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Nuclear Factor kappa B is central to Marek's disease herpesvirus induced neoplastic transformation of CD30 expressing lymphocytes in-vivo.

Kumar S, Kunec D, Buza JJ, Chiang HI, Zhou H, Subramaniam S, Pendarvis K, Cheng HH, Burgess SC - BMC Syst Biol (2012)

Bottom Line: The exact mechanism of neoplastic transformation from CD30(lo) expressing phenotype to CD30(hi) expressing neoplastic phenotype is unknown.Here, using microarray, proteomics and Systems Biology modeling; we compare the global gene expression of CD30(lo) and CD30(hi) cells to identify key pathways of neoplastic transformation.We propose and test a specific mechanism of neoplastic transformation, and genetic resistance, involving the MDV oncogene Meq, host gene products of the Nuclear Factor Kappa B (NF-κB) family and CD30; we also identify a novel Meq protein interactome.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathobiology and Population Medicine, Mississippi State University, MS 39762, USA. skumar@cvm.msstate.edu

ABSTRACT

Background: Marek's Disease (MD) is a hyperproliferative, lymphomatous, neoplastic disease of chickens caused by the oncogenic Gallid herpesvirus type 2 (GaHV-2; MDV). Like several human lymphomas the neoplastic MD lymphoma cells overexpress the CD30 antigen (CD30(hi)) and are in minority, while the non-neoplastic cells (CD30(lo)) form the majority of population. MD is a unique natural in-vivo model of human CD30(hi) lymphomas with both natural CD30(hi) lymphomagenesis and spontaneous regression. The exact mechanism of neoplastic transformation from CD30(lo) expressing phenotype to CD30(hi) expressing neoplastic phenotype is unknown. Here, using microarray, proteomics and Systems Biology modeling; we compare the global gene expression of CD30(lo) and CD30(hi) cells to identify key pathways of neoplastic transformation. We propose and test a specific mechanism of neoplastic transformation, and genetic resistance, involving the MDV oncogene Meq, host gene products of the Nuclear Factor Kappa B (NF-κB) family and CD30; we also identify a novel Meq protein interactome.

Results: Our results show that a) CD30(lo) lymphocytes are pre-neoplastic precursors and not merely reactive lymphocytes; b) multiple transformation mechanisms exist and are potentially controlled by Meq; c) Meq can drive a feed-forward cycle that induces CD30 transcription, increases CD30 signaling which activates NF-κB, and, in turn, increases Meq transcription; d) Meq transcriptional repression or activation of the CD30 promoter generally correlates with polymorphisms in the CD30 promoter distinguishing MD-lymphoma resistant and susceptible chicken genotypes e) MDV oncoprotein Meq interacts with proteins involved in physiological processes central to lymphomagenesis.

Conclusions: In the context of the MD lymphoma microenvironment (and potentially in other CD30(hi) lymphomas as well), our results show that the neoplastic transformation is a continuum and the non-neoplastic cells are actually pre-neoplastic precursor cells and not merely immune bystanders. We also show that NF-κB is a central player in MDV induced neoplastic transformation of CD30-expressing lymphocytes in vivo. Our results provide insights into molecular mechanisms of neoplastic transformation in MD specifically and also herpesvirus induced lymphoma in general.

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QPCR based results of various transcription reporter assays and phylogenetic tree constructed from CD30 promoter sequences of MD resistant and susceptible genotypes. QPCR results from in vitro transcription assay comparing the efficacy of NF-κB subunits and Meq combinations on transcription from the MDV RB-1B Meq promoter (mean ± sem); a-f indicate statistical differences (A). Unrooted phylogenetic tree constructed from CD30 promoter sequences for MD-resistant (61, N) and MD-susceptible genotypes (7, 72, 15I5 and P) (B). Quantitative effects of Meq on transcription from the MD resistant and susceptible genotypes’ CD30 promoter (mean, 95% CI) (C).
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Figure 5: QPCR based results of various transcription reporter assays and phylogenetic tree constructed from CD30 promoter sequences of MD resistant and susceptible genotypes. QPCR results from in vitro transcription assay comparing the efficacy of NF-κB subunits and Meq combinations on transcription from the MDV RB-1B Meq promoter (mean ± sem); a-f indicate statistical differences (A). Unrooted phylogenetic tree constructed from CD30 promoter sequences for MD-resistant (61, N) and MD-susceptible genotypes (7, 72, 15I5 and P) (B). Quantitative effects of Meq on transcription from the MD resistant and susceptible genotypes’ CD30 promoter (mean, 95% CI) (C).

Mentions: Because we proposed a feed-forward loop model of increasing Meq and CD30 expression[18] and our global analysis suggests that NF-κB is central in MD lymphomagenesis, we tested NF-κB isoforms’ transactivation potential on the Meq promoter using in vitro transcription reporter assays (1 Kb 5’ of the Meq ATG; MDV strain RB-1B). We cloned genes RELA [p65], NFKB1 [p105/p50] and NFKB2 [p100/p52] and MEQ (RB-1B) into expression plasmids. SOgE cells[101] were transfected with the reporter plasmid alone or in combination with plasmids expressing different NF-κB isoforms and/or Meq, and transcription was quantified by QPCR. The three NF-κB isoforms differentially transactivated the Meq promoter (Figure5A): p52 was less than p50 and RELA (p65) alone, which produced similar transcription and were less than p50 and RELA (p65) together (in vivo p50 and p65 form the classical dimer). Meq alone transactivated the Meq promoter to similar levels as the positive control cytomegalovirus promoter and, when used together with different NF-κB isoforms, except in the p50-p65 dimer, it further increased transcription. This finding suggests that neoplastic transformation in MD depends fundamentally on CD30 signaling, and may explain why MD neoplastically-transformed cell survival (like that of many human herpesvirus induced lymphomas) critically depends on the lymphoma environment[6,12,15], as well as why MDV co-opted the CD30 signaling pathway.


Nuclear Factor kappa B is central to Marek's disease herpesvirus induced neoplastic transformation of CD30 expressing lymphocytes in-vivo.

Kumar S, Kunec D, Buza JJ, Chiang HI, Zhou H, Subramaniam S, Pendarvis K, Cheng HH, Burgess SC - BMC Syst Biol (2012)

QPCR based results of various transcription reporter assays and phylogenetic tree constructed from CD30 promoter sequences of MD resistant and susceptible genotypes. QPCR results from in vitro transcription assay comparing the efficacy of NF-κB subunits and Meq combinations on transcription from the MDV RB-1B Meq promoter (mean ± sem); a-f indicate statistical differences (A). Unrooted phylogenetic tree constructed from CD30 promoter sequences for MD-resistant (61, N) and MD-susceptible genotypes (7, 72, 15I5 and P) (B). Quantitative effects of Meq on transcription from the MD resistant and susceptible genotypes’ CD30 promoter (mean, 95% CI) (C).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3472249&req=5

Figure 5: QPCR based results of various transcription reporter assays and phylogenetic tree constructed from CD30 promoter sequences of MD resistant and susceptible genotypes. QPCR results from in vitro transcription assay comparing the efficacy of NF-κB subunits and Meq combinations on transcription from the MDV RB-1B Meq promoter (mean ± sem); a-f indicate statistical differences (A). Unrooted phylogenetic tree constructed from CD30 promoter sequences for MD-resistant (61, N) and MD-susceptible genotypes (7, 72, 15I5 and P) (B). Quantitative effects of Meq on transcription from the MD resistant and susceptible genotypes’ CD30 promoter (mean, 95% CI) (C).
Mentions: Because we proposed a feed-forward loop model of increasing Meq and CD30 expression[18] and our global analysis suggests that NF-κB is central in MD lymphomagenesis, we tested NF-κB isoforms’ transactivation potential on the Meq promoter using in vitro transcription reporter assays (1 Kb 5’ of the Meq ATG; MDV strain RB-1B). We cloned genes RELA [p65], NFKB1 [p105/p50] and NFKB2 [p100/p52] and MEQ (RB-1B) into expression plasmids. SOgE cells[101] were transfected with the reporter plasmid alone or in combination with plasmids expressing different NF-κB isoforms and/or Meq, and transcription was quantified by QPCR. The three NF-κB isoforms differentially transactivated the Meq promoter (Figure5A): p52 was less than p50 and RELA (p65) alone, which produced similar transcription and were less than p50 and RELA (p65) together (in vivo p50 and p65 form the classical dimer). Meq alone transactivated the Meq promoter to similar levels as the positive control cytomegalovirus promoter and, when used together with different NF-κB isoforms, except in the p50-p65 dimer, it further increased transcription. This finding suggests that neoplastic transformation in MD depends fundamentally on CD30 signaling, and may explain why MD neoplastically-transformed cell survival (like that of many human herpesvirus induced lymphomas) critically depends on the lymphoma environment[6,12,15], as well as why MDV co-opted the CD30 signaling pathway.

Bottom Line: The exact mechanism of neoplastic transformation from CD30(lo) expressing phenotype to CD30(hi) expressing neoplastic phenotype is unknown.Here, using microarray, proteomics and Systems Biology modeling; we compare the global gene expression of CD30(lo) and CD30(hi) cells to identify key pathways of neoplastic transformation.We propose and test a specific mechanism of neoplastic transformation, and genetic resistance, involving the MDV oncogene Meq, host gene products of the Nuclear Factor Kappa B (NF-κB) family and CD30; we also identify a novel Meq protein interactome.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathobiology and Population Medicine, Mississippi State University, MS 39762, USA. skumar@cvm.msstate.edu

ABSTRACT

Background: Marek's Disease (MD) is a hyperproliferative, lymphomatous, neoplastic disease of chickens caused by the oncogenic Gallid herpesvirus type 2 (GaHV-2; MDV). Like several human lymphomas the neoplastic MD lymphoma cells overexpress the CD30 antigen (CD30(hi)) and are in minority, while the non-neoplastic cells (CD30(lo)) form the majority of population. MD is a unique natural in-vivo model of human CD30(hi) lymphomas with both natural CD30(hi) lymphomagenesis and spontaneous regression. The exact mechanism of neoplastic transformation from CD30(lo) expressing phenotype to CD30(hi) expressing neoplastic phenotype is unknown. Here, using microarray, proteomics and Systems Biology modeling; we compare the global gene expression of CD30(lo) and CD30(hi) cells to identify key pathways of neoplastic transformation. We propose and test a specific mechanism of neoplastic transformation, and genetic resistance, involving the MDV oncogene Meq, host gene products of the Nuclear Factor Kappa B (NF-κB) family and CD30; we also identify a novel Meq protein interactome.

Results: Our results show that a) CD30(lo) lymphocytes are pre-neoplastic precursors and not merely reactive lymphocytes; b) multiple transformation mechanisms exist and are potentially controlled by Meq; c) Meq can drive a feed-forward cycle that induces CD30 transcription, increases CD30 signaling which activates NF-κB, and, in turn, increases Meq transcription; d) Meq transcriptional repression or activation of the CD30 promoter generally correlates with polymorphisms in the CD30 promoter distinguishing MD-lymphoma resistant and susceptible chicken genotypes e) MDV oncoprotein Meq interacts with proteins involved in physiological processes central to lymphomagenesis.

Conclusions: In the context of the MD lymphoma microenvironment (and potentially in other CD30(hi) lymphomas as well), our results show that the neoplastic transformation is a continuum and the non-neoplastic cells are actually pre-neoplastic precursor cells and not merely immune bystanders. We also show that NF-κB is a central player in MDV induced neoplastic transformation of CD30-expressing lymphocytes in vivo. Our results provide insights into molecular mechanisms of neoplastic transformation in MD specifically and also herpesvirus induced lymphoma in general.

Show MeSH
Related in: MedlinePlus