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Male Fitness Model / Muscle Model and Female Diva Fitness Model
All model contestants, male and female, will be required to perform a runway presentation in the competition rounds listed below. Models will also be required to perform quarter turns for the judges. There is short and tall height categories for females 5'2 and under is short class. Anything over 5'2 is tall class.
There are no weight limitations for Male or Female Model categories. Competitors must be at least 18 years of age to compete with the WBFF. We do provide music for the Model categories of the event.
NOTE: To clarify the Fitness Model categories, Male and Female Categories are being named differently, Male Fitness Model - Female Fitness Diva Model. All judging criteria's are the same, nothing has changed but the name of the category.
There is no music required for the male and female models to prepare to hand in for this competition category!
Female Diva Fitness Model Criteria:
Bikini round / Themewear round
Contestants for the Fitness Diva Model category are being judged on their sense of style, poise, overall beauty and stage presence as well as display a fit and toned physique. Fitness Diva Models will be expected to walk on stage in particular pattern which will be explained at the athletes meeting the evening prior to the competition (t walk style walk). Competitors must be able to present themselves in a classy feminine and tasteful way that is marketable. There will be 2 categories to this competition which include:
Round #1: Themewear round, prejudging show (example: Victoria
Round #2: Swimsuit bikini round (two piece any style)
Diva Fitness Model Classes
Short class: up to 1,62 m (5'2″)
40%- Overall Marketability
All rights reserved © 2011-2012 | The WBFF Denmark | Linde allé 6 | 6400 Sønderborg | + 45 42694600 | email@example.com | By skw


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Infiniti G35X Pricing Magically Fixed But Spoiler Disappears Too
Remember yesterday when we told you how the pricing on the Infiniti G35X was a bit funny? How the "Sport Package" appeared to be included in the price of the vehicle yet calls to Infiniti dealers said otherwise? Well today the option doesn't even seem to be listed on the website — not only that but they took away the… »

subset: commoncrawl


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Human T lymphotropic virus type-1 (HTLV-1) and type 2 (HTLV-2) are closely related human retroviruses, but have unique disease associations. HTLV-1 is the causative agent of an aggressive T-cell leukemia known as adult T-cell leukemia (ATL), HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP), and other inflammatory diseases. HTLV-2 infection has not been clearly associated with any disease condition. Although both viruses can transform T cells in vitro, the HTLV-1 provirus is mainly detected in CD4+ T cells whereas HTLV-2 is mainly detected in CD8+ T cells of infected individuals. HTLV-1 and HTLV-2 encode accessory proteins p30 and p28, respectively, which share partial amino acid homology and are required for viral persistence in vivo. The goal of this study was to identify host proteins interacting with p30 and p28 in order to understand their role in pathogenesis.
Affinity-tag purification coupled with mass spectrometric (MS) analyses revealed 42 and 22 potential interacting cellular partners of p30 and p28, respectively. Of these, only three cellular proteins, protein arginine methyltransferase 5 (PRMT5), hnRNP K and 60 S ribosomal protein L8 were detected in both p30 and p28 fractions. To validate the proteomic results, four interacting proteins were selected for further analyses using immunoblot assays. In full agreement with the MS analysis two cellular proteins REGγ and NEAF-interacting protein 30 (NIP30) selectively interacted with p30 and not with p28; heterogeneous nuclear ribonucleoprotein H1 (hnRNP H1) bound to p28 and not to p30; and PRMT5 interacted with both p30 and p28. Further studies demonstrated that reduced levels of PRMT5 resulted in decreased HTLV-2 viral gene expression whereas the viral gene expression of HTLV-1 was unchanged.
The comparisons of p30 and p28 host protein interaction proteome showed striking differences with some degree of overlap. PRMT5, one of the host proteins that interacted with both p30 and p28 differentially affected HTLV-1 and HTLV-2 viral gene expression suggesting that PRMT5 is involved at different stages of HTLV-1 and HTLV-2 biology. These findings suggest that distinct host protein interaction profiles of p30 and p28 could, in part, be responsible for differences in HTLV-1 and HTLV-2 pathobiology. This study provides new avenues of investigation into mechanisms of viral infection, tropism and persistence.
Human T lymphotropic virus type 1 (HTLV-1) and type 2 (HTLV-2) are complex deltaretroviruses that are closely related with approximately 70% nucleotide sequence similarity [1]. HTLV-1 was the first retrovirus linked to human malignancy [2,3]. HTLV-1 can infect T cells, B cells, fibroblasts and macrophages; however, the provirus is predominantly detected in CD4+ T cells [4-6]. HTLV-1 infection causes adult T-cell leukemia (ATL) in approximately 1–5% of infected individuals [7]. In addition, HTLV-1 infection has also been associated with a neurodegenerative disease, HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP) and other immune-mediated inflammatory diseases [8]. Despite being a closely related retrovirus, HTLV-2 unlike HTLV-1, has no clear disease association with the exception of a few cases of HAM/TSP-like neurodegenerative disease [9]. HTLV-2 shows selective tropism towards CD8+ T cells, both in vitro and in vivo[10-13]. Furthermore, although both viruses are capable of transforming T cells in vitro, it is intriguing that they exhibit sharply distinct target cell and transformation tropisms, and only HTLV-1 is associated with malignancy and chronic inflammatory conditions [14,15].
The genome organization of the HTLV-1 and HTLV-2 provirus is very similar with 5' and 3' LTRs. Both viruses encode regulatory and accessory proteins apart from the typical structural and enzymatic proteins Gag, Pol and Env. Two key regulatory gene products are Tax-1/Tax-2 and Rex-1/Rex-2 corresponding to HTLV-1/HTLV-2, respectively.
The Tax proteins encoded by both viruses are considered the primary oncoproteins [16,17] required for T cell transformation. However, Tax is not sufficient for the malignant process; additional proteins encoded by the virus are suggested to play a role as well [18]. For instance, HTLV-1 and HTLV-2 encode HZB and APH-2 proteins from the antisense strand of the proviral genome respectively [19,20], and growing evidence indicates a role for HBZ in the transformation process [21].
HTLV-1 p30 and HTLV-2 p28 are accessory proteins involved in the regulation of viral replication and persistence and could therefore affect the pathogenic outcome [22-27]. Both proteins are encoded by the corresponding viruses from a doubly spliced mRNA from the ORFII of the pX region and are dispensable for in vitro viral infection and T cell transformation. Interestingly, both p30 and p28 are required for the establishment of viral persistence in the rabbit model, underlining the importance of these proteins in vivo[25,27,28]. In addition, p30 has been shown to be required for infection of dendritic cells and non-human primates [26]. Both p30 and p28 are nuclear/nucleolar localizing proteins [29,30]. The nucleolar retention of p30 is linked to RNA transcription and binding to the 60 S ribosomal subunit [31]. Both p30 and p28 are post-transcriptional negative regulators of viral gene transcription that act by retaining tax/rex mRNA in the nucleus [22,30]. However, evidence indicates that p30 also regulates viral gene expression at a transcriptional level by competing with Tax for binding to CBP/p300 [23]. In addition, p30 and Rex may be a part of the nuclear retention mechanism by forming a ribonucleoprotein complex with tax/rex mRNA [32].
Microarray studies and genome-wide screens have shown that p30 differentially modulates cellular gene expression [33,34]. Expression of p30 activates the G2/M cell cycle checkpoint to promote cell survival, and delays entry into S-phase [35,36]. Under genotoxic stress, p30 promotes cell survival by binding and modulating levels of ATM possibly through binding to REGγ [37]. The ability of p30 to bind to the Myc-Tip60 complex and to also promote non-homologous end joining DNA repair support its role in cellular transformation [38,39].
Comparative studies of host protein interactions with HTLV-1 and HTLV-2 proteins have been largely focused on Tax-1 and Tax-2 [18,40-42]. Studies comparing host protein interactions of HTLV-1 and HTLV-2 accessory proteins have not been performed. Herein, we compared the cellular interacting protein profiles of p30 and p28 to better understand their roles in viral infection, persistence and cellular transformation. To this end, we used affinity tag purification of p30 and p28 coupled to mass spectrometry to identify potential interacting cellular proteins. We have confirmed the interaction of p30 with REGγ and identified a new p30 binding partner, NEAF-interacting protein 30 (NIP30). These cellular proteins copurified with p30 and not p28. In contrast, heterogeneous nuclear ribonucleoprotein H1 (hnRNP H1) interacted with p28 and not p30. Our data also reveal that arginine methyl transferase 5 (PRMT5) can interact with both p30 and p28. Knockdown studies of PRMT5 have indicated that this protein is important for effective gene expression of HTLV-2 and not HTLV-1. Our data provide new insights into the comparable host cell protein interactions used by these closely related human retroviruses.
Host protein interaction profiles of HTLV-1 p30 and HTLV-2 p28
In order to sample the p30 and p28 cellular proteome, we employed S-tag affinity purification [43] of ectopically expressed HTLV-1 p30 and HTLV-2 p28 in HEK 293T cells. An amino terminal S-tag and a HA and AU1 tag on the carboxy terminus were added to a CMV driven pTriEx4-Neo plasmid. S-tag affinity pull-down was performed on the lysates of cells, transfected with either empty vector (mock control) or p30 or p28, using S-beads. We analyzed p30- and p28-associated proteins using shotgun proteomics. The proteins that were unique to p30 and p28 purification fractions after subtracting the mock control proteins were considered as their potential interacting partners. The data were further refined by eliminating contaminants and highly abundant proteins, such as keratin, that were detected in controls. Duplicate experiments resulted in the identification of 42 and 22 potential interacting partners of p30 and p28, respectively (Tables 1 and 2). Of these, only three cellular proteins, PRMT5, hnRNP K and 60 S ribosomal protein L8 were detected in both p30 and p28 fractions. In order to validate the results of mass spectrometry-based proteomic experiments, we selected the following four cellular proteins for immunoblotting assays: two proteins REGγ and NEFA-interacting nuclear protein NIP30 (NIP30), which were exclusively found in p30 fractions (Table 1); heterogeneous nuclear ribonucleoprotein H1 (hnRNP H1) that purified with p28 and not with p30; and protein arginine methylate transferase 5 (PRMT5), which was found in both p30 and p28 fractions (Tables 1 and 2). An additional negative control of amino terminal S-tag GFP (S-GFP) expressed from the same expression vector was also tested in the immunoblotting assays.
REGγ and NIP30 interact with p30 and not with p28
We previously identified the interaction of p30 and REGγ using proteomic and molecular biology techniques [37]. However, the interaction of p28 and REGγ was not investigated previously. The proteomic data reported here indicate that REGγ selectively interacts with p30 and not with p28 (Tables 1 and 2). Similarly, we detected NIP30 in p30 (Table 1) and not in p28 (Table 2) fractions. To further evaluate these observations, 293T cells were transfected with mock, S-GFP, S-p30-HA and S-p28-HA. The cell lysates were subjected to S-tag affinity purification and immunoblotted with anti-REGγ and NIP30 antibodies. As shown in the Figure 1A, co-purification of REGγ and NIP30 with p30 and not p28 indicates the specific interaction of these proteins with p30. The expression and S-tag enrichment of p28 and p30 was tested by immunoblotting with anti-HA antibodies, whereas the expression and S-tag enrichment of GFP was evaluated with anti-GFP antibodies (Figure 1A).
Figure 1. Validation of p30 interaction with REGγ and NIP30.A) S-tag affinity purification with Mock, S-GFP, S-p30-HA and S-p28-HA transfected 293T cell lysates. The purified product was analyzed by immunoblotting using indicated antibodies. The expression of individual protein and S-tag purification was confirmed using the indicated antibodies. B) 293T cells were transfected with S-p30-HA and S-p28-HA and immunoprecipitated with non-specific IgG (NSIgG) or anti-REGγ antibody and probed with anti-HA antibody. Immunoblotting of input using anti-REGγ and anti-HA antibodies confirmed the expression of the interacting proteins. C) 293T cells lysates expressing S-p30-HA and S-p28-HA were immunoprecipitated with non-specific IgG (NSIgG) or anti-NIP30 antibody and probed with anti-HA antibody. The expression of p30, p28 and NIP30 was confirmed with indicated antibodies.
The interaction of p30 with REGγ was further confirmed by immunoprecipitating REGγ from lysates of cells transfected with S-p30-HA and S-p28-HA and immunoblotted with anti-HA antibody. REGγ was able to co-immunoprecipitate p30 but not p28 (Figure 1B). These results are fully consistent with our proteomic data (Tables 1 and 2). Similarly, we confirmed the interaction of NIP30 with p30 by immunoprecipitating NIP30 from S-p30-HA and S-p28-HA transfected cell lysates. The results shown in Figure 1C indicate that NIP30 can selectively co-immunoprecipitate p30 but not p28. Immunoblotting of the cell lysate with anti-REGγ, anti-NIP30 and anti-HA antibodies confirmed the expression of endogenous REGγ and NIP30 along with transfected p30 and p28 (Figure 1B, C). To evaluate the functional significance of these interactions we knocked down REGγ and NIP30 using siRNA, then transfected HTLV-1 molecular proviral clone plasmid, and measured p24 production. We observed that reduced levels of REGγ and NIP30 had no effect on p24 levels ( Additional file 1: Figure S1).
Additional file 1. Figure S1. Effects of REGγ and NIP30 knockdown on HTLV-1 gene expression. A) Two independent experiments (labeled 1 and 2) of control and REGγ siRNA treated 293T cells were transfected with Ach (HTLV-1 molecular clone). The knockdown of REGγ was confirmed by immunoblotting with anti-REGγ. The expression of HTLV p24 was monitored by immunoblotting. B) Negative siRNA and NIP30 siRNA treated 293 T cells were transfected with HTLV-1 molecular clone (Ach). The levels of NIP30 were tested by immunoblotting with anti-NIP30 antibodies. The expression of HTLV-1 p24 was monitored by anti-HTLV-1 p24 antibody. Equal loading of samples was confirmed by anti-actin antibodies. (JPEG 51 kb)
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p28 exclusively interacts with hnRNP H1
To confirm the interaction of p28 with hnRNP H1, lysates from cells with transient expression of mock, S-GFP, S-p30-HA and S-28-HA were S-tag affinity purified and immunoblotted with anti-hnRNP H1 antibody. The presence of hnRNP H1 only in the p28 purification and not in GFP or p30 purifications (Figure 2A) indicates that hnRNP H1 specifically binds to p28. Immunoblotting with anti-GFP and anti-HA to evaluate the expression and affinity purification of transfected GFP, p30 and p28 is shown in Figure 1A. In order to further confirm the interaction between p28 and hnRNP H1, lysates from cells transfected with S-p30-HA and S-p28-HA were immunoprecipitated with anti-hnRNP H1 antibodies and probed with anti-HA antibodies. Selective co-immunoprecipitation of p28 and not p30 by hnRNP H1 (Figure 2B) indicates that only p28 interacts with hnRNP H1.
Figure 2. Validation of p28 interaction with hnRNP H1.A) Ectopically expressed Mock, S-GFP, S-p30-HA, S-p28-HA in 293T cells were S-tag affinity purified and probed with anti-hnRNP H1 antibodies. Endogenous expression of hnRNP H1 was confirmed by immunoblotting with anti-hnRNP H1 antibody. B) The immunoprecipitation assay was performed on cell lysates transfected with S-p30-HA and S-p28-HA. Non-specific IgG (NSIgG) or anti-hnRNP H1 antibodies were used for immunoprecipitation and subsequently probed with anti-HA antibody. Immunoblotting using anti-HA and anti-hnRNP H1 was used to validate the expression of all proteins.
p30 and p28 interact with PRMT5
The mass spectrometry-based proteomic data indicate that both p30 and p28 interact with PRMT5. It should be noted that these experiments were conducted using S-tag affinity purification of lysates of 293T cells transfected with mock, S-GFP, S-p30-HA or S-p28-HA. While it has been reported [44] that PRMT5 can non-specifically associate with Flag-beads, our data show that with the S-tag affinity pull-down approach PRMT5 was detected in p30 and p28 fractions but not with mock or GFP purification. In order to confirm this interaction, S-tag affinity purification was performed on cell lysates from empty, S-GFP, S-p30-HA or S-p28-HA transfected 293T cells and analyzed by Western blotting using PRMT5 antibody. The ability of both p30 and p28 to co-purify PRMT5 (Figure 3) indicates that PRMT5 interacts with both p30 and p28. Immunoblots (Figure 1A) probed with anti-GFP and anti-HA antibodies confirmed the expression and S-tag purification of GFP, p30 and p28. To further validate the interaction of PRMT5 with p30 and p28, Flag tagged PRMT5 (Flag-PRMT5) was cotransfected with p30 or p28. Immunoprecipitation of Flag-PRMT5 using anti-Flag antibodies was able to co-immunoprecipitate p30 (Figure 3B). However, Flag-immunoprecipitation of Flag-PRMT5 was not able to co-immunoprecipitate p28 (Figure 3B). It is possible that the structural features of p28 allow the detection of PRMT5 binding only by S-tag affinity purification and not by immunoprecipitation. Looking at p30 proteomics data, it is also possible that interaction of p30 with PRMT5 binding proteins such as methylosome subunit pICln and methylosome 50 facilitate co-immunoprecipitation of p30 with PRMT5. The expression of Flag-PRMT5, p30 and p28 was confirmed using anti-Flag and anti-HA antibodies (Figure 3B).
Figure 3. Validation of p30 and p28 interaction with PRMT5. Mock, S-GFP, S-p30-HA and S-p28-HA expressed in 293T cells were purified by S-tag affinity purification and immunoblotting with anti-PRMT5 antibody. The expression of PRMT5 was confirmed by immunoblotting the cell lysates with anti-PRMT5 antibody. B) 293T cells co- transfected with p30 or p28 or Flag PRMT5 as indicated in the figure. Immunoprecipitation was performed using anti-Flag antibodies and subsequently probed for p30 and p28 using anti-HA antibodies. The expression of Flag-PRMT5, p30 and p28 was validated by immunoblotting as shown in the figure.
PRMT5 is required for HTLV-2 gene expression
To investigate a potential role of PRMT5 in HTLV biology, we downregulated the protein levels in 293T cells using the shRNA approach. Cells were then transfected with either HTLV-1 or HTLV-2 molecular clone plasmids, and viral gene expression was measured by immunoblotting for intracellular p24 and by ELISA for p19 production. The levels of PRMT5 had little effect on HTLV-1 gene expression in terms of intracellular p24 and p19 production as shown in Figure 4A. However, lower levels of PRMT5 resulted in a significant reduction in intracellular HTLV-2 p24 (Figure 4B). Consistent with this result HTLV-2 p19 measured by ELISA was also reduced 3 fold in PRMT5 knockdown cells (Figure 4B). These results indicate that PRMT5 plays a role in HTLV-2 gene expression, but has no measurable effect on HTLV-1 gene expression.
Figure 4. PRMT5 knockdown and HTLV gene expression.A) 293T cells expressing normal and knockdown levels of PRMT5 were transfected with HTLV-1 molecular clone. Immunoblotting was performed using anti-PRMT5 antibodies to confirm the knockdown and with anti-HTLV p24 antibodies to monitor p24 levels. The amount of p19 Gag production was analyzed by p19 ELISA. B) 293T cells transduced with scrambled shRNA or shRNA against PRMT5 were transfected with HTLV-2 molecular clone. Knockdown of PRMT5 was confirmed by immunoblotting with anti-PRMT5 antibodies. The intracellular p24 and extracellular p19 Gag production was analyzed by immunoblotting with anti-HTLV p24 antibodies and with p19 ELISA respectively. Student t-test resulted in a p value of 0.011 (*) indicating that decrease in p19 production with lower levels of PRMT5 is significant. Immunoblotting with anti-Actin antibodies was used to validate equal loading.
HTLV-1 and -2 are closely related human retroviruses with HTLV-1 being pathogenic whereas a clear correlation between HTLV-2 and pathogenesis has not been established. HTLV-1 and HTLV-2 have different pathological outcomes that cannot be attributed to a single viral protein, but are likely due to the contribution of viral proteins and their interactions with the host cellular machinery. Both viruses encode accessory proteins that are required for viral persistence in vivo. We hypothesize that differences in viral infection, tropism and pathological outcome may result in part from how these accessory proteins interact with host proteins. Recently host-pathogen interactome study comparing HTLV-1 and HTLV-2 was reported to propose novel hypothesis to explain differential pathobiology of these closely related viruses [45]. However, in this study yeast-two hybrid screening did not result in any p30 or p28 interacting proteins. In our study we conjugated over expression and mass spectrometry to compare the host protein interaction proteome of HTLV-1 p30 and HTLV-2 p28 accessory proteins. There is approximately 77% homology between the last and first 49 amino acids of p30 and p28, respectively, whereas the remaining amino acid sequences of p30 and p28 differ markedly. Therefore, it is logical to propose that these two viral proteins may exhibit some similarities and differences in their interactions with cellular proteins.
We adapted S-tag purification of p30 and p28 and combined it with mass spectrometry to identify potential host protein interactions. The approach yielded the list of proteins (Table 1 and 2), with the number of host proteins interacting with p30 being higher than p28. These findings are consistent with the observation that p30 is a multifunctional protein regulating transcription and post transcriptional gene expression, cell cycle/cell survival and DNA damage [35-39,46]. In contrast, p28 has only been implicated in post transcriptional gene regulation [30]. However, both proteins are important for high proviral loads and persistence in the rabbit model.
Detailed analysis of differences and similarities between p30 and p28 host protein-interacting profiles was performed in this study. For the analysis of host proteins that interact with p30, we combined the proteins identified in this study (Table 1) and previously identified proteins (Table 3). We then analyzed the molecular processes in which these interacting proteins are directly or indirectly involved. The graphical representation of molecular processes in which the host proteins interact with p30 (Figure 5A) and p28 (Figure 6A) suggests that p30 is involved in a wider variety of molecular processes than p28. The p30 pie graph (Figure 5A) indicates that 32% of p30 interacting proteins are involved in protein processes such as post translational modification (phosphorylation, ubiquitination and methylation), folding, and transport. Proteins involved in DNA damage repair and cell cycle constitute 14%, whereas proteins that are involved in energy metabolism contributed to 14% of the p30 interactome. Transcriptional proteins accounted for 4%, and only 2% of the proteins are involved in mRNA processing.
Table 3. Previously identified HTLV-1 p30 host interacting proteins
Figure 5. Analysis of p30 interacting host proteins.A) Functional distribution of HTLV-1 p30-interacting proteins summarized in the pie chart graph as percentages. B), C) and D) Ingenuity pathways analysis results in three network pathways with broad cellular functions with the involvement of p30-interacting proteins. The proteins that were identified in this study to interact with p30 are indicated in bold, and previously identified p30-interacting proteins are indicated in bold italics. The interaction between two proteins is indicated with a straight line; arrows indicate action upon in the direction of the arrow; and dashed lines indicate indirect interactions.
Figure 6. Analysis of p28 interacting host proteins.A) Functional distribution of HTLV-2 p28-interacting proteins summarized in the pie chart graph as percentages. B) and C) Ingenuity pathways analysis results in two network pathways with broad cellular functions with the involvement of p28-interacting proteins. The proteins that were identified in this study to interact with p28 are indicated in bold. The interaction between two proteins is indicated with a straight line; arrows indicate action upon in the direction of the arrow; and dashed lines indicate indirect interactions.
The distribution of p28 interacting processes from Table 3 resulted in a lower number of molecular processes. The major molecular processes (23%) are RNA processing, while 27% of the proteins are involved in regulation of protein phosphorylation, folding and transportation. Translation contributes to about 10 and 18% of the p30 and p28 interactome, respectively. Binding of both p30 and p28 to phosphatases suggests that both proteins might be phosphorylated or modulate the phosphorylation status of their binding partners. The ability of p30 to interact with ubiquitin and with proteins having ubiquitin conjugation activity suggests that p30 might be ubiquitinated. p30 is involved in DNA damage repair mechanisms, and regulation of phosphorylation and ubiquitination play a critical role in DNA damage repair. A large number of p30-interacting proteins are involved in energy metabolism like electron transport, glycolysis and fatty acid synthesis, and further investigation should be carried out to understand the functional significance of these interactions.
Furthermore, we evaluated indirect involvement of p30 and p28 interacting host proteins in various cellular processes using Ingenuity Pathways Analysis (Ingenuity Systems, http://www.ingenuity.com webcite). The analysis of all proteins that interact with p30 from Table 1 and with previously known p30-interacting proteins (Table 3) yielded three networks with high scores. The first network (Figure 5B) suggests significant involvement of p30-interacting proteins with DNA recombination, repair, cell cycle and cancer with pathways clustering around ATM, cyclin E, cyclin A and CDK2. It is consistent with various studies indicating the role of p30 in DNA damage repair, cell survival and cell cycle alteration. The second network (Figure 5C) revealed the influence of p30-interacting proteins on cancer, gastrointestinal disease and drug metabolism. The major convergence of the pathway was on ERK 1/2, Hsp 90 and 14–3–3 family proteins (YWHAZ). The last pathway (Figure 5D) is involved in neurological disease, developmental and hereditary disorders with central proteins being UBC, p38 MAPK and Akt, which are involved in various pathological conditions.
Similar analysis was performed using the list of p28-interacting proteins (Table 3) that resulted in two network pathways with high scores. The first pathway (Figure 6B) with histone proteins and hnRNP K at the center of the pathway suggested the involvement of p28-interacting proteins in RNA post transcriptional modification, amino acid metabolism and post translational modifications. The second pathway (Figure 6C) suggested the involvement of p28-interacting protein in neurological disease, skeletal muscle and development disorders. The central molecules of the second pathway were UBC and TNF, which are important for protein regulation and immune responses, respectively. The analysis suggests that p30 influences various pathways involved in processes such as cell cycle regulation, cell signaling and cancer biology. Interestingly, these pathways are also modulated by Tax and/or HBZ oncoproteins during cellular transformation. On the other hand, p28 has limited involvement in the kinds of cellular process that facilitate transformation.
We previously reported that p30 interacts with REGγ [37], which binds and activates the 20 S proteasome [48]. While REGγ targets a number of cellular proteins to proteasome degradation [49-52], we have shown that the levels of p30 correlate with the levels of REGγ indicating that p30 is not targeted to degradation via its interaction with REGγ [37]. We propose that the cell survival effect of p30 under naïve and genotoxic conditions is likely to be mediated through its interaction with REGγ to allow the viral infected cell to proliferate. Furthermore, our previous studies suggest that p30, ATM and REGγ are part of a multiprotein complex, and reduced levels of ATM under genotoxic stress might be due to REGγ binding to promote cell survival [37]. siRNA knockdown of REGγ had no effect on HTLV-1 viral gene expression ( Additional file 1: Figure S1A) suggesting that interaction of p30 with REGγ could play a role at other stages of the virus life cycle such as viral spread. In contrast with p30, p28 does not interact with REGγ. This difference between p30 and p28 could have possible long term effects during latency and clonal expansion of infected cells leading to the fully transformed or malignant state.
Another cellular protein NIP30, which selectively interacted with p30 and not with p28, is predicted to bind the DNA binding/EF hand/Leucine zipper protein (NEFA). NEFA was initially thought to bind DNA; however, it has been shown to be a Golgi localized calcium-binding protein [53,54]. The existence of NIP30 has been confirmed at the protein level and was initially identified by cDNA screens. It is localized to the nucleus, which is also the cellular compartment of p30 [55]. The biological role of NIP30 is not yet clear; however, based on its NEFA interaction it might play a role in calcium homeostasis. As calcium plays a crucial role in the immune response, the binding of p30 to NIP30 might influence the immune response to facilitate viral infection or replication [56]. Similarly to REGγ, NIP30 siRNA knockdowns had no effect on HTLV-1 viral gene expression ( Additional file 1: Figure S1B) suggesting that NIP30 might be involved in viral biology other than gene expression.
Although both p30 and p28 specifically bind tax/rex mRNA (potentially at exon 2/exon 3 spice junction) and retain it in the nucleus and thus regulate viral gene expression at the post transcriptional level, our findings in Figure 2 reveal that p28 and not p30 interacts with hnRNP H1, which is a part of spliceosome and involved in regulating splicing of mRNAs [57]. At the same time, the proteomics data show that both p30 and p28 bind to another host protein, hnRNP F, which is not only involved in mRNA processing but also has the ability to bind DNA [58,59]. Thus there is some degree of convergence and divergence in the mechanisms of action of p30 and p28. The processing of mRNA plays a critical role in viral infection and spread. The data suggest that p28 is more closely associated with mRNA processing compared to p30. However, both proteins interact and retain tax/rex mRNA in the nucleus as a part of the spliceosome to regulate viral gene expression. Further studies should provide insights into the mechanism of p30 and p28 function and avenues to develop novel therapeutic targets.
Both p30 and p28 interact with PRMT5, a type II PRMT enzyme that symmetrically dimethylates arginines [60]. The interaction of PRMT5 is of particular interest because it is up-regulated in B cell leukemia [61]. In the nucleus, PRMT5 is recruited to various transcription repressor complexes and also methylates histones H2A, H4R3 and H3R8 [62-65]. The methylation preference of PRMT5 is shifted to H4R3 upon binding to cooperator of PRMT5 (COPR5) [66]. A correlation between symmetrical methylation of H4R3 and H4R8 and transcription repression indicates the role of PRTM5 in transcription repression. PRMT5 also promotes cell survival via regulating p53 expression [67]. Upon DNA damage PRMT5 is known to methylate p53 through Strap co-factor to affect gene specificity of p53 [68]. In addition, PRMT5 functions in spliceosome formation and mRNA processing. PRMT5 is required for proper methylation of the mammalian cleavage factor 1 (mCF1), which is responsible for RNA 3'end processing [69]. PRMT5 is also required for Sm U snRNP methylation, which is important for in vivo for SMN complex formation to join proteins and RNA [70].
To investigate the possible role of PRMT5 interaction with p30 in transcriptional regulation, we performed PRMT5 knockdowns. HTLV-1 gene expression did not change significantly under low levels of PRMT5 (Figure 4A). The expression of p30 has been linked to increased cell survival and cell cycle deregulation. PRMT5 also has been linked to increased cell survival and to altering p53 function. Therefore, we posit that p30 and PRMT5 interaction might be relevant for cell survival to facilitate either viral spread or transformation. However, p28 interaction with PRMT5 might be relevant at transcriptional level by modulating the interaction of PRMT5 with transcription machinery. In addition p28 and PRMT5 interaction could also play a role at the post-transcriptional level in spliceosome formation, protein-mRNA recruitment or RNA processing. In both cases, PRMT5 would be affecting viral gene expression. This notion is consistent with our findings showing that downregulation of PRMT5 resulted in the reduced levels of HTLV-2 gene expression (Figure 4B). The interaction of p28 with PRMT5 might be regulating PRMT5 to modulate viral gene expression at either a transcriptional or post-transcriptional level. Furthermore, it is noteworthy that PRMT5 levels are up-regulated in B cell leukemia suggesting that PRMT5 might have a role to play in T cell transformation of HTLV-1 infected T cells. The study of the interaction of p30 and p28 with PRMT5 would provide a molecular model to understand the role of PRMT5 in cellular transformation.
In summary, we identified 42 and 22 potential cellular proteins interacting with HTLV-1 p30 and HTLV-2 p28, respectively. Of these, only three cellular proteins overlapped indicating markedly different interacting profiles for these two viral proteins. PRMT5 interacted with both p30 and p28, but had a differential effect on the gene expression of HTLV-1 and HTLV-2 indicating that PRMT5 might have different roles in HTLV-1 and HTLV-2 viral biology. The differences in protein and cellular processes with which p30 and p28 engage could be one reason for the difference in viral infection outcomes. Elucidating the role of PRMT5 in HTLV-1 viral biology would provide insights into how HTLV-1 and HTLV-2 are utilizing a host protein differently to potentially result in different pathological outcomes. Further investigation to understand the functional relevance of these interactions is warranted to better understand the HTLV life cycle and its role in T-cell transformation in order to develop new therapeutic drug targets.
Cell culture and plasmid transfection
HEK 293T cells were cultured in DMEM containing 10% FBS, 2 mM L-glutamine, 100 mg/ml streptomycin, and 100 units/ml penicillin. Plasmids were transfected using SuperFect (Qiagen, Valencia, CA) according to the manufacturer's instructions. Briefly 2x106 cells were plated in 100 mm dishes and after 24 hr, cells were transfected with 10 μg of pTriEx4-Neo (Mock), 10 μg of pTriEX-Neo-S-GFP (S-GFP), 10 μg of pTriEx4-Neo-S-p30-HA (S-p30-HA), 10 μg of pTriEx4-Neo-S-p28-AU1 or 4 μg of pTriEx4-Neo-S-p28-HA(S-p28-HA) from Novagen, Madison, WI. For Flag-PRMT5 immunoprecipitation 5 μg of pBabe (empty vector) or 5 μg of Flag-PRMT5 (generous gift from Dr. Robert Baiocchi's laboratory, Ohio State University) was co-tranfected with 5 μg of S-p30-HA or S-p28-HA. The cells were harvested and lysed in 1x passive lysis buffer after 24 hr (Promega, Madison, WI). 293T cells treated with siRNA or lentiviral particles expressing scrambled shRNA or PRMT5 shRNA were transfected with the HTLV-1 molecular clone, Ach (1.8 μg/well) and HTLV-2 molecular clone, Ph6neo (1.8 μg/well) of a six well plate using SuperFect (Qiagen, Valencia, CA). Oligofectamine (Invitrogen, Grand Island, NY) was used to transfect 293T cells with 100 nM of control siRNA or smart pool siRNA against REGγ and NIP30 (Dharmacon, Lafayette, CO). Stable knockdowns were produced by transducing 293T cells with lentiviral particles expresssing scramble or PRMT5 shRNAs (generous gift from Dr. Robert Baiocchi, Ohio State University) and selected for puromycin resistance.
HTLV-1 and HTLV-2 gene expression
Molecular proviral clones of HTLV-1 and HTLV-2 were transfected 24 hrs after siRNA treatment and incubated for 24 hrs. Supernatant was collected for measure of p19 Gag by ELISA (Zeptometrix, Buffalo, NY) and cells were lysated and immunoblotted for p24 production.
S-tag affinity purification and immunoprecipitation
Cell lysates were prepared with 1X passive lysis buffer (Promega, Madison, WI) in the presence of protease inhibitor mixture (Roche Applied Science, Indianapolis, IN). S-tag purification was performed by rocking cell lysates with S-beads (Novagen, Madison, WI) overnight at 4°C. The S-beads were washed once with high salt (1 M NaCl) containing radioimmunoprecipitation buffer (150 mM NaCl, 0.01 M Sodium pyrophosphate, 10 mM EDTA, 10 mM sodium fluoride, 50 mM Tris, 0.1% SDS, 12.8 mM deoxycholic acid, 10% glycerol and 1% Nonidet P-40 (pH 8.0)), three times with radioimmunoprecipitation buffer and once with PBS. The beads were suspended in ddH2O and subjected to shotgun proteomics. For subsequent immunoblotting the proteins were eluted by boiling the beads in SDS loading buffer. Immunoprecipitation was performed by rocking cell lysates overnight with specific antibody at 4°C and then with Sepharose protein A beads for 2 hrs (GE Sweden). Sepharose protein A beads were washed twice with high salt (1 M NaCl) containing radioimmunoprecipitation assay buffer and thrice with low salt (150 nM NaCl) radioimmunoprecipitation assay buffer and once with PBS. For western blotting proteins were eluted by boiling the beads in SDS loading buffer.
Shotgun proteomics
Capillary-liquid chromatography-tandem mass spectrometry (Capillary-LC/MS/MS) for global protein identification was performed on a Thermo Finnigan LTQ Orbitrap mass spectrometer equipped with a microspray source (Michrom Bioresources Inc, Auburn, CA) operated in positive ion mode. Samples were separated on a capillary column (0.2X150 mm Magic C18AQ 3 μ 200A, Michrom Bioresources Inc, Auburn, CA) using an UltiMateTM 3000 HPLC system from LC-Packings A Dionex Co. (Sunnyvale, CA). The scan sequence of the mass spectrometer was based on the data dependant TopTenTM method. The resolution of a full scan was set at 30000 to achieve a high mass accuracy MS determination.
The RAW data files collected on the mass spectrometer were converted to mzXML and MGF files by use of MassMatrix data conversion tools (version 1.3, http://www.massmatrix.net/download webcite). The resulting MGF files were searched using Mascot Daemon by Matrix Science version 2.2.2 (Boston, MA) and the database searched against the full SwissProt database version 57.5 (471472 sequences; 167326533 residues) or NCBI database version 20091013 (9873339 sequences; 3367482728 residues). Considered modifications (variable) were methionine oxidation and the presence of carbamidomethyl cysteine. Three missed cleavages for the enzyme were permitted with a peptide tolerance of 1.2 Da, and the MS/MS ion tolerance was 0.8 Da. Mock transfected (empty pTriEx4-Neo) cell lysates treated similarly served as negative control. Search results were compiled and visualized using the Scaffold 3 sofware. Unweighted spectrum count and percent coverage provided semi-quantitative data analyses. Protein identifications were assigned using PeptideProphet. Proteins with 80% confidence were accepted with a minimum of one peptide displaying 95% threshold confidence level.
Immunoblotting and antibodies
To confirm the MS/MS data of selected p28- and p30-interacting proteins we performed immunoblotting analysis. Cell lysate-derived proteins and proteins from S-Tag purification or immunoprecipitation assays were resolved by 4–20% gradient SDS-PAGE and transferred to nitrocellulose membranes prior to immunoblotting using the following primary and secondary antibodies: mouse anti-HA monoclonal antibodies (1:1000) (Covance Research Products, Princeton, NJ); rabbit anti-hnRNPH1 monoclonal (1:1000), mouse anti-hnRNPH1 (1:1000), mouse anti-HTLV p24 (1:1000) (Abcam, Cambridge, MA); Rabbit anti-NIP30 monoclonal (1:1000), rabbit anti-REGγ (1:1000) (Proteintech Group, Chicago, IL); mouse-anti β-actin (1:2000) (Sigma-Aldrich, St. Louis, MO); rat anti-Flag (1:1000) horse anti-mouse (1:2000), and goat anti-rabbit antibodies (1:2000) anti-rat (1:2000) (Cell Signaling, Danvers, MA).
NIP30, NEFA interacting protein 30; PRMT5, Protein Arginine methlytransferase 5; hnRNP H1, Heterogeneous nuclear ribonucleoprotein H1; HTLV-1 and -2, Human T-cell lymphotropic virus type 1 and 2; ATL, Adult T cell leukemia; HAM/TSP, HTLV-1 associated myelopathy/tropical spastic paraparesis.
Rami Doueiri and Rajaneesh Anupam contributed equally to this work.
Competing interests
Authors have no competing interests involved in data collection, materials and methods or conclusions.
Authors' contributions
RD and RA contributed equally to designing and collecting the data presented in the paper. KBG headed the shot-proteomics data collection. MK helped with mass spectrometry-based proteomic data analysis and finalizing the manuscript. MDL and PLG conceived, coordinated and helped in writing the manuscript. All authors read and approved the final manuscript.
We thank Kate Hayes-Ozello for editorial comments on the manuscript. This work was supported by a National Cancer Institute Program Grant CA100730 to MDL, PLG and MK.
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Despite these cases, the fact of the matter is, Noel Kearne approved Mary Beth Whitehead as a surrogate mother when she did not pass a psychological evaluation. He was told she would have a difficult time giving up the baby, and he did not care.
Surrogate mother Judy Stiver of Lansing, Michigan, sued him for impregnating her with the semen of a man who carried a herpes-related virus. She delivered a baby with a damaged brain because of it, which she was forced to keep.
Surrogate mother Patty Nowakowski, of Michigan, gave birth to twins. The intended parents decided that they only wanted the girl, and left Patty with their baby boy. Patty eventually got custody of both children, not something she had ever intended to do, but something she felt she needed to do based on the coldness of the people raising her biological daughter.
There were cases of surrogate mothers finding out after pregnant that their intended mother was actually a man, rumors of coercion regarding surrogacy applicants, and more.
Home==>What is Surrogacy==>Baby M
The History of Surrogacy Celebrities Using Surrogate Mothers Surrogacy in India About this Website About the Author


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Amtrak is adding more trains to its Wolverine route between Ann Arbor and Chicago in anticipation of a busy Thanksgiving weekend.
The passenger train service has deployed additional rail cars for the holiday, but spokesman Marc Magliari said this is the first time in his 13 years with Amtrak that it's offering four more trains along the Wolverine route for Thanksgiving.
Post Link: http://www.mlive.com/business/index.ssf/2012/10/amtrak_adding_trains_through_m.html


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Shen Yun is what could safely be described as a "really big show."
There are platoons of dancers, video projection, and a live orchestra, all dedicated to bringing to life 5,000 years of Chinese cultural traditions. The critic for the Pittsburgh Tribune-Review wrote of a recent performance, "The costumes were stunning, and most, particularly with the women dancers, were integral in the eye-popping movements. Sleeves contracted and unfurled, while dresses helped the dancers look like colorful pinwheels. The men displayed impressive acrobatic stunts."
There's also a political, pro-Falun Gong and anti-Chinese government point of view in a few of the set pieces.
So, now that you know what you're getting into, why not join more than 100 artists wearing 400 costumes and a full orchestra tonight and experience the sights and sounds of a long-lost age—and a continuing cultural tradition.
Shen Yun will be presented tonight (Thursday, May 9) at 7:30 PM on the Mainstage at Proctors (432 State St., Schenectady). Tickets are $50-$120. For more info, call the box office at 346-6204.


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If you are planning on an exemplary family vacation, North Georgia must be on your list. It is one of those destinations around the world which allows you to connect with nature. You can be assured to enjoy quality family time with activities available for everyone in the family. In terms of accommodation, the ideal option you have are North Ga cabins. They are renowned to comprise of all the amenities necessary to guarantee a satisfactory vacation. They are offered at competitive costs over the internet and can be easily afforded by you. Surf online to get best results for accommodation.
Located in the cold mountains, the North Ga cabins give you an opportunity to connect with nature. There are not too many locations as this one which suits people of all ages, be it aged or the young. You can be assured that everyone in the family will have some activity or the other to engage in. When you conduct research online, make sure that you find a reliable source. Visit different websites that offer Georgia cabin rentals and read through their reviews and testimonials. Eventually, you will come across a service provider that will give you good value for money.
The staff at the cabins is courteous and hospitable and will be responsible for catering to all your family needs. They possess necessary experience to make sure that there is no shortage of amenities being offered to their customers. Inside the cabins, you will find amenities such as Wi-Fi connection, flat screen TV, telephone, etc. You can also plan North Ga activities where you have plenty of options in shopping destinations as well as hiking and other adventure sports. Other activities which can be taken advantage of include swimming, trout fishing, kayaking, etc.
To add to the outdoor Georgia mountain activities, you can go in to the drive-in movies. You can take your car to the destination and along with your family catch up on a recent movie or a classic one. The theatre operates all around the year and you can enjoy the film from the comfort and coziness of your car. The North Ga cabins also have spacious kitchens and complimentary gadgets where the missus can cook delicious food for the entire family. With so much indoor as well as outdoor activities to engage in, you will surely find the cabins worth investing in.
Since there are a number of companies online, it is advisable that you do not rush in your search process. You must have learnt about instance wherein families have been duped by companies about delivery of exceptional packages. You do not want to experience this which is why it is recommended that you find the best company in business for North Ga cabins. When you find a trustworthy source, you will come across a range of packages offered. Take time to learn about such packages and accordingly invest in them. This memory will be cherished for a long time by everyone in the family.
North Georgia Cabin Rentals is the author of this article on Georgia Cabin Rentals.
Find more information, about Georgia mountain activities here


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Sunday: A storm system will kick up our winds and usher in cold air overnight. El Paso 66/33. Las Cruces 64/29. West 15 to 25 mph. Gusts will increase with front arriving to 40+ mph for west facing slope areas.
Monday: The cold front will keep our temperatures below average and give us cold nights. El Paso 50/28. Las Cruces 48/23. Northeast winds 5 to 15 mph.
Tuesday: Temperatures will warm some in the afternoon but still remain below average. El Paso 54/25. Las Cruces 53/21. Southeast winds 5 to 10 mph.


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|Colonial morphology of treponemes observed by electron microscopy.
MedLine Citation:
|PMID: 6727709 Owner: NLM Status: MEDLINE
|The colonial morphology of three strains of cultivable, nonpathogenic treponemes including a human oral treponeme was examined by light and electron microscopy. Treponema phagedenis strains Kazan and Reiter produced large white colonies on the surface of solid media composed of sterility test broth, 0.9 to 3.1% agar, rifampin, and 12.5% rabbit or horse serum. A human oral treponeme, strain G7201 , grew as diffused white zones on 0.9 to 3.1% agar plates. Under the cultural conditions employed agar concentrations slightly affected the time of appearance of colonies of the three strains of treponemes. When the colonies of these three strains were viewed by scanning electron microscopy, differences in their colonial morphology were observed. The 11-day-old colonies of human oral strain G7201 were very small, 5 to 15 micron in diameter, and had a slight irregular border. Kazan treponemes developed circular, entire and low convex colonies. Scanning and transmission electron microscopy revealed that the colonies of Reiter treponemes contained spherical forms almost up to 5 micron in diameter, each consisting of an outer membrane and a treponemal main body. They were very similar to the spherical bodies produced by strain G7201 in sucrose-containing broth.
|T Umemoto; I Namikawa; M Yamamoto
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Publication Detail:
|Type: Comparative Study; Journal Article
Journal Detail:
|Title: Microbiology and immunology Volume: 28 ISSN: 0385-5600 ISO Abbreviation: Microbiol. Immunol. Publication Date: 1984
Date Detail:
|Created Date: 1984-07-09 Completed Date: 1984-07-09 Revised Date: 2006-11-15
Medline Journal Info:
|Nlm Unique ID: 7703966 Medline TA: Microbiol Immunol Country: JAPAN
Other Details:
|Languages: eng Pagination: 11-22 Citation Subset: IM
Export Citation:
|APA/MLA Format Download EndNote Download BibTex
|MeSH Terms
Culture Media
Microscopy, Electron
Microscopy, Electron, Scanning
Treponema / cytology*, isolation & purification, ultrastructure
Reg. No./Substance:
|0/Culture Media; 9002-18-0/Agar
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
Previous Document: Drug resistance and R plasmids in Vibrio anguillarum isolated in cultured ayu (Plecoglossus altiveli...
Next Document: Cytotoxicity and calcium-dependent antigen of Yersinia.


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A larger version of Baltimore's beloved Inner Harbor ice rink opens today in the Amphitheatre.
The new rink, http://innerharboricerink.org, is 6,000 square feet, or 20 percent larger than the previous skating park at McKeldin Square, and will have stunning views of the Inner Harbor. It will be located at the top of the amphitheatre at 201 E. Pratt St.
Mayor Stephanie Rawlings-Blake will open the new rink Friday afternoon, according to a news release issued by the Waterfront Partnership of Baltimore.
A spokesman for the Partnership explained that the area atop the amphitheatre became available after Harborplace decided to...


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Address: 15740 Pines Blvd, Pembroke Pines, FL 33027 Map It!
About Tropical Financial Credit UnionEstablished in 1935, Tropical Financial Credit Union has grown to manage the time and money of more than 54,000 members in Miami and other areas of South Florida. Topical FCU's field of membership includes anyone that lives or works within Miami-Dade, Broward, or Palm Beach County and additional zipcodes in its operating territory or anyone who is an employee of one of Member-Companies (see website for details). Members enjoy the convenience of access to nine local branches as well as thousands of surcharge-free, network ATMs located across the nation.
Tropical Financial Credit Union Rates
Local Credit Union
Rates are current up to 30 days and show the best rate available, verify rate through institution site. The accuracy cannot be guaranteed.
Tropical Financial Credit Union Health Savings Account Rates at 3.00% APY
A tax-free, high-interest way to save for medical expenses.
Read More »
4 Reasons You Should Get a Tropical Financial Credit Union Mortgage in 2014
Get a home loan affordably and easily at Tropical Financial Credit Union this year.
Read More »
4 Best Credit Cards for Improving Your Credit in Fort Lauderdale
If you live in Fort Lauderdale, these credit cards could actually boost your poor credit score.
Read More »
Mortgage Interest Rates Today: Tropical Financial Credit Union at 3.375%
A mortgage is likely the largest loan that you will ever take out. Choosing a fixed interest rate will lock in your monthly payment and save...
Read More »
Renew Your Mortgage This Spring With Tropical Financial CU
Can't wait to get into your new home or refinance this spring?
Read More »
Tropical Financial Credit Union Reviews
Tropical Financial Credit Union Has Very Low Mortgage Interest Rates!
I couldn't be happier with my mortgage loan from Tropical Financial Credit Union. They were super helpful and got me an extremely low interest rate, lower than any bank could offer me!


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Book your room at Argosy On The Beach
Please enter your travel dates.
Get an overview of this hotelDelightful beach apartment with a spa tub
Nestled on the beach, this Clifton Beach apartment is 0 mi (0 km) from Clifton Beach and 2.7 mi (4.3 km) from Cairns Tropical Zoo. Palm Cove Beach is 2.1 mi (3.4 km) away.
Property Features
Guests of this beach apartment will appreciate convenient onsite amenities such as a spa tub and a sauna.
Room Amenities
All 16 individually furnished apartments feature WiFi and Select Comfort beds. Guests will find kitchens with refrigerators, stovetops, microwaves, and dishwashers. Weekly housekeeping is available.Information missing or incorrect? Tell us!
Hotel Amenities
Hotel Amenities
In addition to an outdoor pool, Argosy On The Beach provides a spa tub and a sauna. High-speed wireless Internet access is available for a surcharge. For a surcharge, the property provides an area shuttle within 20.00 km and a roundtrip airport shuttle (available 24 hours). This beach apartment also offers a library, tour/ticket assistance, and barbecue grills. Complimentary self parking is available on site.
Argosy On The Beach is a smoke-free property.
- Babysitting or childcare (surcharge)
- Airport transportation (surcharge)
- Airport transportation - drop-off (surcharge)
- Total number of rooms - 16
- Number of floors - 4
- Barbecue grill(s)
- Dry cleaning/laundry service
- Elevator/lift
- Front desk (limited hours)
- Tours/ticket assistance
- Library
- Laundry facilities
- Limo or Town Car service available
- Spa treatment room(s)
- Free self parking
- Year Built 1982
- Area shuttle (surcharge)
- Sauna
- Smoke-free property
- Garden
- Number of buildings/towers - 1
- Spa tub
- Outdoor pool
- WiFi (surcharge)
Family Friendly Amenities
- Family Hotel
- Outdoor pool
- Kitchen
- Laundry facilities
- Babysitting or childcare (surcharge)
- Full-sized refrigerator/freezer
- Microwave
- DVD player
Room Amenities
- Premium TV channels
- DVD player
- Premium bedding
- Furnished balcony or patio
- Access via exterior corridors
- Air conditioning
- Ceiling fan
- Climate control
- Kitchen
- Coffee/tea maker
- Microwave
- Weekly housekeeping provided
- Private bathroom
- Designer toiletries
- Hair dryer
- Iron/ironing board
- Desk
- Cookware, dishware, and utensils
- Blackout drapes/curtains
- Dishwasher
- Full-sized refrigerator/freezer
- Stovetop
- Cable TV service
- WiFi (surcharge)
- Individually furnished
- Sleep Number by Select Comfort mattress
- Cribs/infant beds (surcharge)
- Rollaway/extra beds (surcharge)
Nearby Things to Do
An outdoor pool and a spa tub are on site. Other recreational amenities include a sauna.
The recreational activities listed below are available either on site or nearby; fees may apply.
- Golfing nearby
- Snorkeling nearby
- Surfing/boogie boarding nearby
- Windsurfing nearby
- Kayaking nearby
- Sailing nearby
- Scuba diving nearby
- Water skiing nearby
- Rafting nearby
- Scooter/moped rentals nearby
- Bicycle rentals nearby
- Hiking/biking trails nearby
- Mountain biking nearby
- Skydiving nearby
- Parasailing nearby
- Ecotours nearby
- Golf lessons available nearby
- Boat tours nearby
To make requests for specific accessibility needs, complete the "Special or Accessibility Requests" section of "Trip Preferences" when you book your hotel.
- Accessible bathroom
- Accessible path of travel
- In-room accessibility
Hotel Policies
Check-in time ends at 5:30 PM
Check-in time starts at 2 PM
Minimum check-in age is 18
Special check-in instructions:
- Monday - Saturday: 8 AM - 5 PM
Check-out time is 10 AM
Payment types
You need to know
Extra-person charges may apply and vary depending on hotel policy.
Government-issued photo identification and a credit card are required at check-in for incidental charges.
Special requests are subject to availability upon check-in and may incur additional charges. Special requests cannot be guaranteed.
- Fee for in-room wireless Internet: AUD 10 (for 10 hours, rates may vary)
- Fee for wireless Internet in public areas: AUD 10 (for 10 hours, rates may vary)
- Airport shuttle fee: AUD 25 per person
- Crib (infant bed) fee: AUD 8.00 per night
- Rollaway bed fee: AUD 28.00 per night
We should mention
This property offers transfers from the airport (surcharges may apply). Guests must contact the property with arrival details before travel, using the contact information on the booking confirmation.


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You are here:
With the increase in call centres and other-telephone based businesses, more and more companies are using telephone interviews to check the telephone manner and skills of potential staff. Telephone interviewing is not usually a substitute for a face-to-face interview, however, they're a quick and easy way to identify and discount unsuitable applicants.
A phone interview is the best way to check on a candidate's telephone manner, especially where telephone manner and customer contact are key parts of the role.
Even though no-one can see you, smiling will help you to relax and sound positive. Some people also find that standing up while talking on the phone makes them feel more confident.
Make sure your surroundings are quiet and that you won't be disturbed. Shut the door, turn off the radio - and your mobile.
Make notes of what is covered - you may be asked the same questions if invited to a second interview.
Plan what might be asked in the interview beforehand, e.g. personal details, education, career history, experience. The interviewer will be aiming to match candidates against the job description and person specification so re-read thoroughly.
Just the way you answer the phone has an impact on the person calling. Talk distinctly and clearly. If you're not confident in your speaking voice, you cannot compensate for it in other ways.
Try to establish something in common. Ask about the caller's experience with the company or mention something you have read about the company.
Watch out for awkward silences, 'umms' and 'errrs', and unintentional interruptions - by either the candidate or interviewer. If you need a minute to compose your thoughts, don't be afraid to ask for a little time before answering.
You should be given the opportunity to ask questions - make sure you have some in mind.
Thank the caller for their time and express interest in the opportunity. (A good touch is to send an e-mail, reinforcing this.) Don't be afraid to ask what the next stage is.The advantages of a telephone interview are that it can be arranged with little delay, with little disruption to your existing job and you'll know the outcome quite quickly. However, what you say will be more important than in a face-to-face interview, so be prepared!
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Diversity & Equality | Disclaimer | Vacancy Policy | Data Protection


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Cambridge United sign John Sullivan
Posted: January 06, 2014|
Cambridge United have snapped up keeper John Sullivan on loan from Portsmouth until the end of the season.
Sullivan, 25, is set to make his debut in tomorrow's game at Braintree after he agreed terms this morning.
It follows the end of Chris Maxwell's loan deal from Fleetwood and the departure of Jason Brown after his short-term deal came to a close.
Will Norris has also returned from his loan spell with Royston to provide back-up to Sullivan.
Richard Money said he had tried to sign Sullivan last season only to bring in Nick Pope from Charlton on loan as well as making another attempt last summer with the keeper instead moving to Pompey.
He said: "We worked really hard to keep Chris here, but the key thing here is we made it perfectly clear for over a month to everybody else what we could pay.
"When it became pretty obvious it wasn't going to be the case, we were very resourceful and because we work extremely hard on our recruitment, we either revisit stuff we've looked at in the past or look another way.
"When we heard John might be available, it put us in a pretty relaxed mode.
"Chris has been outstanding in the first half of the season and I think we've been good for each other.
"He's disappointed, but he understands the parameteers we have at this club and we've all parted on good terms."


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Titanic Sails Into Cape Town's Waterfront
Rob PetersThe Titanic has always held me in its thrall. The ill-fated voyage of the most famous ship in history is a story that you could not make up – complete with villains, heroes and ultimately tragedy. So when the Titanic Expo sailed into Cape Town this month, I was first in line to snap up a ticket.
Cape Town a top eco-friendly destination
Rob PetersInternational travel magazine, Conde Nast Traveler, has named Cape Town in its top five list of Eco-Friendly cities in the world and said it is at the "forefront of the continent's green living movement".
Hit the Mother City Trails with Scootours
Rob PetersIf you're looking for something new, something unique and something exciting, look no further, because your chariot awaits and it's a scooter. But not just any scooter. These are scooters on steroids.
The Mount Nelson's Long and Storied History
Bianca ColemanThe Belmond Mount Nelson, affectionately known as 'the Nellie', or 'the Pink Lady' – the distinctive rosy hue was mixed specially for the hotel in the 1920s – has a fascinating history which dates back to when it was opened in March 1899.
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Cape Town a top eco-friendly destination
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Titanic Sails Into Cape Town's Waterfront
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#InstagramDiary: My Stellenbosch Wine Festival Experience
The Stellenbosch Wine Festival and Expo, presented by Pick 'n Pay and held from January 24 to February 2, 2014, is yet another reason we Capetonians have to celebrate Cape Town.
Wine canapés and French words in Cape Town's Constantia Winelands
And there's me thinking I'd seen all the beautiful wine farms this amazing city of Cape Town has to offer. Think again Diva. Beau Constantia est arrivé (by the way, when I see French words like 'beau' and ' pas de nom', I can't help but throw my French about a bit).
#Instagram Diary: Groote Post, Groote Delight
Monday 23 September, 2013, saw me heading out of the Mother City along the West Coast road and bouncing in a mini van through the veld towards the Darling Hills where, between the Kapokberg on one side and the Atlantic Ocean on the other, lies a gem of a wine farm called Groote Post.
My Big Bottle Festival Experience in Instagram
Saturday 24 August, 2013 found me wandering around the elegant Cellars-Hohenhort Hotel in Constantia at the third annual Big Bottle Festival, a rather a posh event, what with the Bentley and Lamborghini flanking the entrance way and the sound of high heels clicking across the conservatory floor (at least my sneakers were from Paul Smith!).
My Montagu Adventure on Instagram
Route 62 is one of the most stunning drives in the Western Cape, possibly in the whole of South Africa, and it's one of my favourites. It leads to some of the most picturesque towns and villages in the province, like Montagu.
Five amazing places for tea in Cape Town
Drinking tea is a gentle, languid pastime and there's no shortage of amazing places to indulge in an excellent blend in our gorgeous Mother City. I did a tea meander in the Cape Town CBD and surrounds, and these are a few amazing places to sip and savour the leaves – be they from Ceylon, China, prepared by the Earl, or plucked in the Cederberg, like our local pride and joy, rooibos.
My Good Food & Wine Show experience in Instagram
I confess this was my first visit to the Good Food & Wine Show in a long time. With more than 200 stands in the vast CTICC venue over four days from Thursday 23 to Sunday 26 May, 2013, I was amazed at the event's growth over the last few years. The stalls, some far more fabulous than others in both product and appearance, offered everything from food and wine to everything in between that relates to the home.
Top 5 delectable places to eat in tasty Cape Town: an Instagram essay
I love seeking out the best food and drink in tasty Cape Town, and then letting fellow foodies know when I find it. It can be anything from haute cuisine or a plate of salad at a deli to a craft beer, a glass of wine or a cup of coffee. If it's great, I'll Instagram it immediately it and show it off to my followers.


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Donate a book, receive a bouquet.
Royer's Flowers and Gifts' "Bouquets for Books" kicks off Friday and runs through Nov. 9. The annual book drive, which runs in York, Berks, Cumberland, Dauphin, Lancaster and Lebanon counties, encourages community members to donate a new book to their local library.
For each book, donors receive a free bouquet of flowers, up to three per family per visit.
Read more on the program and York County Libraries' wishlist at Book Buzz.


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1:28 pm CST - January 28, 2009
Posted under On The Record
Bikers Rally at Capitol to Gain Equal Recognition
Over 1500 bikers across Texas came together on Monday to lobby Texas legislators for name and club recognition.
Austin Bandido Gimmi Jimmy, State Chairman for United Clubs of Texas and U.S. Commander for the U.S. Defenders, told Texas Insider their ultimate goal was to "take congress away from congress, and give it back to the people." The goal for Monday however, was to make headway in gaining state recognition for Texas biker clubs.
"We're tired of being pushed off as second class citizens, and we're uniting as man power across the United States, as the U.S. Defenders, to take our rights back and get congress to recognize us," explained Jimmy.
"We want our votes to be heard, our names to be recognized and
our clubs to be recognized as people who do hold jobs. We are regular
citizens, we're fathers, grandmothers etc. and we're fighting for our
rights just like every citizen out there."
Bikers were eager to
tackle several issues in the legislative meeting including HIPAA
(Health Insurance Portability and Accountability Act), the Road Block
bill, and the Equal Access bill.
"HIPAA is a joke," said
Jimmy, "You fall down on your bike and your insurance company doesn't
have to pay for it because it was a recreational activity."
bill motorcyclists feel strongly against is the Road Block bill, also
being called Alcohol Checkpoint. According to the bikers on Monday,
this bill puts officers at liberty to charge anyone in a vehicle with
any crime they choose once they stop a driver at a Road Block or Check
Point. For the well-being and fairness of all drivers, bikers hope to
put this bill to a stop.
came to the capitol to stop not only HIPAA and the Road Block bill, but
to also end discrimination. The Texas Motorcycle Rights Association
wrote an Equal Access bill that would allow bikers equal protection
under the law in public places. Motorcyclists protested that they have
not been allowed equal parking spaces in public parking areas, as well
as having been turned away from public entities because of their dress.
The Equal Access bill, which bikers hope to pass, remedies this problem
by stating, "If a business wishes to have total control over who they
may and may not serve they can open as a Private Club rather than a
Public Accommodation."
As bikers went in to battle for their
rights, Jimmy summed their purpose with, "Give the power to the people
and let's put God back into our country and let the country be run by
the people again."


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Three more on Shasta County's "Most Wanted" list have been apprehended, the Redding Police Department reported today.
So far 89 suspects have been arrested since the county's weekly Most Wanted program geared toward spotlighting offenders that failed to appear in court launched in September. Another 19 remain at large.
The most recent arrests included:
- Jared Mylon French, 28, of Redding, who was picked up by Redding police in the 2200 block of Pine Street in Redding.
- Anthony Joseph Messina, 33, of Redding, who was arrested near the intersection of Placer and California streets by RPD.
- Theodore Salvatore Curato, 50, of Redding, who Shasta Interagency Narcotics Task Force agents arrested in the 9500 block of Yuki Yama Drive in Redding.
Anyone with information on most wanted subjects should call SHASCOM at 530-245-6540. You may also call Shasta County Secret Witness at 530-243-2319, or go to scsecretwitness.com/tip.htm.
Secret Witness is offering up to $250 for information that leads to the arrest of most wanted subjects.
Subscribe to the Redding Record Searchlight
Need Help? Call us at 1-877-710-6185.
Monday-Friday: 6am-6pm / Saturday & Sunday: 7am-10am


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I have changed thousands of diapers between my two children and all the babies I worked with as a NICU nurse. I have a neat and efficient diaper changing system. I have taught my husband, and he believes other people would benefit from learning my technique as well.
STEP 1: Choose your changing area
- Should be a flat stable surface such as a changing table, a couch, a bed, or even the floor
- Line the changing area with a lap pad, baby blanket, disposable changing pad, etc. for a clean changing surface
STEP 2: Prep all the supplies: diaper, wipes, creams, etc.
- Open up the clean diaper
- Remove 1-2 wipes, keep the rest nearby just in case
- Pre-load the diaper with A&D ointment, Desitin, etc. if necessary. Add 1/2-1 inch strip to the center of the open diaper
- If you are changing your baby's clothes as well, have all the new clothes within reach
STEP 3: Prep your baby
- Give your baby something to look at or play with (can be a toy, a picture, a mirror, or even your face!)
- Lay your baby on his back on the changing surface
- Remove all clothing covering the diaper area
- Unfasten the Velcro or stickers holding the diaper closed
- Fold the straps onto themselves so they don't get stuck to anything
STEP 4: Change the diaper Be quick here, you don't want to get a golden shower!
- Slowly pull the top of the diaper open towards your baby's feet
- Hold up your baby's butt slightly by lifting up his ankles with one hand
- Check for poop. If poop is present, gently remove the bulk of it by wiping your baby's butt with the part of the diaper that was covering his belly. By doing this, you will reduce the number of wipes used to clean him up.
- Tuck the used diaper in half under your baby. All of the poop and pee should be contained inside the dirty diaper at this point.
- Your baby should now be lying on his back with the used diaper underneath him. Why keep the dirty diaper under baby during cleaning? To catch any additional "deposits" your baby might make during cleaning!
- Clean your baby thoroughly with baby wipes. Drop the used wipe on top of the used diaper, and repeat until clean
- For girls, wipe from front to back, using a clean part of the wipe or a new wipe with each stroke
- For boys with foreskin, gently pull back the foreskin and clean gently
- For boys with a fresh circumcision site, apply petroleum jelly after cleaning
- For newborns with cord still attached, squeeze a few drops of alcohol onto the cord stump
- Remove the used diaper and wipes from under your baby's bottom, set aside (out of your baby's reach)
- Replace quickly with the new diaper, keeping the fasteners toward your baby's back
- If diaper cream was pre-applied, rub it onto your baby's skin with the diaper
- Pull the front of the diaper over your baby's belly and hold it snug
- Gently open the fasteners from the back and secure them over the front of the diaper
- Re-adjust the fasteners as needed
STEP 5: Replace your baby's clothes
- Don't forget to refasten all those snaps and buttons.
STEP 6: Dispose of dirty diaper
- Discard the dirty diaper and wipes in a diaper pail, garbage can, etc. Enclose in a plastic baggie to contain the smell, if desired.
STEP 7: Wash your hands
- If soap is not available, use an antibacterial gel such as Purell.
especially when on a changing table.
I hope this article helps you decrease the amount of time it takes for you to change a diaper. It is all about preparation and expecting the unexpected.
What is your diaper changing technique? Please feel free to share yours in the comments section below.


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Transition dressing is no mean feat, especially when dealing with that finicky period between winter and spring. Beyond color palette changes, the fabric, cut, layering, and accessories have to be taken into consideration so as to emerge from the closet chic and stylish, not confused, cold, and miserable.
You can't expect winter to transition smoothly into pleasant spring because there could be days of freezing temperature, despite what the calendar says. What can be done to address bipolar weather conditions is to keep a few closet essentials, and add or subtract accessories accordingly.
A selection of LBDs (little black dresses)
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