Characterizing the Interaction between the HTLV-1 Transactivator Tax-1 with Transcription Elongation Factor ELL2 and Its Impact on Viral Transactivation.

Stephan Kohrt,Andrea Thoma-Kress,Heinrich Sticht,Melanie Mann,Sarah Strobel,Bernhard Fleckenstein

doi:10.3390/ijms222413597

Abstract

The human T-cell leukemia virus type 1 (HTLV-1)-encoded transactivator and oncoprotein Tax-1 is essential for HTLV-1 replication. We recently found that Tax-1 interacts with transcription elongation factor for RNA polymerase II 2, ELL2, which enhances Tax-1-mediated transactivation of the HTLV-1 promotor. Here, we characterize the Tax-1:ELL2 interaction and its impact on viral transactivation by confocal imaging, co-immunoprecipitation, and luciferase assays. We found that Tax-1 and ELL2 not only co-precipitate, but also co-localize in dot-like structures in the nucleus. Tax-1:ELL2 complex formation occurred independently of Tax-1 point mutations, which are crucial for post translational modifications (PTMs) of Tax-1, suggesting that these PTMs are irrelevant for Tax-1:ELL2 interaction. In contrast, Tax-1 deletion mutants lacking either N-terminal (aa 1–37) or C-terminal regions (aa 150–353) of Tax-1 were impaired in interacting with ELL2. Contrary to Tax-1, the related, non-oncogenic Tax-2B from HTLV-2B did not interact with ELL2. Finally, we found that ELL2-R1 (aa 1–353), which carries an RNA polymerase II binding domain, and ELL2-R3 (aa 515–640) are sufficient to interact with Tax-1; however, only ELL2-truncations expressing R1 could enhance Tax-1-mediated transactivation of the HTLV-1 promoter. Together, this study identifies domains in Tax-1 and ELL2 being required for Tax-1:ELL2 complex formation and for viral transactivation.

Highlights

Human T-cell leukemia virus type 1 (HTLV-1) is a highly oncogenic delta-retrovirus predominantly infecting CD4+ T cells and causing an aggressive type of cancer called adult T-cell leukemia/lymphoma (ATL) or an inflammatory neurological disease leading to damage of the spinal cord, known as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) [1,2,3,4]
Upon transfection of 293T cells with EF1adriven Tax-1, ELL2, both Tax-1 and ELL2 expression plasmids, or empty vector control cells were stained at 24 h post transfection with anti-Tax, anti-ELL2, and the respective fluorescently labeled secondary antibodies
Upon transfection of 293T cells with EF1a-driven Tax-1, ELL2, both Tax-1 and ELL2 expression plasmids, or empty vector contro3lof 24 cells were stained at 24 h post transfection with anti-Tax, anti-ELL2, and the respective fluorescently labeled secondary antibodies

Summary

Introduction

Human T-cell leukemia virus type 1 (HTLV-1) is a highly oncogenic delta-retrovirus predominantly infecting CD4+ T cells and causing an aggressive type of cancer called adult T-cell leukemia/lymphoma (ATL) or an inflammatory neurological disease leading to damage of the spinal cord, known as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) [1,2,3,4]. Due to very inefficient cell-free transmission, the virus is mainly transmitted via cell–cell contacts at the virological synapse, via viral biofilms or via cellular protrusions [6,7,8]. HTLV-1 integrates in vivo into the host cell genome in its provirus form HTLV-1 infection is not part of sexual health screening in most countries; asymptomatic carriers are mainly unaware of their infection and may pass the virus to other people [12]. HTLV-1 shows reduced gene expression without expression of the immunodominant viral Tax protein, allowing the virus to escape from the CD8+ cytotoxic T-cell response (CTL) of the host, which is directed towards the immunogenic Tax-1 protein [13,14,15,16]

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Results

Conclusion