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The transcription profile of Tax-3 is more similar to Tax-1 than Tax-2: insights into HTLV-3 potential leukemogenic properties.

Chevalier SA, Durand S, Dasgupta A, Radonovich M, Cimarelli A, Brady JN, Mahieux R, Pise-Masison CA - PLoS ONE (2012)

Bottom Line: These results demonstrate that Tax-3 and Tax-1 are closely related in terms of cellular gene deregulation.The majority of those up-regulated genes are functionally linked in biological processes characteristic of HTLV-1-infected T-cells expressing Tax such as regulation of transcription and apoptosis, activation of the NF-κB cascade, T-cell mediated immunity and induction of cell proliferation and differentiation.In conclusion, our results demonstrate for the first time that, in T- and non T-cells types, Tax-3 is a functional analogue of Tax-1 in terms of transcriptional activation and suggest that HTLV-3 might share pathogenic features with HTLV-1 in vivo.

View Article: PubMed Central - PubMed

Affiliation: Virus Tumor Biology Section, Laboratory of Cellular Oncology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America. sebastien.chevalier@ens-lyon.fr

ABSTRACT
Human T-cell Lymphotropic Viruses type 1 (HTLV-1) is the etiological agent of Adult T-cell Leukemia/Lymphoma. Although associated with lymphocytosis, HTLV-2 infection is not associated with any malignant hematological disease. Similarly, no infection-related symptom has been detected in HTLV-3-infected individuals studied so far. Differences in individual Tax transcriptional activity might account for these distinct physiopathological outcomes. Tax-1 and Tax-3 possess a PDZ binding motif in their sequence. Interestingly, this motif, which is critical for Tax-1 transforming activity, is absent from Tax-2. We used the DNA microarray technology to analyze and compare the global gene expression profiles of different T- and non T-cell types expressing Tax-1, Tax-2 or Tax-3 viral transactivators. In a T-cell line, this analysis allowed us to identify 48 genes whose expression is commonly affected by all Tax proteins and are hence characteristic of the HTLV infection, independently of the virus type. Importantly, we also identified a subset of genes (n = 70) which are specifically up-regulated by Tax-1 and Tax-3, while Tax-1 and Tax-2 shared only 1 gene and Tax-2 and Tax-3 shared 8 genes. These results demonstrate that Tax-3 and Tax-1 are closely related in terms of cellular gene deregulation. Analysis of the molecular interactions existing between those Tax-1/Tax-3 deregulated genes then allowed us to highlight biological networks of genes characteristic of HTLV-1 and HTLV-3 infection. The majority of those up-regulated genes are functionally linked in biological processes characteristic of HTLV-1-infected T-cells expressing Tax such as regulation of transcription and apoptosis, activation of the NF-κB cascade, T-cell mediated immunity and induction of cell proliferation and differentiation. In conclusion, our results demonstrate for the first time that, in T- and non T-cells types, Tax-3 is a functional analogue of Tax-1 in terms of transcriptional activation and suggest that HTLV-3 might share pathogenic features with HTLV-1 in vivo.

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Intracellular localization of the Flag-Tax proteins.293 T cells were transduced by Lenti-IRES-GFP or Lenti-Flag-Tax lentiviruses for 72 h. Cells were fixed on Lab-tek slides and stained with the mouse monoclonal antibody anti-Flag-M2 (Sigma) followed by a goat anti-mouse CY3-conjugated secondary antibody (Amersham Biosciences). Nucleic acids were stained with DAPI-containing mounting medium (Vectashield, Vector Laboratories). Cells were vizualized using with a Zeiss Axioplan 2 imaging microscope, X40, and the SimplePCI software (Hamamatsu).
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pone-0041003-g002: Intracellular localization of the Flag-Tax proteins.293 T cells were transduced by Lenti-IRES-GFP or Lenti-Flag-Tax lentiviruses for 72 h. Cells were fixed on Lab-tek slides and stained with the mouse monoclonal antibody anti-Flag-M2 (Sigma) followed by a goat anti-mouse CY3-conjugated secondary antibody (Amersham Biosciences). Nucleic acids were stained with DAPI-containing mounting medium (Vectashield, Vector Laboratories). Cells were vizualized using with a Zeiss Axioplan 2 imaging microscope, X40, and the SimplePCI software (Hamamatsu).

Mentions: To verify that Flag-Tax protein localization was similar to that of previous publications, we performed imaging of 293 T cells transduced with the different Flag-Tax lentiviral particles. As expected, these proteins exhibited an intracellular pattern characteristic of the Tax proteins i.e. Tax-1 and Tax-3 were localized mainly in the nucleus but also in the cytoplasm, whereas Tax-2 displayed mainly a cytoplasmic distribution (Figure 2, see Flag panel) [51], [52], [53], [54], [55]. Together these data demonstrate that the Flag tag does not alter Tax intracellular localization. Next we tested the ability of the Flag-Tax proteins to activate transcription from the HTLV-1-LTR and from a NF-κB-dependent promoter in transduced cells. Tax proteins displayed a transcriptional activity between 89 to 275 fold over the control on the HTLV-1-LTR (Figure 3A). Of note, although Tax-1 had a lower expression (Figures 3C and D), it displayed the highest transcriptional activity on this promoter. The three proteins also showed an activation of over 4000 fold on the NF-κB-responding promoter (Figure 3B). Hence, these results demonstrate that these proteins are well expressed, localized as their untagged counterparts, and transcriptionally active.


The transcription profile of Tax-3 is more similar to Tax-1 than Tax-2: insights into HTLV-3 potential leukemogenic properties.

Chevalier SA, Durand S, Dasgupta A, Radonovich M, Cimarelli A, Brady JN, Mahieux R, Pise-Masison CA - PLoS ONE (2012)

Intracellular localization of the Flag-Tax proteins.293 T cells were transduced by Lenti-IRES-GFP or Lenti-Flag-Tax lentiviruses for 72 h. Cells were fixed on Lab-tek slides and stained with the mouse monoclonal antibody anti-Flag-M2 (Sigma) followed by a goat anti-mouse CY3-conjugated secondary antibody (Amersham Biosciences). Nucleic acids were stained with DAPI-containing mounting medium (Vectashield, Vector Laboratories). Cells were vizualized using with a Zeiss Axioplan 2 imaging microscope, X40, and the SimplePCI software (Hamamatsu).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3401231&req=5

pone-0041003-g002: Intracellular localization of the Flag-Tax proteins.293 T cells were transduced by Lenti-IRES-GFP or Lenti-Flag-Tax lentiviruses for 72 h. Cells were fixed on Lab-tek slides and stained with the mouse monoclonal antibody anti-Flag-M2 (Sigma) followed by a goat anti-mouse CY3-conjugated secondary antibody (Amersham Biosciences). Nucleic acids were stained with DAPI-containing mounting medium (Vectashield, Vector Laboratories). Cells were vizualized using with a Zeiss Axioplan 2 imaging microscope, X40, and the SimplePCI software (Hamamatsu).
Mentions: To verify that Flag-Tax protein localization was similar to that of previous publications, we performed imaging of 293 T cells transduced with the different Flag-Tax lentiviral particles. As expected, these proteins exhibited an intracellular pattern characteristic of the Tax proteins i.e. Tax-1 and Tax-3 were localized mainly in the nucleus but also in the cytoplasm, whereas Tax-2 displayed mainly a cytoplasmic distribution (Figure 2, see Flag panel) [51], [52], [53], [54], [55]. Together these data demonstrate that the Flag tag does not alter Tax intracellular localization. Next we tested the ability of the Flag-Tax proteins to activate transcription from the HTLV-1-LTR and from a NF-κB-dependent promoter in transduced cells. Tax proteins displayed a transcriptional activity between 89 to 275 fold over the control on the HTLV-1-LTR (Figure 3A). Of note, although Tax-1 had a lower expression (Figures 3C and D), it displayed the highest transcriptional activity on this promoter. The three proteins also showed an activation of over 4000 fold on the NF-κB-responding promoter (Figure 3B). Hence, these results demonstrate that these proteins are well expressed, localized as their untagged counterparts, and transcriptionally active.

Bottom Line: These results demonstrate that Tax-3 and Tax-1 are closely related in terms of cellular gene deregulation.The majority of those up-regulated genes are functionally linked in biological processes characteristic of HTLV-1-infected T-cells expressing Tax such as regulation of transcription and apoptosis, activation of the NF-κB cascade, T-cell mediated immunity and induction of cell proliferation and differentiation.In conclusion, our results demonstrate for the first time that, in T- and non T-cells types, Tax-3 is a functional analogue of Tax-1 in terms of transcriptional activation and suggest that HTLV-3 might share pathogenic features with HTLV-1 in vivo.

View Article: PubMed Central - PubMed

Affiliation: Virus Tumor Biology Section, Laboratory of Cellular Oncology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America. sebastien.chevalier@ens-lyon.fr

ABSTRACT
Human T-cell Lymphotropic Viruses type 1 (HTLV-1) is the etiological agent of Adult T-cell Leukemia/Lymphoma. Although associated with lymphocytosis, HTLV-2 infection is not associated with any malignant hematological disease. Similarly, no infection-related symptom has been detected in HTLV-3-infected individuals studied so far. Differences in individual Tax transcriptional activity might account for these distinct physiopathological outcomes. Tax-1 and Tax-3 possess a PDZ binding motif in their sequence. Interestingly, this motif, which is critical for Tax-1 transforming activity, is absent from Tax-2. We used the DNA microarray technology to analyze and compare the global gene expression profiles of different T- and non T-cell types expressing Tax-1, Tax-2 or Tax-3 viral transactivators. In a T-cell line, this analysis allowed us to identify 48 genes whose expression is commonly affected by all Tax proteins and are hence characteristic of the HTLV infection, independently of the virus type. Importantly, we also identified a subset of genes (n = 70) which are specifically up-regulated by Tax-1 and Tax-3, while Tax-1 and Tax-2 shared only 1 gene and Tax-2 and Tax-3 shared 8 genes. These results demonstrate that Tax-3 and Tax-1 are closely related in terms of cellular gene deregulation. Analysis of the molecular interactions existing between those Tax-1/Tax-3 deregulated genes then allowed us to highlight biological networks of genes characteristic of HTLV-1 and HTLV-3 infection. The majority of those up-regulated genes are functionally linked in biological processes characteristic of HTLV-1-infected T-cells expressing Tax such as regulation of transcription and apoptosis, activation of the NF-κB cascade, T-cell mediated immunity and induction of cell proliferation and differentiation. In conclusion, our results demonstrate for the first time that, in T- and non T-cells types, Tax-3 is a functional analogue of Tax-1 in terms of transcriptional activation and suggest that HTLV-3 might share pathogenic features with HTLV-1 in vivo.

Show MeSH
Related in: MedlinePlus