Transforming growth factor beta 1 stimulates expression of the Epstein-Barr virus BZLF1 immediate-early gene product ZEBRA by an indirect mechanism which requires the MAPK kinase pathway
H. Fahmi et al., Transforming growth factor beta 1 stimulates expression of the Epstein-Barr virus BZLF1 immediate-early gene product ZEBRA by an indirect mechanism which requires the MAPK kinase pathway, J VIROLOGY, 74(13), 2000, pp. 5810-5818
Disruption of Epstein-Barr virus (EBV) latency is mediated by ZEBRA the pro
tein product of the immediate-early EBV gene, BZLF1. In vitro, phorbol It-m
yristate 13-acetate (PIMA), a potent activator of protein kinase C (PKC), i
nduces reactivation of EBV. However, the physiological stimuli responsible
for the disruption of viral latency are not well characterized. Transformin
g growth factor beta 1 (TGF-beta 1) has also been shown to trigger the reac
tivation of EBV in Burkitt Lymphoma cell lines; however, the effect of TGF-
beta 1 on ZEBRA expression has not been reported. To further understand thi
s phenomenon, we have investigated the effect of TGF-beta 1 on ZEBRA expres
sion. Our results indicate that the treatment of different EBV-positive Bur
kitt's lymphoma cell lines with TGF-beta 1 induces a time-dependent activat
ion of BZLF1 transcription with a corresponding increase in the production
of the protein ZEBRA. TGF-beta 1 has been shown to exert its effects throug
h a wide range of intracellular routes; in the present study, we have explo
red these pathways. Transient expression of Smad proteins on their own had
no effect on ZEBRA expression, A specific inhibitor of p38 mitogen-activate
d protein kinase (MAPK), SB203580, did not affect TGF-beta 1-induced ZEBRA
expression, whereas treatment with the MAPK/ERK kinase inhibitors, PD98059
and U0126, dramatically decreased this induction, This suggests that TGF-be
ta 1 effect on BZLF1 expression requires the MAPK pathway. However, in Raji
and B95-8 cells additional routes can be used, as (i) the inhibition of ZE
BRA induction by PD98059 or U0126 was incomplete, whereas these inhibitors
completely abolished PMA-induced ZEBRA expression, (ii) TGF-beta 1 inductio
n of ZEBRA expression occurs in PKC-depleted cells, (iii) in Raji and in B9
5-8 cells, the effect of TGF-beta 1 and PMA are additive. Transient transfe
ction of the EBV-negative B-cell line DG75 with a BZLF1 promoter-fusion con
struct (Zp-CAT) showed that under conditions where the BZLF1 promoter is ac
tivated by PMA treatment, TGF-beta 1 had no significant effect on the expre
ssion of the chloramphenicol acetyltransferase gene. Furthermore, TGF-beta
1 induction of BZLF1 transcripts is dependent on de novo protein synthesis,
which suggests that TGF-beta 1 induces BZLF1 expression by an indirect mec
hanism.