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Induction of p16INK4a is the major barrier to proliferation when Epstein-Barr virus (EBV) transforms primary B cells into Lymphoblastoid cell lines
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posted on 2023-06-15, 14:53 authored by Lenka Skalska, Robert E White, Gillian A Parker, Ernest Turro, Alison Sinclair, Kostas Pachos, Martin J AlldayTo explore the role of p16INK4a as an intrinsic barrier to B cell transformation by EBV, we transformed primary B cells from an individual homozygous for a deletion in the CDKN2A locus encoding p16INK4a and p14ARF. Using recombinant EBV-BAC viruses expressing conditional EBNA3C (3CHT), we developed a system that allows inactivation of EBNA3C in lymphoblastoid cell lines (LCLs) lacking active p16INK4a protein but expressing a functional 14ARF-fusion protein (p14/p16). The INK4a locus is epigenetically repressed by EBNA3C – in cooperation with EBNA3A – despite the absence of functional p16INK4a. Although inactivation of EBNA3C in LCLs from normal B cells leads to an increase in p16INK4a and growth arrest, EBNA3C inactivation in the p16INK4a-null LCLs has no impact on the rate of proliferation, establishing that the repression of INK4a is a major function of EBNA3C in EBV-driven LCL proliferation. This conditional LCL system allowed us to use microarray analysis to identify and confirm genes regulated specifically by EBNA3C, independently of proliferation changes modulated by the p16INK4a-Rb-E2F axis. Infections of normal primary B cells with recombinant EBV-BAC virus from which EBNA3C is deleted or with 3CHT EBV in the absence of activating ligand 4-hydroxytamoxifen, revealed that EBNA3C is necessary to overcome an EBV-driven increase in p16INK4a expression and concomitant block to proliferation 2–4 weeks post-infection. If cells are p16INK4a-null, functional EBNA3C is dispensable for the outgrowth of LCLs.
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Publication status
- Published
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- Published version
Journal
PLoS PathogensISSN
1553-7366Publisher
Public Library of ScienceExternal DOI
Issue
2Volume
9Article number
e1003187Department affiliated with
- Biochemistry Publications
Full text available
- Yes
Peer reviewed?
- Yes
Legacy Posted Date
2013-10-22First Open Access (FOA) Date
2013-10-22First Compliant Deposit (FCD) Date
2013-10-22Usage metrics
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