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This study was funded by the Academy of Finland (Centre of Excellence Program 2014-2019: Translational Cancer Biology grant no. 307366), Finnish Cancer Foundation, and Sigrid Juselius Foundation. E.E. and S.G. were funded by the Academy of Finland postdoctoral research grants (no. 324487 and 309544). The funders had no role in the study design, data collection, interpretation or decision to submit the work for publication.

Analysis of institutional authors

Balboa, DAuthor

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November 24, 2020
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Kaposi's Sarcoma-Associated Herpesvirus Reactivation by Targeting of a dCas9-Based Transcription Activator to the ORF50 Promoter

Publicated to:Viruses-Basel. 12 (9): 952- - 2020-09-01 12(9), DOI: 10.3390/v12090952

Authors: Elbasani, E; Falasco, F; Gramolelli, S; Nurminen, V; Gunther, T; Weltner, J; Balboa, D; Grundhoff, A; Otonkoski, T; Ojala, PM

Affiliations

Barcelona Inst Sci & Technol, Ctr Genom Regulat, Barcelona 08003, Spain - Author
Imperial Coll London, Dept Infect Dis, London W2 1NY, England - Author
IRCCS, IOV, Veneto Inst Oncol, Surg Oncol Unit, I-35128 Padua, Italy - Author
Karolinska Inst, Dept Clin Sci Intervent & Technol, SE-14186 Stockholm, Sweden - Author
Karolinska Univ Sjukhuset, Div Obstet & Gynecol, SE-14186 Stockholm, Sweden - Author
Leibniz Inst Expt Virol, Heinrich Pette Inst, D-20251 Hamburg, Germany - Author
Univ Helsinki, Fac Med, Stem Cells & Metab Res Program, Helsinki 00290, Finland - Author
Univ Helsinki, Fac Med, Translat Canc Med Res Program, Helsinki 00290, Finland - Author
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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.

Keywords

Capsid proteinsCas9 endonuclease streptococcus pyogenesCrisprCrispr-associated protein 9CrispraDcas9Dd-dcas9-vp192Dna-sequencesEndothelial-cellsGene expression regulation, viralGene-expressionHerpesvirus 8, humanHumansImmediate-early proteinsInflammatory syndromeKaposi's sarcoma-associated herpesvirusKaposi’s sarcoma-associated herpesvirusKshvKshv lytic cycleKshv reactivationLatencyLytic replicationMediated controlOrf50Promoter regions, geneticProteinProtein domainsRecombinant fusion proteinsRtaRta protein, human herpesvirus 8Sarcoma, kaposiTrans-activatorsTranscriptional activationVirusVirus activationVirus latencyVp19 protein, human herpesvirus 1

Quality index

Bibliometric impact. Analysis of the contribution and dissemination channel

The work has been published in the journal Viruses-Basel due to its progression and the good impact it has achieved in recent years, according to the agency Scopus (SJR), it has become a reference in its field. In the year of publication of the work, 2020, it was in position , thus managing to position itself as a Q1 (Primer Cuartil), in the category Infectious Diseases.

Independientemente del impacto esperado determinado por el canal de difusión, es importante destacar el impacto real observado de la propia aportación.

Según las diferentes agencias de indexación, el número de citas acumuladas por esta publicación hasta la fecha 2025-07-16:

  • WoS: 4
  • Europe PMC: 4

Impact and social visibility

From the perspective of influence or social adoption, and based on metrics associated with mentions and interactions provided by agencies specializing in calculating the so-called "Alternative or Social Metrics," we can highlight as of 2025-07-16:

  • The use, from an academic perspective evidenced by the Altmetric agency indicator referring to aggregations made by the personal bibliographic manager Mendeley, gives us a total of: 14.
  • The use of this contribution in bookmarks, code forks, additions to favorite lists for recurrent reading, as well as general views, indicates that someone is using the publication as a basis for their current work. This may be a notable indicator of future more formal and academic citations. This claim is supported by the result of the "Capture" indicator, which yields a total of: 14 (PlumX).

With a more dissemination-oriented intent and targeting more general audiences, we can observe other more global scores such as:

  • The Total Score from Altmetric: 1.75.
  • The number of mentions on the social network X (formerly Twitter): 3 (Altmetric).

It is essential to present evidence supporting full alignment with institutional principles and guidelines on Open Science and the Conservation and Dissemination of Intellectual Heritage. A clear example of this is:

  • The work has been submitted to a journal whose editorial policy allows open Open Access publication.

Leadership analysis of institutional authors

This work has been carried out with international collaboration, specifically with researchers from: Finland; Germany; Italy; Sweden; United Kingdom.