Kaposi's Sarcoma-Associated Herpesvirus Reactivation by Targeting of a dCas9-Based Transcription Activator to the ORF50 Promoter

Show full item record



Permalink

http://hdl.handle.net/10138/324034

Citation

Elbasani , E , Falasco , F , Gramolelli , S , Nurminen , V , Günther , T , Weltner , J , Balboa , D , Grundhoff , A , Otonkoski , T & Ojala , P M 2020 , ' Kaposi's Sarcoma-Associated Herpesvirus Reactivation by Targeting of a dCas9-Based Transcription Activator to the ORF50 Promoter ' , Viruses-Basel , vol. 12 , no. 9 , 952 . https://doi.org/10.3390/v12090952

Title: Kaposi's Sarcoma-Associated Herpesvirus Reactivation by Targeting of a dCas9-Based Transcription Activator to the ORF50 Promoter
Author: Elbasani, Endrit; Falasco, Francesca; Gramolelli, Silvia; Nurminen, Veijo; Günther, Thomas; Weltner, Jere; Balboa, Diego; Grundhoff, Adam; Otonkoski, Timo; Ojala, Päivi M.
Contributor: University of Helsinki, CAN-PRO - Translational Cancer Medicine Program
University of Helsinki, Faculty of Medicine
University of Helsinki, CAN-PRO - Translational Cancer Medicine Program
University of Helsinki, CAN-PRO - Translational Cancer Medicine Program
University of Helsinki, Centre of Excellence in Stem Cell Metabolism
University of Helsinki, Centre of Excellence in Stem Cell Metabolism
University of Helsinki, Helsinki One Health (HOH)
University of Helsinki, Department of Pathology
Date: 2020-09
Language: eng
Number of pages: 14
Belongs to series: Viruses-Basel
ISSN: 1999-4915
URI: http://hdl.handle.net/10138/324034
Abstract: CRISPR activation (CRISPRa) has revealed great potential as a tool to modulate the expression of targeted cellular genes. Here, we successfully applied the CRISPRa system to trigger the Kaposi's sarcoma-associated herpesvirus (KSHV) reactivation in latently infected cells by selectively activating ORF50 gene directly from the virus genome. We found that a nuclease-deficient Cas9 (dCas9) fused to a destabilization domain (DD) and 12 copies of the VP16 activation domain (VP192) triggered a more efficient KSHV lytic cycle and virus production when guided to two different sites on the ORF50 promoter, instead of only a single site. To our surprise, the virus reactivation induced by binding of the stable DD-dCas9-VP192 on the ORF50 promoter was even more efficient than reactivation induced by ectopic expression of ORF50. This suggests that recruitment of additional transcriptional activators to the ORF50 promoter, in addition to ORF50 itself, are needed for the efficient virus production. Further, we show that CRISPRa can be applied to selectively express the early lytic gene, ORF57, without disturbing the viral latency. Therefore, CRISPRa-based systems can be utilized to facilitate virus-host interaction studies by controlling the expression of not only cellular but also of specific KSHV genes.
Subject: KSHV
dCas9
CRISPRa
DD-dCas9-VP192
ORF50
RTA
KSHV lytic cycle
KSHV reactivation
Kaposi's sarcoma-associated herpesvirus
GENE-EXPRESSION
DNA-SEQUENCES
INFLAMMATORY SYNDROME
LYTIC REPLICATION
ENDOTHELIAL-CELLS
MEDIATED CONTROL
LATENCY
PROTEIN
CRISPR
VIRUS
3111 Biomedicine
11832 Microbiology and virology
Rights:


Files in this item

Total number of downloads: Loading...

Files Size Format View
viruses_12_00952_v2.pdf 5.365Mb PDF View/Open

This item appears in the following Collection(s)

Show full item record