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Transcriptional networks in plasmacytoid dendritic cells stimulated with synthetic TLR 7 agonists.

Birmachu W, Gleason RM, Bulbulian BJ, Riter CL, Vasilakos JP, Lipson KE, Nikolsky Y - BMC Immunol. (2007)

Bottom Line: A large number of chemokines and co-stimulatory molecules as well as the chemokine receptor CCR7 were increased in expression indicating maturation and change in the migratory ability of pDC.Further analysis of the data using the pathway analysis tool MetaCore gave insight into molecular and cellular processes impacted.The analysis also revealed increased expression of a network of genes important for protein ISGylation as well as an anti-apoptotic and pro-survival gene expression program.

View Article: PubMed Central - HTML - PubMed

Affiliation: Pharmacology, 3M Pharmaceuticals, St Paul, Minnesota, USA. wbirmachu@comcast.net

ABSTRACT

Background: Plasmacytoid Dendritic Cells (pDC) comprise approximately 0.2 to 0.8% of the blood mononuclear cells and are the primary type 1 interferon (IFN), producing cells, secreting high levels of IFN in response to viral infections. Plasmacytoid dendritic cells express predominantly TLRs 7 & 9, making them responsive to ssRNA and CpG DNA. The objective of this study was to evaluate the molecular and cellular processes altered upon stimulation of pDC with synthetic TLR 7 and TLR 7/8 agonists. To this end, we evaluated changes in global gene expression upon stimulation of 99.9% pure human pDC with the TLR7 selective agonists 3M-852A, and the TLR7/8 agonist 3M-011.

Results: Global gene expression was evaluated using the Affymetrix U133A GeneChip(R) and selected genes were confirmed using real time TaqMan(R) RTPCR. The gene expression profiles of the two agonists were similar indicating that changes in gene expression were solely due to stimulation through TLR7. Type 1 interferons were among the highest induced genes and included IFNB and multiple IFNalpha subtypes, IFNalpha2, alpha5, alpha6, alpha8, alpha1/13, alpha10, alpha14, alpha16, alpha17, alpha21. A large number of chemokines and co-stimulatory molecules as well as the chemokine receptor CCR7 were increased in expression indicating maturation and change in the migratory ability of pDC. Induction of an antiviral state was shown by the expression of several IFN-inducible genes with known anti-viral activity. Further analysis of the data using the pathway analysis tool MetaCore gave insight into molecular and cellular processes impacted. The analysis revealed transcription networks that show increased expression of signaling components in TLR7 and TLR3 pathways, and the cytosolic anti-viral pathway regulated by RIG1 and MDA5, suggestive of optimization of an antiviral state targeted towards RNA viruses. The analysis also revealed increased expression of a network of genes important for protein ISGylation as well as an anti-apoptotic and pro-survival gene expression program.

Conclusion: Thus this study demonstrates that as early as 4 hr post stimulation, synthetic TLR7 agonists induce a complex transcription network responsible for activating pDC for innate anti-viral immune responses with optimized responses towards RNA viruses, increased co-stimulatory capacity, and increased survival.

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Stimulation of pDC with 3M-852A and 3M-011 results in the induction of an anti-apoptotic gene expression program. Protein-interaction network was generated for expression changes in pDC 4 hr post stimulation with 3M-852A and 3M-011. The network was generated from a list of genes with GO process classification of anti-apoptosis, using the shortest path algorithm. The network highlights the large number of anti-apoptotic genes that are increased in expression with a concomitant decrease in expression of key pro-apoptotic genes including several that are transscriptionally regulated by p53. Remaining network details are as described in Figure 6.
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Figure 8: Stimulation of pDC with 3M-852A and 3M-011 results in the induction of an anti-apoptotic gene expression program. Protein-interaction network was generated for expression changes in pDC 4 hr post stimulation with 3M-852A and 3M-011. The network was generated from a list of genes with GO process classification of anti-apoptosis, using the shortest path algorithm. The network highlights the large number of anti-apoptotic genes that are increased in expression with a concomitant decrease in expression of key pro-apoptotic genes including several that are transscriptionally regulated by p53. Remaining network details are as described in Figure 6.

Mentions: The transcription factors CREB1, c-Myc and NFkB regulate the transcription of genes important in cell proliferation, differentiation, and apoptosis. Whereas most of the genes regulated by NFkB and CREB1 are increased in expression, a significant percentage of those regulated by c-Myc are decreased in expression. Several of the down regulated genes are involved in the regulation of cell cycle and of apoptosis and include Caspase 8 and BTG2 [45,46]. Genes increased in expression include pro-inflammatory genes, genes with growth factor activity and or proliferative activity including GRO1, GROG, GROB, MCL-1 [47,48] and anti-apoptotic activity, BCL2, c-IAP2, CFLIP, SERPINB9 [49-51]. Thus, the transcription regulatory network shown in Figure 7 suggests that 4 hr post stimulation with TLR7 agonists, pDC are programmed for cell survival. This is shown more clearly in Figure 8; a protein regulatory network built using a list of genes with GO classification of anti-apoptosis as the input nodes.


Transcriptional networks in plasmacytoid dendritic cells stimulated with synthetic TLR 7 agonists.

Birmachu W, Gleason RM, Bulbulian BJ, Riter CL, Vasilakos JP, Lipson KE, Nikolsky Y - BMC Immunol. (2007)

Stimulation of pDC with 3M-852A and 3M-011 results in the induction of an anti-apoptotic gene expression program. Protein-interaction network was generated for expression changes in pDC 4 hr post stimulation with 3M-852A and 3M-011. The network was generated from a list of genes with GO process classification of anti-apoptosis, using the shortest path algorithm. The network highlights the large number of anti-apoptotic genes that are increased in expression with a concomitant decrease in expression of key pro-apoptotic genes including several that are transscriptionally regulated by p53. Remaining network details are as described in Figure 6.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2175514&req=5

Figure 8: Stimulation of pDC with 3M-852A and 3M-011 results in the induction of an anti-apoptotic gene expression program. Protein-interaction network was generated for expression changes in pDC 4 hr post stimulation with 3M-852A and 3M-011. The network was generated from a list of genes with GO process classification of anti-apoptosis, using the shortest path algorithm. The network highlights the large number of anti-apoptotic genes that are increased in expression with a concomitant decrease in expression of key pro-apoptotic genes including several that are transscriptionally regulated by p53. Remaining network details are as described in Figure 6.
Mentions: The transcription factors CREB1, c-Myc and NFkB regulate the transcription of genes important in cell proliferation, differentiation, and apoptosis. Whereas most of the genes regulated by NFkB and CREB1 are increased in expression, a significant percentage of those regulated by c-Myc are decreased in expression. Several of the down regulated genes are involved in the regulation of cell cycle and of apoptosis and include Caspase 8 and BTG2 [45,46]. Genes increased in expression include pro-inflammatory genes, genes with growth factor activity and or proliferative activity including GRO1, GROG, GROB, MCL-1 [47,48] and anti-apoptotic activity, BCL2, c-IAP2, CFLIP, SERPINB9 [49-51]. Thus, the transcription regulatory network shown in Figure 7 suggests that 4 hr post stimulation with TLR7 agonists, pDC are programmed for cell survival. This is shown more clearly in Figure 8; a protein regulatory network built using a list of genes with GO classification of anti-apoptosis as the input nodes.

Bottom Line: A large number of chemokines and co-stimulatory molecules as well as the chemokine receptor CCR7 were increased in expression indicating maturation and change in the migratory ability of pDC.Further analysis of the data using the pathway analysis tool MetaCore gave insight into molecular and cellular processes impacted.The analysis also revealed increased expression of a network of genes important for protein ISGylation as well as an anti-apoptotic and pro-survival gene expression program.

View Article: PubMed Central - HTML - PubMed

Affiliation: Pharmacology, 3M Pharmaceuticals, St Paul, Minnesota, USA. wbirmachu@comcast.net

ABSTRACT

Background: Plasmacytoid Dendritic Cells (pDC) comprise approximately 0.2 to 0.8% of the blood mononuclear cells and are the primary type 1 interferon (IFN), producing cells, secreting high levels of IFN in response to viral infections. Plasmacytoid dendritic cells express predominantly TLRs 7 & 9, making them responsive to ssRNA and CpG DNA. The objective of this study was to evaluate the molecular and cellular processes altered upon stimulation of pDC with synthetic TLR 7 and TLR 7/8 agonists. To this end, we evaluated changes in global gene expression upon stimulation of 99.9% pure human pDC with the TLR7 selective agonists 3M-852A, and the TLR7/8 agonist 3M-011.

Results: Global gene expression was evaluated using the Affymetrix U133A GeneChip(R) and selected genes were confirmed using real time TaqMan(R) RTPCR. The gene expression profiles of the two agonists were similar indicating that changes in gene expression were solely due to stimulation through TLR7. Type 1 interferons were among the highest induced genes and included IFNB and multiple IFNalpha subtypes, IFNalpha2, alpha5, alpha6, alpha8, alpha1/13, alpha10, alpha14, alpha16, alpha17, alpha21. A large number of chemokines and co-stimulatory molecules as well as the chemokine receptor CCR7 were increased in expression indicating maturation and change in the migratory ability of pDC. Induction of an antiviral state was shown by the expression of several IFN-inducible genes with known anti-viral activity. Further analysis of the data using the pathway analysis tool MetaCore gave insight into molecular and cellular processes impacted. The analysis revealed transcription networks that show increased expression of signaling components in TLR7 and TLR3 pathways, and the cytosolic anti-viral pathway regulated by RIG1 and MDA5, suggestive of optimization of an antiviral state targeted towards RNA viruses. The analysis also revealed increased expression of a network of genes important for protein ISGylation as well as an anti-apoptotic and pro-survival gene expression program.

Conclusion: Thus this study demonstrates that as early as 4 hr post stimulation, synthetic TLR7 agonists induce a complex transcription network responsible for activating pDC for innate anti-viral immune responses with optimized responses towards RNA viruses, increased co-stimulatory capacity, and increased survival.

Show MeSH
Related in: MedlinePlus