Comparative functional analysis of Jembrana disease virus Tat protein on lentivirus long terminal repeat promoters: evidence for flexibility at its N-terminus.
BACKGROUND
Jembrana disease virus (JDV) encodes a potent regulatory protein Tat that strongly stimulates viral expression by transactivating the long terminal repeat (LTR) promoter. JDV Tat (jTat) promotes the transcription from its own LTR as well as non-cognate LTRs, by recruiting host transcription factors and facilitating transcriptional elongation. Here, we compared the sequence requirements of jTat for transactivation of JDV, bovine immunodeficiency virus (BIV) and human immunodeficiency virus (HIV) LTRs.


RESULTS
In this study, we identified the minimal protein sequence for LTR activation using jTat truncation mutants. We found that jTat N-terminal residues were indispensable for transactivating the HIV LTR. In contrast, transactivation of BIV and JDV LTRs depended largely on an arginine-rich motif and some flanking residues. Competitive inhibition assay and knockdown analysis showed that P-TEFb was required for jTat-mediated LTR transactivation, and a mammalian two-hybrid assay revealed the robust interaction of jTat with cyclin T1. In addition, HIV LTR transactivation was largely affected by fusion protein at the jTat N-terminus despite the fact that the cyclin T1-binding affinity was not altered. Furthermore, the jTat N-terminal sequence enabled HIV Tat to transactivate BIV and JDV LTRs, suggesting the flexibility at the jTat N-terminus.


CONCLUSIONS
This study showed the distinct sequence requirements of jTat for HIV, BIV and JDV LTR activation. Residues responsible for interaction with cyclin T1 and transactivation response element are the key determinants for transactivation of its cognate LTR. N-terminal residues in jTat may compensate for transactivation of the HIV LTR, based on the flexibility.
Su
Yang
Y
Key Laboratory of Molecular Microbiology and Biotechnology (Ministry of Education), College of Life Sciences, Nankai University, Tianjin 300071, China. tenlions@mail.nankai.edu.cn
Deng
Gang
G
Gai
Yuanming
Y
Li
Yue
Y
Gao
Yang
Y
Du
Jiansen
J
Geng
Yunqi
Y
Chen
Qimin
Q
Qiao
Wentao
W
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D056906:Cyclin T / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D015696:Gene Products, tat / Q000378:metabolism*; D006678:HIV / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D016087:Immunodeficiency Virus, Bovine / Q000502:physiology*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D025941:Protein Interaction Mapping; D017384:Sequence Deletion; D020079:Terminal Repeat Sequences; D020798:Two-Hybrid System Techniques
D056906:Cyclin T; D015696:Gene Products, tat