The Transcriptional Regulatory Functions of the SV40 Small t Ag

• Main Authors: LIU, Ning-Chun, 留寧群
• Other Authors: Wang Won-Bo
• Format: Others
• Language: zh-TW
• Published: 1998
• Online Access: http://ndltd.ncl.edu.tw/handle/44674642100715460554

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.