okay, lets us get the negatives out of the way first. The story really feels like it has been thrown together. Some of the threads and seeds that were sown in 'The Force Awakens' and 'The Last Jedi' are completely overlooked and abandoned. It's kind of a slapdash approach in respect to the film's writing and one that, on first viewing, I had a problem with. The problem does diminish somewhat on a second viewing but the problems still remain. It is common knowledge that the script was almost completely rewritten after Colin Trevorrow's departure from the film but Chris Terrio and J J Abrams' screenplay has some major flaws within its pages and this is reflected during the course of the film.
Again, many characters are sidelined and this is not exclusive to the classic characters. Many of the new or newer characters have barely enough screentime to shine or be given enough to do. Kelly Marie Tran's character of Rose for example. In the previous film, she was shown as a tough, determined young woman. Here, she is relegated to minor player status, a slap in the face to the young actress after all the turmoil she faced from the internet trolls after the last film. Her role in the film is so limited that if she didn't appear at all, we wouldn't really notice. And that is completely unfair to Ms Tran. Likewise Keri Russell, a newcomer to the franchise as 'Zorri Bliss'. Her role and appearance are similar to Zam Wesell from 'Attack Of The Clones'. Her screen time is around the same too. R2-D2 again is pushed into the shadows as is Chewbacca himself. It is always going to be hard to juggle enough screentime for all the characters that the franchise contains but some are almost ignored completely.
It pains me to say that John Williams' score for the film, a staple and something that we all hold dear is considerably lacking this final time around. Of course, we get the blasts of the classic themes as the film deserves but they are few and far between. Classic motifs such as The Imperial March get a brief look in during proceedings but a fair part of the score contains the Emperor's theme, which is haunting as it should be. However, surrounding these welcome and expected themes, we have a score that hardly suits the film. The new music is disappointing in general. It was reported that Williams wrote 135 minutes worth of new music for the film. I can report that, apart from around three minutes during the course of the film and part of the end credits montage that brings a huge smile to the viewers face, the rest of the score is a letdown. It is a score that lets the film down and that is one of the biggest shames the film could have offered us.
Now for the positives, which I am relieved to say are many. Not all can be reported here for reasons that will become clear during the film but when you see them, you will know what I mean. The late Carrie Fisher shines in her limited role in the film, giving us a sad thought of what might have been if our beloved Princess was still with us in the real world. Her appearance is tactful and handled with huge sensitivity and Abrams must be given huge credit for how he handles such a delicate matter. Mark Hamill, in what is really only a cameo role but thankfully has dialogue and longer screentime than 'The Force Awakens', is a delight as the now Force ghost Luke Skywalker. His appearance gained a massive cheer at both screenings I attended and is completely deserved. The acting is once again pretty top-notch and shows what a great future Daisy Ridley and John Boyega have in front of them post-Star Wars. Adam Driver this time around plays Ren less whiny than we have seen him before and is a welcome addition this time. The special effects, though I did notice one that was a bit under-par, is once again a marvel to behold, The Falcon chase near the start of the film is a standout. The locations are beautifully presented to us in all their colourful glory and the production design is once again, without reproach. But apart from things I cannot and WILL not mention, the main thrill came when we finally see the great Billy Dee Williams return to the screen once again as Lando Calrissian. Finally, the fourth and final part of the wheel comes back into play, after the returns of Luke, Leia and Han in the previous movies. A really large cheer was reserved pacifically for the first screen appearance of Lando in the film and is totally deserved. Williams slips on the persona of Lando as easily as one of Lando's cloaks slips on. It is effortless.
Some of the surprises the film contains that I won't reveal top everything. The viewer will need to keep their eyes and ears out for everything throughout the events of the film from the start right up to the end credits. The thrill and excitement and knowing smiles will flow naturally from the audience when these surprises happen.
Is the film the perfect ending to the Skywalker saga? No, It's not. There are certain elements and plotlines that are lacking to make the film the completely satisfying ending to a journey that started 42 years ago. However, what we are given is good old fashioned entertainment, a throwback to sitting in a movie screen many years ago and marvelling at a new kind of science fiction movie, the likes of which somehow wormed its way into the hearts and minds of audiences the world over. Did I shed a tear or two at the end? I'm not ashamed to say, yes I did. It wasn't for what I saw on the screen (though the final shot helped) but for the finality of something that I grabbed as a seven-year-old child, Something that has been with me for the years since until now, as an adult, I viewed the final act, the curtain call, the swan song of something that has been with me almost all of my life.
In 1983, we all thought it was over. In 2005, we felt the same way again.
Now, in 2019, we can say goodbye to what is, in essence, the greatest family saga of all time, once and for all. The Star Wars saga as we know it is over. We will never see the likes of it again, most certainly not in my lifetime. As I exited the theatre as the final strains of the end credits music faded, I looked back at the screen one final time and wished a fond farewell to an old friend. Was it a perfect goodbye? No. But make no bones about it, this IS the final goodbye. The saga may be over but the legend will live forever.
'The Force will be with you, always'
Maybe not. But the legend of 'Star Wars' always will be.
Star Wars: The Rise Of Skywalker is distributed by Walt Disney Pictures and is playing in cinemas NOW!
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Carl Roberts is a Senior Staff Writer and Books and Literature Correspondent for Future of the Force. He is passionate about Star Wars, Marvel, DC, Indiana Jones and Horror movies. Follow him on Twitter @CarlRoberts2 where he uses the force frequently!
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Kalwaria Zebrzydowska - the Mannierist Architectural and Park Landscape Complex and Piligrimage Park placed 50 km from Cracow.
One of the most beautiful sanctuaries In Europe. It contains Basilica of Our Lady of the Angels which is late baroque building with elements of rococo. Interior is decorated with polychrome, stalls, rich inlaid furniture and numerous paintings. Monastery is three-storied building located around two patios. Interior is characterized by rococo and baroque style and adorned with the stucco decorations, portraits of founders and benefactors. System of paths known as Piligrimage Park consisting of a group of churches and chapels built in baroque and mannerism style. Complex is built In the midst of natural landscape. Park is inspired by the descriptions pf the Holy Land.
Our driver will pick you up from the hotel, apartment or any other place suitable for you straight to Kalwaria. We will be waiting for you, as long as you wish. After seeing Kalwaria our driver will take you back to Krakow, wherever you wish.


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Search and Recover es el quinto episodio de la tercera temporada y vigésimo quinto episodio a lo largo de la serie de drama y ciencia ficción de TNT, Falling Skies; fue escrito por Jordan Rosenberg y dirigido por Sergio Mimica-Gezzan y salió al aire el 30 de junio de 2013 en Estados Unidos.

Tom y Pope deben confiar el uno en el otro para sobrevivir después del accidente que sufrieron. Mientras tanto, en Charleston, Weaver y compañía buscan desesperadamente a dos miembros de la 2nd Mass que han desaparecido, y Marina toma el asunto en sus propias manos sin tener en cuenta la autoridad de Tom.

Argumento
Tom despierta después de que el avión donde viajaba se estrellara en el bosque y se encuentra con que al General Bressler lo ha atravesado una de las ramas de un árbol en el corazón y a Pope inconsciente en la parte posterior. Tom escucha a una nave acercarse y con trabajo logra sacar a Pope y poner a salvo a ambos, inmediatamente después, la aeronave explota con el General Bressler dentro. Mientras tanto, en Charleston, Ben le informa a Hal y a Maggie que Anne y Alexis han desaparecido. Poco después, se reúnen con Matt, el Coronel Weaver y Marina para escuchar lo que el Dr. Kadar y Lourdes sobre lo ocurrido con Anne y Kadar les revela que Alexis es un híbrido. Weaver insiste en formar un equipo de búsqueda y rescate, sin embargo, Marina, en su papel de vicepresidente, le niega la autorización para disponer de los militares, entonces Weaver le dice que formará un equipo con los Mason, a lo que Marina no se niega.

Pope y Tom pelean por lo que deben hacer cuando unas naves llegan a inspeccionar el lugar. Tom insiste en que deben moverse mientras Pope dice que deben quedarse; Tom comienza a andar cuando Pope lo derriba justo antes que una nave pueda verlo, entonces, Tom decide quedarse también. Mientras tanto, Weaver, Hal, Ben, Matt, Maggie y Jeanne salen en busca de Anne y Alexis y se encuentran con una mujer muerta por la que Matt se compadece y sugiere sepultarla, a lo que todos acceden, a excepción de Hal que se muestra un poco receloso con todo lo relacionado con la búsqueda. Por otra parte, Marina acude al Dr. Kadar y le pide que investigue para qué sirve realmente el arma que los Volm están construyendo mediante unas fotografías de la misma.

Pope le revela a Tom el motivo por el cual dejó de ver a sus hijos, cinco años antes de la invasión extraterrestre: después de que su hijo Brandon y él arreglaban la motocicleta, el chico dio un paseo pero estuvo a punto de ser atropellado por un hombre que manejaba inconscientemente, entonces Pope, queriendo darle una lección al hombre, lo golpea, provocando que su cabeza "rebotara como pelota en el pavimento", según las palabras de Pope, causándole la muerte. Por su parte, Tom le cuenta que su juventud no fue fácil debido a que su padre era un hombre difícil y que lo único que le alegra de su juventud es haber sobrevivido a ella.

Mientras continúan la búsqueda de Anne y Alexis, Jeanne le pide a su padre que nunca la deje caer en las manos de los Skitters y Maggie nota que Hal tiene una extraña actitud cuando descubren unas huellas de Skitters, sin embargo, Weaver decide que es tiempo de regresar. Mientras tanto, Tom y Pope se enfrascan en una pelea que termina cuando descubren la presencia de unos Skitters que están en busca de algún sobreviviente del accidente aéreo. Una de las criaturas se da cuenta de la presencia de los humanos y los ataca pero Tom le dispara atrayendo la atención del resto de ellos, por lo que se ven obligados a correr y saltan al agua. Cuando llegan al otro extremo del río, Tom y Pope continúan con la pelea pero el profesor Mason resulta lastimado de un pie.

En Charleston, tras haber encontrado a la mujer muerta, Matt le revela a Ben que no desea morir solo como ella. Ben le asegura que eso no sucederá. Pope y Tom encuentran un refugio para poder entablillar el pie del profesor. Pope insiste en seguir, pero Tom desea quedarse pues sigue sin poder apoyar su pie lastimado por lo que se enfrascan en una nueva discusión que termina cuando Tom le asegura a Pope que es un cobarde y que tiene miedo de seguir solo. Pope se marcha sin decir nada, pero regresa después cuando unos Skitters encuentran a Tom y le dice que "lo único que no puede soportar además de él, son a los Skitters" y lo lleva hasta una cabaña donde encontró una camioneta abandonada que los ayudará a regresar a Charleston.

Finalmente, Pope y Tom llegan a Charleston y se desmayan en la entrada. Marina regresa con el Dr. Kadar para saber si ya tiene alguna respuesta a su solicitud y Kadar le revela que el arma de los Volm no es lo que dicen que es. Dos días después, Tom despierta en el hospital con Pope a su lado y después se encuentra con su familia y Maggie y el coronel Weaver, cuando nota la ausencia de Anne y Alexis pregunta por ellas por lo que Weaver le revela que han desaparecido.

Elenco

Personajes principales
 Noah Wyle como Tom Mason.
 Drew Roy como Hal Mason.
 Connor Jessup como Ben Mason.
 Maxim Knight como Matt Mason.
 Seychelle Gabriel como Lourdes Delgado.
 Sarah Sanguin Carter como Maggie.
 Colin Cunningham como John Pope.
 Will Patton como Coronel Weaver.

Personajes recurrentes
 Laci Mailey como Jeanne Waver.

Estrellas invitadas
 Gloria Reuben como Marina Perlata.
 Matt Frewer como el General Bressler.
 Robert Sean Leonard como Roger Kadar.

Continuidad
 El episodio comienza la misma noche de la desaparición de Anne y Alexis y momentos después del accidente del avión en donde viajaban Tom, Pope y el General Bressler.
 El General Bressler muere en este episodio.
 Pope revela a Tom el nombre de sus hijos y la razón por la que no los ha visto desde cinco años antes de la invasión.
 El Dr. Kadar realiza pruebas de ADN a los bebés recién nacidos en Charleston para saber si hay más híbridos como Alexis.

Producción
El episodio está basado en la película de suspense de 1958, The Defiant Ones.

A partir de este episodio, Moon Bloodgood se ausenta temporalmente de la serie y la trama debido a que la actriz se encontraba embarazada en ese entonces. En un reportaje exclusivo con TV Line sobre los acontecimientos venideros en la trama, Remi Aubuchon, uno de los productores ejecutivos de la serie, habló al respecto de la decisión tomada en torno a la situación:

Recepción

Recepción de la crítica
Chris Carabott de ING califica al episodio de satisfactorio y le da una puntuación de 6.0, diciendo: "Search and Recover era una muy buena oportunidad de evolucionar la relación inestable entre Pope y Tom Mason pero se desperdició por el melodrama innecesario, lo que conduce finalmente a que los dos personajes terminen exactamente en el mismo lugar en el que comenzaron. La búsqueda de Anne Glass produce pocos resultados también. Aunque nos hacemos una idea de la situación emocional de algunos personajes, no es nada que no se podría haber conjeturado ya por nuestra cuenta. El único relato concreto es el desarrollo de momentos que vienen en la forma de la muerte de Bressler y los motivos potencialmente siniestros de Marina".

Recepción del público
En Estados Unidos, Search and Recover fue visto por 3.21 millones de espectadores, de acuerdo con Nielsen Media Research.

Véase también
 Falling Skies
 Tercera temporada
 Lista de episodios
 Personajes de Falling Skies

Referencias

Enlaces externos
 Página web oficial (en inglés).
 Página web oficial (Latinoamérica).
 Sitio oficial  (España)

Episodios de la tercera temporada de Falling Skies

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Say it like you mean it in this dope Thank you for Nothing Dad Hat in white for men. This cap is great to wear with any outfit for effortless style either during the week or on the weekends. Details include Thank you for Nothing embroidered on the front, adjustable back strap with clasp, duck bill front is 2.5" and eyelets on top.


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Dickens's 1839 account book shows that he, rather than Catherine, paid their servants' wages. This was an unusual level of involvement in domestic affairs for a male head of the household.
Account book, 1839.
AA26. © Charles Dickens Museum
Purchased with support from the National Heritage Monument Trust, the Art Fund, Friends of the National Libraries and the Dickens Fellowship.
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A Conversation with Lois Main Templeton
with Lois Main Templeton, Contemporary Artist
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https://comcastnewsmakers.com/Videos/2018/6/15/A-Conversation-with-Lois-Main-Templeton
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Born in 1928, artist Lois Main Templeton experienced a Wisconsin childhood. Painting only arrived in the 1970's when, she was living near San Francisco, the materials themselves called out and her fingers itched to get a hold of them. The high horizons and rimrock, typical of the West, can still be the first thing to climb on her canvases. Comcast Newsmakers Field Reporter Talor Whitaker sits down with Lois from inside her very own exhibit at the Indiana State Museum for a thorough discussion on her life and tenure within the artist community.
Hosted by: Talor Whitaker Produced by: Heartland Newsmakers Team
#Indiana #General Interest #Local #Community
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Making House a Home: The Evolution of Tinker House Events
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Skyros - Geographic co-ordinates and position
The island of Skyros, "the heart of the Aegean Sea", is the bigger island of North Sporades having an extent of 215 square kilometres and its resident population is about 3.500 people.
The island is halfway or so from the east coasts of Euboea, in a distance of 23 miles north-east from Kymi and 26 miles south from Alonnessos.
Skyros belongs, for administrative purposes, to the Prefecture of Euboea. It participates in the Prefectural Council having three elected representatives, who constitute the provincial council of the island.
Skyros, as a whole, constitutes a Municipality, which is geographically identified with the County of Skyros, the responsibilities of which are different from those of a Municipality.
There are a lot of smaller islands in the surrounding area. Some of these small islets are used as pasturelands, while this medley of land and sea is a marvellous fishery.
We could mention several islands like Skyropoula, Sarakiniko or the Island of the Despot, Renea, Plateia and Koulouri. There are also a lot of desert islands as well as some rocky islands such as Vrahonissia in Achilli, St. Armolaos, Diavates, Aloni, Myrmigonissia, Cameles, Vrykolakonissia etc.
The island of Skyros consists of two massifs; between these massifs, there is a level ground that stretches for 3,5 kilometres and is 2,5 kilometres in width.
This level ground, which once was sunk in the sea, begins from Achilli and goes up to Kalamitsa, dividing the island in the north-western and south-eastern parts. The north-western part of Skyros is relatively level and fertile, with spring sources, a profuse vegetation and it is full of pinewoods.
On the contrary, the south-eastern part of the island is mountainous, stony and barren. The people of Skyros call this part of the island "the Mountain", not for its altitude, but because it is completely stony.


H.E. Ambassador Genaro V. Pappalardo's letters of credence presentation on 27/06/2017 to H.M. Queen Elizabeth II.
Jesuit Mission of Jesus & Trinidad, founded in 1685.
Train Station of Sapucai, 85 kms from Asunción, founded in 1910.
Galopera – Traditional Paraguayan Dance.
Representation of Kambá Ra'angá in Altos.
Festivity of Saint Francisco Solano on 24th July.
The Embassy of the Republic of Paraguay to the United Kingdom of Great Britain and Northern Ireland welcomes you to its official website, hoping that this will be an efficient mean of information and a useful channel of communication, where visitors can access clear and concise information about the multiple services that are carried out by the Embassy and the Consulate.


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Posted on October 11, 2016 by FugueBurg
(Originally posted December 2011 at Cinemachine)
A nighttime Halloween party. As children laugh, run and play in their costumes, one child clad in ghost sheets invites another boy and girl to come see the "bag of jewels" he's found in the nearby forest. We're shown via flashback that this gullible couple are bullies, and the ghost kid was apparently the target of their cruelty. As the boy bully nears a bag placed precariously next to some sort of ditch, ghost kid sneaks up from behind and gently pushes the tricked bully down the hole and out of sight.
Abruptly, the swelling musical score to this scene cuts off in mid-note and flashing lightning accompanies two shots alternating back and forth: a moving teddy bear with glowing eyes – and a deep hole full of some shifting, hairy…things. The movie's title is superimposed, fading from yellow to red: THE PIT.
Persons experiencing confusion at this obtuse introduction, take heart. The film is, indeed, somewhat incompetently made and totally unsure of itself. There is a kid, and a pit full of things, and a teddy bear, but anyone seeking reason or answers for them will be sorely disappointed.
The Pit is far and away one of the most baffling and intriguing titles from the 80s independent exploitation era; an adolescent revenge fantasy (not uncommon to the genre) by way of fantastical fairy tale (there are monsters and a living doll) filtered through the uncomfortably sleazy eye of a hacky director whose primary concern seemed to be including gratuitous nudity, or to at least show his actresses doing aerobics or jogging, as often as possible.
Jamie (Sammy Snyders, fresh from playing Tom Sawyer on the TV series Huckleberry Finn and His Friends) is a young boy who doesn't fit in. He also has an active imagination – not only having conversations with his teddy bear, but also a deep dark pit full of monsters in the local woods. Or are they real? At first the viewer rationalizes that both aberrations are figments of a troubled boy's imagination, until the film shows Jamie taking revenge on local bullies and troublesome adults by tricking them into the pit where they become the creatures' lunch. Meanwhile, Jamie's babysitter Sandy (Jeannie Elias, soon to become a successful voice actress in animated TV shows like The Super Mario Bros. Super Show) is the only adult watching him while his parents are out of town – and in a recipe for disaster, she's Jamie's first crush.
Sammy Snyders as Jamie in The Pit
The monsters at the bottom of the pit are believed by Jamie to be prehistoric troglodytes. These trogs are played by little people in fur costumes with immobile pig-like masks, and they're filmed with barely any regard to covering up the obvious shortcomings of cheap makeup special effects. The burden of their effectiveness is left to the sound effects of their continual roars (which sound more like the MGM lion than primordial beasts) and the bombastic strings and horns of the musical score by Victor Davies.
Jamie's teddy bear "Teddy" speaks to him in Sammy Snyders' voice with an echo added, and Teddy seems to know a lot more about Jamie's stirrings of manhood than he does. Shoving people down the pit to feed the trogs is also Teddy's idea. What's more, Teddy is indeed alive. This is only confirmed by his glowing eyes at the opening title, and another shot twenty minutes into the film where the bear's head turns a few degrees and Victor Davies lays the music on extra thick to sell the shock.
"Teddy" as himself
Compounding the strange nature of all these story details is a wildly inconsistent tone by the director that veers back and forth from bombastic shock to daft whimsy. Which constantly begs the question, for what type of horror fan was the film intended? Even amongst the few examples of Bad Seed or Village of the Damned type horror films about mischievously murderous tots from this era, there are none so unique as this hallucinatory tale involving trogs, teddy bears and hormonal stirrings directed toward babysitters.
My personal interest in The Pit began from reading a review published on the long running cult film website BadMovies.org. Like those reading about the film for the first time at this moment, my initial response to descriptions of such a strange film was incredulous. Knowing I'd need to see the movie for myself to believe it, I kept an eye out until eventually happening across the VHS. Sitting down with a likeminded fan of the cinema du bizarro, the effect was even more bewildering than we expected. How was this cheesy horror film ever constructed around such a singularly surreal story?
Many years later in a Toronto used bookstore, I happened upon the spine of a paperback entitled Teddy, by John Gault. Taking a look at the cover to pass judgment I was struck by the illustration of a child holding a teddy bear with demonic lights shining from its eyes, and a very familiar tagline. Then I looked at the back cover and nearly fainted – who else but Teddy's unforgettable askew-eyed face was staring back at me?
Teddy, the novel by John Gault based on Ian A. Stuart's screenplay
Buying and reading the novel immediately, I found that John Gault's version of the story "based on an original screenplay" by Ian A. Stuart was an astonishingly different take on the same material. While the characters and events were all essentially the same, Gault set a tone of genuine eerie dread that the slapdash film never accomplished, as well as giving some much needed additional focus on the relationship between young Jamie and his evil toy. Gault's novel Teddy was The Pit as a deadly serious horror story, and the disparity between the two made me all the more curious as to what went awry in the making of what seemed to have been originally intended as a serious horror film.
The novel was published in Toronto. Living there at the time, I attempted to locate Mr. John Gault in the hope that he could shed some light on the secret history of The Pit, having worked on his novel from the original screenplay. Unexpectedly, my efforts led to the contact information of the original screenwriter himself, Mr. Ian A. Stuart – whose initial vision was not only meant to be a serious chiller, but one that would have given the sexual undercurrent a very different context and ended with a twist explaining the fantastic elements of trogs and living dolls.
Mr. Stuart was kind enough to grant the following interview.
How did The Pit get produced?
The idea was originally to produce a Canadian-made horror picture on a relatively low budget with a simple story. I had written such a script, and it was purchased by Amulet Pictures. It was originally called Teddy, but marketed as The Pit because that sounded more dramatic for a horror picture. The producer was Bennet Fode and the executive producer was the late Johnny F. Bassett, who used to own the Toronto Telegram. He was from the Bassett family who controlled the CTV television empire and he was in the film business for a short while as an executive producer.
Just as we were finished putting the pieces of the puzzle together to get the picture ready for production, either Bennet or Johnny Bassett hired Lew Lehman to direct. He was an American director – they were going to shoot in Wisconsin – so he was hired to do the job. Bassett later admitted that they had never seen anything he'd done previously.
Variety ad for distribution under the original title Teddy
How did your original screenplay differ from the film?
It was never meant to be funny – except that we have a tendency to laugh at children who do amusing things. Jamie has a rather dark imagination, he's discovered a huge hole in the ground in the forest at the bottom of which live these things. He doesn't know what to call them but he's heard about cave dwelling early human beings called Troglodytes and mispronounces the word as "troglodies" – not the "tra-la-logs" we heard in the film, which was downright silly. He believes in their reality, and he tells the babysitter who's living with him about them. She of course doesn't believe him. Then as he becomes more insistent that they're real, she becomes more and more annoyed until one day she slaps him in the face, saying he has to stop this nonsense.
From that moment to the end of the film, nothing the audience sees is really happening. It's all in his mind. There's a hole in the forest, but there's nothing at the bottom of it. By slapping his face she's cut herself off from him. The creatures in his imagination dispose of her. She falls into the pit accidentally, at least in his mind, because he couldn't have pushed her. That would be an act of agression. It had to be an accident.
Only at the end of the film do you see a psychiatrist saying, "This boy is very disturbed." The camera pans over and wait a minute, there's the babysitter. She's not dead, this has all been in his mind. So he's taken out into the country to the grandparents, where he meets a little girl and she takes him out in the woods and says "Look what I found, a big hole in the ground where these little creatures live." He says they're troglodies, and she says "I know," and push! He goes down the hole. The audience should walk out of the theater wondering, what just happened there? Was that real or unreal? He gets his just desserts, but in whose mind did it take place?
The traditional structure of a horror story isn't the novel, it's the short story that begins very realistically, introduces an element of the fantastic, and then has to wrap up relatively quickly. But the wrap-up is in the last phrase, the last sentence, even the last word, where the story really gives you a jolt. Jamie's institutionalized, goes out into the woods, meets this other child just like him who says "I want to show you something, I found a big hole in the ground and look what's at the bottom, troglodies!" He says they eat people, and she says "Yes, I know." Push! It's all in the last word. "Push."
the freeze-framed final shot
What was the initial inspiration for the story?
From a literary genre point of view, the story is what you'd call the "Demon Child" story – an apparently innocent child who is actually demonic. That story has been told many times. This scenario came from a very real incident of a child's mental illness, described to me by a friend who's a child psychologist and knew a boy who'd draw creatures like Jamie's "troglodies" that he could send after people who did harm to him. This friend also told me, "I've had to sign commitment orders for children who are 8, 9 and 10 years old, who are not really children. They're little balls of hate and fury. And the only reason they haven't killed somebody yet is they're not big enough and strong enough. But, someday. Unless you deal with that problem, you have that next murderer, next rapist. That child who's full of hate and fury is going to react violently against the world."
I also had a ventriloquist friend who used to talk to an autistic boy using his dummy, the kid would talk to the dummy, but as far as he was concerned, my friend didn't exist. Putting those children's perceptions together gave me the idea for Jamie's particular circumstances in the film. John Bassett actually had the script read by another child psychiatrist who said it was the best depiction he had ever read of the mind a psychotic child. Then, after the film was made he came to me and said "I'm sorry, Ian. You wrote a great script, and all we've produced is Grade-B garbage."
When the decision was made that Teddy and the Trogs in the pit would be real, were you asked to write those changes?
No, Lew did whatever Lew did. When he came on board he sort of said "I'm here now, I'm in charge, I make the decisions, whatever has been done before this is the past and I'm taking charge of the project." So what you see is Lew's interpretation.
If the trogs were all in Jamie's imagination, were they originally meant to escape as they do in the film?
Yes, he has to let them out because he's run out of nasty people. He's not accepting the fact he's created his little creatures in his mind, so how does he manage to look after them? As their creator he has to look after them, so the only thing he can do is put a rope down and let them climb out and look after themselves. And because they're doing bad things to good people, it's out of Jamie's hands, he has to ask for help. The help comes in the form of the adults in the community getting together and destroying this evil that he created. But all this take place entirely in his mind.
Was it your idea to have Jamie see the ghost of Sandy after her apparent death?
Yes, he cannot get her out of his mind and feels responsible for her death even though it was accidental and imaginary.
How did you feel about the way Jamie was portrayed?
The difficulty with making the picture was that you needed a boy to play the lead role, who had to be a good actor and yet be only 8 or 9 years of age. So we found five boys through pretty extensive casting, any one of which could play the role, so that they could all be presented to the director to select one of them. He never interviewed even one of those boys. He started all over again and picked a boy who was 12 years old, looked 14, and was almost muscular. That was his choice to play the role, and this to me was a fundamental mistake. Because this was a story from the mind of a psychotic child and to make him older, put him in a situation where he has almost a romantic relationship with the babysitter rather than that of a child to a young woman, changed the whole nature of the film.
Jamie was meant to be played by a younger child, and when a child is 8 or 9 years old they're interested in the mechanics, how the plumbing works and they first get interested in sexuality, but they're not sure why they have the feelings they have. Jamie has some kind of feelings for his babysitter Sandy, and he peeks on her, because children's sexuality at that level is usually at the peeping and showing stage. A 9 year old relating to a teenage babysitter like that, he's intimidated because he wants to experience something that he's not sure what it is because he's still a child.
The minute you make that child 12, as happened in the film, you change that utterly. Now a relationship between the two of them is possible, and we've seen examples of that in the media. The film was changed all together because it's no longer slightly confused childhood curiosity in which he's trying to express feelings he doesn't quite understand. In the scene where she's giving him a bath, that's a very different bath from the bath you give a child.
If you say to a 12 year old, "I want you to play a 9 year old," it's almost impossible for them to do that, they can't remember what it was like to be themselves a few years ago. They're maturing so rapidly, pretending to be less mature is difficult. A 12 year old is closer to a 17 year old than a 9 year old. At one point he tricks a woman who's mean to him into undressing, to cause her embarrassment, thinking it's very funny. But that's typical of an 8 or 9 year old. When Jamie was cast older, the basis for the entire film was changed and instead of being sort of cute, what he's doing is a little bit sinister and creepy.
Age difference aside, how did you feel about Sammy Snyders' performance?
I thought Sammy did a pretty good job from his perspective of an older child.
During the bath scene, Jamie seems to make a reference to molestation by his mother, asking Sandy if he knows why his mother washes him so much. Was that your intention?
I don't remember this line, and there was never the slightest suggestion that his mother did this, so it may have been something Lew had Sammy insert.
How did you feel about the way the trogs in the pit were depicted?
I'm not the first one who's noted that something you imagine is more horrible than something you're shown. Almost every horror movie's anticipation of seeing the monster is blown out the window when the monster finally appears. Of course, modern special effects can produce something quite scary and realistic, but to put suits on dwarves and have them running around pretending to be monster – if you see that for long enough you begin to laugh. You've done the worst thing a horror movie can do, which is step over the line between fantasy and the ludicrous.
So making someone believe for a long time in monsters that are running around, if they're shadowy things that you can hardly see – little yellow eyes in the dark – something scuttling around in the leaves, this you can believe in because you don't know what it is and your mind creates the image. The minute it's staring you in the face, it's a dwarf in a suit. And their eyes were lights! They actually put lights behind their eyes. Unbelievable.
How did you feel about the way Teddy was depicted?
With the teddy bear it was pretty close to what was originally written. It just depends how effective you are with the special effect, how realistic is the teddy bear. If you think it's just a kid with a stuffed toy and the stuffed toy moves on its own, that should startle you and give you a little shiver up and down the spine.
Was there supposed to be as much nudity in the film as there is?
The inclusion of nude scenes was almost mandatory at the time to get an "R" rating which was presumed to be required for the commercial horror audience. Only in one scene was it really essential, when Jamie was peeping on Sandy in the shower, she wouldn't be wearing clothes. Probably the most ludicrous fact about the shoot was that the director's wife refused to let him shoot the nude scenes, so I had to shoot them. I was a director with several films to my credit so it wasn't technically difficult, but the only scene involving nudity the director was allowed to film was the "skinny dipping" scene because the actress he hired for the part was his daughter!
Have you read the novelization by John Gault, under your original title "Teddy"?
Yes, and it was good for what it was. I actually met with Gault, but the producer had me writing a completely different script at the time, he didn't want me to be bothered with the novelization. I was busy working on the next script while Lew was shooting and Gault was writing the book.
What are your thoughts on The Pit today?
People still contact me after seeing the film – which amazes me to no end – to say that although it wasn't a good film they felt there was something else going on under the surface. I tell them that's the thing the director should have got, but didn't get because he really didn't care. He just saw it as a job, making a Grade-B horror film. It could've been a better film, but enough of the original script remained, I'm happy to say, that some people are still talking about this long after the fact.
Sincerest thanks to Mr. Ian A. Stuart for this interview!
80s Canada Horror Ian A. Stuart Sammy Snyders Screenwriting The Pit


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Carbon Neutral Alliance
Rüdersdorf, March 17, 2021
The global acting CEMEX, S.A.B. de C.V. has announced March 17th 2021, that, as part of its "Future in Action" program, it is developing a carbon neutral alliance in Germany to test and accelerate innovative technologies to reach carbon neutrality at its Rüdersdorf plant.
CEMEX is joining pioneering industrial consortiums with the objective of developing industrial-scale demonstration projects utilizing ground-breaking technologies to form this alliance, which aspires to achieve carbon neutrality at the Rüdersdorf plant by 2030. Several technologies, including waste heat recovery, carbon capture and transformation into building materials, synthetic fuels, and green hydrocarbons, are being evaluated, including hydrogen production with renewable energy.
The technological learnings derived from the Rüdersdorf "Carbon Neutral Alliance" should gradually be shared with the CEMEX global portfolio of cement plants and adapted to local conditions to accelerate its decarbonization goals of the cement production process. CEMEX is also applying to several European funds, such as the Innovation Fund & Green Deal Call, and national funding schemes for financial support and funding for these innovations, potentially providing best practices to other industrial players.
Aligned to the vision of the European Commission's Green Deal and climate aspirations, CEMEX communicated its vision for carbon neutrality last year.
In 2020, CEMEX's aspirations became known for the first time as "Vision Rüdersdorf". In order to place these in the global context and within the framework of the international goals, the "Carbon Neutral Alliance" has now been created.
Back to News Room
#CO2nversion
#ConcreteChemicals
© 2021 CEMEX Innovation Holding


Hi Friends! I'm in Indy this weekend for my high school reunion. Indiana just recently opened the first of their showrooms at their new Indiana Design Center and I just couldn't resist going to check it out. It is a beautiful building and will be a spectacular resource for Indiana designers when all the tenants fill their showroom space.
I met a really nice guy, Eric Reynolds, that runs his business Outre. They have wonderful upholstered pieces and some really pretty fabrics to put on those pieces. He used to have a retail furniture business in downtown Indy, but has moved his now 'To the Trades' showroom to the new design center. Go see him and his beautiful modern style furniture. I am sure you will find him to be as charming as I did. Here are some pictures from his space.
I went to The Trade Source. It's a beautiful showroom and the two Susans were dolls. They were so sweet and ordered a couple of memos for me. I can't wait to get them. I am so jealous that they have so many beautiful fabric lines all in one showroom. We have all those lines in Scottsdale, but I have to drive around to several design centers to access them all. Here are some of the photos I took of The Trade Source.
Nice stuff, right! Next, it was lunch time so we visited a new cafe that just opened in the Indiana Design Center on Rangeline Road in Carmel. It is called the Blu Moon Cafe and it was a great place to eat. The decor was nice too. Here are a few pictures.
Even their chairs were cool. However, when you are eating it is all about the food and the food didn't disappoint. I had their vegetable wrap. Two big thumbs up! My sweet mother had their chicken salad and said it was delish too. Brian and Michelle, the owners, met in culinary school and have a gem of a restaurant. Here's a picture of my food and a picture of my mother and I at the restaurant.
I'm the one on the right. I hope you've enjoyed my relaxing design day and aren't too jealous. Time to get cleaned up and head off to my reunion!
Oooh! I live in IL and totally see a road trip in my future.
I loved the chairs from the Trade source. So many fun options and styles. And I totally want the ottoman for my bedroom...I am jealous you got to go there!
So much wonderful furniture! I LOVE that headboard in the third picture from the top. So pretty!
Thanks so much for stopping by Outré while you were in town. It was my pleasure to chat with you and your mother. I hope you had safe travels back to Arizona. Stop by anytime.
That does look like a great day out. Gorgeous stuff.
Please stop by for a visit soon - I have an award for you : ) I hope you'll accept it!


Down a sunny dirt road, over a log bridge, up a grassy hill, deep in Bear Country, lived a family of bears— Papa Bear, Mama Bear and Small Bear.
They lived in a large tree which Papa Bear had hollowed out and made into a house.
It was a very fine house. This is what it looked like inside.
helping Mama bring the vegetables in from the garden.
There were all sorts of interesting things for a small bear to do and see in Bear Country.
Small Bear felt good growing up in a tree... in his own room... in the snug little bed that Papa Bear had made for him when he was a baby.
But one morning, it did not feel so good. Small Bear woke up with pains in his knees and aches in his legs.
"Small Bear, you have outgrown your little bed," said Papa Bear, as he hitched up his overalls and buttoned his shoulder straps.
"Today, we shall go out into the woods and make you a bigger one!"
took up his ax and was out the door. "But, Papa," called Small Bear, following after him. "What will happen to my little bed?"
"Don't worry about that, Small Bear," said Mama Bear as she closed the door after him.
She smiled and patted her front, which had lately grown very big and round. "You've outgrown that snug little bed just in time!"
Papa was sharpening his ax on his grinding stone and didn't hear.
not give you pains in your knees and aches in your legs."
then headed off into the woods.
"What will happen to my little bed?" Small Bear asked again as he caught up with Papa Bear in the woods. Papa had chopped down a tree and was splitting it into boards. "We will have a new baby soon who will need that little bed," said Papa Bear as he whacked off another board.
"A new baby? " asked Small Bear. (He hadn't noticed that Mama Bear had grown very round lately, although he had noticed it was harder and harder to sit on her lap.) "And it's coming soon?" "Yes, very soon!" said Papa Bear. With a final whack he split off the last board, which gave him enough wood to make a bigger bed for Small Bear.
They made the bed a good size and took the rest of the day to chip and shave it smooth and neat. Then they carried it back to the tree and up to Small Bear's room.
When they got there, Small Bear noticed right away that his old bed wasn't there anymore. "My little bed!" said Small Bear. "It's already gone!"
"You outgrew it just in time," called Mama Bear from the next room. "Come and see."
It was true! There was his snug little bed with a new little baby in it. Small Bear had outgrown his snug little bed just in time for his new baby sister. And now he was a big brother!
She was very little but very lively. As Small Bear leaned over for a closer look, she popped him on the nose with a tiny fist.
"Hmm," said Small Bear. "She has a pretty good punch for a little baby."
That night he stretched out proudly in his bigger bed. "Aah!" he said. "Being a big brother is going to be fun."
The next morning he woke up feeling fine, with no pains in his knees or aches in his legs. His nose was a little tender, though.
Report "The Berenstain Bears' New Baby"


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Cole Anthony sinks game-winner as Magic rally past Timberwolves to end skid
Venezolanos de Florida tildan orden de Trump como "un acto político, no humanitario"
TV Guy
Buddy Dyer explains vision for downtown Orlando
By Hal Boedeker
Staff writer |
UPDATED: In local public-affairs programming this weekend:
Orlando Mayor Buddy Dyer discusses his vision for downtown Orlando on "Orlando Matters" at 7 a.m. Saturday on WRBW-Channel 65. Dyer's topics include traffic, new developments and Lake Eola in an interview with host John Brown, a WOFL-Channel 35 anchor. (WOFL and the Orlando Sentinel have a content-sharing agreement.)
Rep. Rene Plasencia, R-Orlando, talks about the upcoming Special Session on "Political Connections" at 11 a.m. and 8 p.m. Sunday on News 13. Ybeth Bruzual conducts the interview. Lou Frey and Dick Batchelor provide analysis about the Special Session. The program uses PolitiFact's Truth-O-Meter to rate a claim by Sen. Marco Rubio, R-Fla., about the status of U.S. nuclear weapons.
Rasesh Thakkar, senior managing director of Tavistock Group, is a guest on "Flashpoint" at 8 a.m. Sunday on WKMG-Channel 6. Topics include Lake Nona's Medical City, which the Tavistock Group owns; Tavistock founder Joe Lewis' goals; and the development of the Medical City concept. Mike Holfeld hosts.
Greg Warmoth recaps WFTV-Channel 9's investigative reports for May on "Central Florida Spotlight" at 12:30 p.m. Sunday on the ABC affiliate. The topics include subsidized housing cheats; elaborate homeless camp sites in Orlando; sex offenders on Facebook and an App to track them; and a controversial law enforcement tool that some say may be an invasion of privacy. Other reports focused on a human trafficking victim's struggle to clear her name; a questionable land deal that cost the city of Orlando millions; and the alarming number of domestic violence-related deaths in Florida over the last decade.
"Metro Center Outlook" and "Global Perspectives" will be preempted Sunday morning for WUCF-Channel 24's spring membership drive.
Buddy Dyer


T&E platform Rydoo can automatically populate Uber for Business receipts into Rydoo's expense app. Employees then can submit those expenses with a single click. In August, Uber for Business announced its intention to integrate with Rydoo, along with Expensya, Happay, Zeno by Serko and Zoho Expense. The ride-hailing giant previously integrated with Concur, Certify, Chrome River and Expensify. According to Certify's latest SpendSmart study, Uber was the most-frequently expensed brand by business travelers in North America in the third quarter of 2018, accounting for nearly 11 percent of expenses Certify processed during the quarter.
Rydoo was launched in June 2018 by Sodexo, which built the platform from the technology acquired via Sodexo's 2017 acquisitions of Belgian expense management company Xpenditure and business travel booking tool Maya. Sodexo combined the tools for the desktop version of its platform, but Rydoo Expense and Rydoo Travel are separate apps. However, the company does plan to merge them into a single app eventually.