| Summary: | 碩士 === 國立臺灣大學 === 微生物學研究所 === 86 === Previously we found that simian virus 40 (SV40) small t antigen
(t) possesses both transcriptional activation and
transcriptional repression functions. Regarding the former
function, we have shown that t could somehow promote pRB
phosphorylation, release the E2F transcription factor from pRB
complex, and consequently activate the transcription of some E2
F-regulated promoters. In this thesis, We studied whether the
ability of t to promote pRB phosphorylation and to stimulate E2
F-regulated genes is correlated t
o its ability to inhibit protein phosphatase 2A (PP2A)
activity. We found that t, indeed, could stimulate the
transcription of endogenous E2F-regulated genes, including DNA
polymerase and dihydrofolate reductase (DHFR) genes, and
that this stimulatory function of t correlated with its ability
to inhibit PP2A. Furthermore, we also showed that the ability
of t to inhibit PP2A was required for its ability to promote
pRB phosphorylation.
Regarding transcriptional repression function of t, we tried to
study the mechanism which accounts for t''s ability to repress
many promoters/enhancers. We found that both t and T/t-common,
which encodes N-terminal 82 amino acids of t, could repress the
coactivation activity of p300, a transcriptional coactivator.
This inhibition was mediated by a specific interaction between
t and p300, because a p300 mutant, which sustains deletion
mutation at C/H3 domain involved in adenovirus E1A binding,
could no longer b
e inhibited by t. We also found that t and T/t-common could
repress the transcriptional activation function of p300
directly when the latter was brought to the promoter by fusing
with the DNA-binding domain of the papillomaviral E2 protein.
These results suggested that t and T/t-common could exert their
repression function by functionally interacting with
transcriptional coactivator p300 and thereby inhibiting the
latter''s activity.
Among various mechanisms which may account for the repression
of p300 activity by t, one is that t can physically interact
with p300 and block the latter''s activity. Indeed, when we used
a mammalian two-hybrid assay to study t-p300 interaction, we
found that t could physically interact with both N-terminal and
C-terminal portions of p300. This finding is reminiscent of
adenoviral E1A protein which also can repress many promoters/
enhancers by physically interacting with and inhibiting p300.
p300 is an important cell regulator involved in regulation of
gene transcription, cell proliferation and differentiation. t
has been shown by us to be able to repress a variety of
promoters/enhancers, and is known to be able to enhance
cellular transformation induced by SV40 large T. However the
mechanisms underlying these functions of t remains elusive. Our
finding that t could physically interact with p300 and block
the latter''s activity, could shed light on these questions.
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