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Release of human cytomegalovirus from latency by a KAP1/TRIM28 phosphorylation switch.

Rauwel B, Jang SM, Cassano M, Kapopoulou A, Barde I, Trono D - Elife (2015)

Bottom Line: Here, we reveal that HCMV latency in human CD34(+) HSC reflects the recruitment on the viral genome of KAP1, a master co-repressor, together with HP1 and the SETDB1 histone methyltransferase, which results in transcriptional silencing.Correspondingly, HCMV can be forced out of latency by KAP1 knockdown or pharmacological induction of KAP1 phosphorylation, and this process can be potentiated by activating NFkB with TNF-α.These results suggest new approaches both to curtail CMV infection and to purge the virus from organ transplants.

View Article: PubMed Central - PubMed

Affiliation: School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne, Lausanne, Switzerland.

ABSTRACT
Human cytomegalovirus (HCMV) is a highly prevalent pathogen that induces life-long infections notably through the establishment of latency in hematopoietic stem cells (HSC). Bouts of reactivation are normally controlled by the immune system, but can be fatal in immuno-compromised individuals such as organ transplant recipients. Here, we reveal that HCMV latency in human CD34(+) HSC reflects the recruitment on the viral genome of KAP1, a master co-repressor, together with HP1 and the SETDB1 histone methyltransferase, which results in transcriptional silencing. During lytic infection, KAP1 is still associated with the viral genome, but its heterochromatin-inducing activity is suppressed by mTOR-mediated phosphorylation. Correspondingly, HCMV can be forced out of latency by KAP1 knockdown or pharmacological induction of KAP1 phosphorylation, and this process can be potentiated by activating NFkB with TNF-α. These results suggest new approaches both to curtail CMV infection and to purge the virus from organ transplants.

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Pharmacological activation of HCMV does not trigger CD34+ cells differentiation.After 7 days of treatment with chloroquine and TNF-α, cells were analyzed by FACS for CD34 surface expression. Representative of experiments performed on cells from three different donors.DOI:http://dx.doi.org/10.7554/eLife.06068.028
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fig8s1: Pharmacological activation of HCMV does not trigger CD34+ cells differentiation.After 7 days of treatment with chloroquine and TNF-α, cells were analyzed by FACS for CD34 surface expression. Representative of experiments performed on cells from three different donors.DOI:http://dx.doi.org/10.7554/eLife.06068.028

Mentions: These data demonstrate that inducing KAP1 phosphorylation results in releasing HCMV from its latent state by relieving repressive marks on the viral chromatin. Nevertheless, levels of viral gene expression achieved in this setting remain lower than observed when HCMV-harboring precursors are differentiated into macrophages or mature dendritic cells, where NF-κB has been demonstrated to drive viral transcription. We thus exposed latently infected CD34+ cells to chloroquine and TNF- α (5 ng/ml), either alone or in combination (Figure 8). With TNF alone, HCMV transcription was not induced, indicating that the virus remained latent. With chloroquine alone, significant induction of cell-associated viral transcripts (approx. 100-fold) and DNA (approx. fivefold) was measured, and viral particles were released in the supernatant (10,000 PFU/ml in the experiment depicted in Figure 8). With the further addition of TNF, levels of CMV-specific RNAs increased by another 10-fold, the amounts of cell-associated viral DNA doubled, and virion production was boosted by a factor 10. Importantly, this dual treatment did not trigger the differentiation of the HSC (Figure 8—figure supplement 1). Taken together, these results suggest a model whereby chloroquine treatment renders the HCMV chromatin permissive for transcription through KAP1 S824 phosphorylation, which then allows TNF to induce full viral gene expression via NF-kB activation.10.7554/eLife.06068.027Figure 8.Combining KAP1 phosphorylation and NF-κB induction increases HCMV activation from latently infected HSC.


Release of human cytomegalovirus from latency by a KAP1/TRIM28 phosphorylation switch.

Rauwel B, Jang SM, Cassano M, Kapopoulou A, Barde I, Trono D - Elife (2015)

Pharmacological activation of HCMV does not trigger CD34+ cells differentiation.After 7 days of treatment with chloroquine and TNF-α, cells were analyzed by FACS for CD34 surface expression. Representative of experiments performed on cells from three different donors.DOI:http://dx.doi.org/10.7554/eLife.06068.028
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4384640&req=5

fig8s1: Pharmacological activation of HCMV does not trigger CD34+ cells differentiation.After 7 days of treatment with chloroquine and TNF-α, cells were analyzed by FACS for CD34 surface expression. Representative of experiments performed on cells from three different donors.DOI:http://dx.doi.org/10.7554/eLife.06068.028
Mentions: These data demonstrate that inducing KAP1 phosphorylation results in releasing HCMV from its latent state by relieving repressive marks on the viral chromatin. Nevertheless, levels of viral gene expression achieved in this setting remain lower than observed when HCMV-harboring precursors are differentiated into macrophages or mature dendritic cells, where NF-κB has been demonstrated to drive viral transcription. We thus exposed latently infected CD34+ cells to chloroquine and TNF- α (5 ng/ml), either alone or in combination (Figure 8). With TNF alone, HCMV transcription was not induced, indicating that the virus remained latent. With chloroquine alone, significant induction of cell-associated viral transcripts (approx. 100-fold) and DNA (approx. fivefold) was measured, and viral particles were released in the supernatant (10,000 PFU/ml in the experiment depicted in Figure 8). With the further addition of TNF, levels of CMV-specific RNAs increased by another 10-fold, the amounts of cell-associated viral DNA doubled, and virion production was boosted by a factor 10. Importantly, this dual treatment did not trigger the differentiation of the HSC (Figure 8—figure supplement 1). Taken together, these results suggest a model whereby chloroquine treatment renders the HCMV chromatin permissive for transcription through KAP1 S824 phosphorylation, which then allows TNF to induce full viral gene expression via NF-kB activation.10.7554/eLife.06068.027Figure 8.Combining KAP1 phosphorylation and NF-κB induction increases HCMV activation from latently infected HSC.

Bottom Line: Here, we reveal that HCMV latency in human CD34(+) HSC reflects the recruitment on the viral genome of KAP1, a master co-repressor, together with HP1 and the SETDB1 histone methyltransferase, which results in transcriptional silencing.Correspondingly, HCMV can be forced out of latency by KAP1 knockdown or pharmacological induction of KAP1 phosphorylation, and this process can be potentiated by activating NFkB with TNF-α.These results suggest new approaches both to curtail CMV infection and to purge the virus from organ transplants.

View Article: PubMed Central - PubMed

Affiliation: School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne, Lausanne, Switzerland.

ABSTRACT
Human cytomegalovirus (HCMV) is a highly prevalent pathogen that induces life-long infections notably through the establishment of latency in hematopoietic stem cells (HSC). Bouts of reactivation are normally controlled by the immune system, but can be fatal in immuno-compromised individuals such as organ transplant recipients. Here, we reveal that HCMV latency in human CD34(+) HSC reflects the recruitment on the viral genome of KAP1, a master co-repressor, together with HP1 and the SETDB1 histone methyltransferase, which results in transcriptional silencing. During lytic infection, KAP1 is still associated with the viral genome, but its heterochromatin-inducing activity is suppressed by mTOR-mediated phosphorylation. Correspondingly, HCMV can be forced out of latency by KAP1 knockdown or pharmacological induction of KAP1 phosphorylation, and this process can be potentiated by activating NFkB with TNF-α. These results suggest new approaches both to curtail CMV infection and to purge the virus from organ transplants.

Show MeSH
Related in: MedlinePlus