The complex human T-cell leukemia virus type 1 (HTLV-1) retrovirus encodes several proteins that are unique to the virus within its 3′-end region. Among them, the viral transactivator Tax and posttranscriptional regulator Rex are well characterized, and both positively regulate HTLV-1 viral expression. Less is known about the other regulatory proteins encoded in this region of the provirus, including the recently discovered HBZ protein. HBZ has been shown to negatively regulate basal and Tax-dependent HTLV-1 transcription through its ability to interact with specific basic-leucine zipper (bZIP) proteins. In the present study, we found that HBZ reduces HTLV-1 transcription and virion production. We then characterized the interaction between HBZ and the cellular transcription factor CREB. CREB plays a critical role in Tax-mediated HTLV-1 transcription by forming a complex with Tax that binds to viral cyclic AMP-response elements (CREs) located within the viral promoter. We found that HBZ and CREB interact in vivo and directly in vitro, and this interaction occurs through the bZIP domain of each protein. We also found that CREM-Ia and ATF-1, which share significant homology in their bZIP domains with the bZIP domain of CREB, interact with HBZ-bZIP. The interaction between CREB and HBZ prevents CREB binding to the viral CRE elements in vitro and in vivo, suggesting that the reduction in HTLV-1 transcription by HBZ is partly due to the loss of CREB at the promoter. We also found that HBZ displaces CREB from a cellular CRE, suggesting that HBZ may deregulate CREB-dependent cellular gene expression.

Activation of human T cell leukemia virus type 1 (HTLV-1) transcription is established through the formation of protein complexes on the viral promoter that are essentially composed of the cellular basic leucine zipper (bZIP) transcription factor cAMP-response element-binding protein (CREB (or certain other members of the ATF/CREB family), the HTLV-1-encoded transactivator Tax, and the pleiotropic cellular coactivators p300/CBP. HTLV-1 bZIP factor (HBZ) is a protein encoded by HTLV-1 that contains a bZIP domain and functions to repress HTLV-1 transcription. HBZ has been shown to repress viral transcription by dimerizing with CREB, which occurs specifically through the bZIP domain in each protein, and preventing CREB from binding to the DNA. However, we previously found that HBZ causes only partial removal of CREB from a chromosomally integrated viral promoter, and more importantly, an HBZ mutant lacking the COOH-terminal bZIP domain retains the ability to repress viral transcription. These results suggest that an additional mechanism contributes to HBZ-mediated repression of HTLV-1 transcription. In this study, we show that HBZ binds directly to the p300 and CBP coactivators. Two LXXLL-like motifs located within the NH₂-terminal region of HBZ are important for this interaction and specifically mediate binding to the KIX domain of p300/CBP. We provide evidence that this interaction interferes with the ability of Tax to bind p300/CBP and thereby inhibits the association of the coactivators with the viral promoter. Our findings demonstrate that HBZ utilizes a bipartite mechanism to repress viral transcription.

The homologous cellular coactivators p300 and CBP contain intrinsic lysine acetyl transferase (termed HAT) activity. This activity is responsible for acetylation of several sites on the histones as well as modification of transcription factors. In a previous study, we found that HBZ, encoded by the Human T-cell Leukemia Virus type 1 (HTLV-1), binds to multiple domains of p300/CBP, including the HAT domain. In this study, we found that HBZ inhibits the HAT activity of p300/CBP through the bZIP domain of the viral protein. This effect correlated with a reduction of H3K18 acetylation, a specific target of p300/CBP, in cells expressing HBZ. Interestingly, lower levels of H3K18 acetylation were detected in HTLV-1 infected cells compared to non-infected cells. The inhibitory effect of HBZ was not limited to histones, as HBZ also inhibited acetylation of the NF-κB subunit, p65, and the tumor suppressor, p53. Recent studies reported that mutations in the HAT domain of p300/CBP that cause a defect in acetylation are found in certain types of leukemia. These observations suggest that inhibition of the HAT activity by HBZ is important for the development of adult T-cell leukemia associated with HTLV-1 infection.

Human T-cell leukemia virus type-1-encoded protein HBZ represses p53 function by inhibiting the acetyltransferase activity of p300/CBP and HBO1