Expression of HTLV-1 Genes in T-Cells Using RNA Electroporation
Human T-cell leukemia virus type 1 (HTLV-1) infects about 20 million people world-wide. Around 5% of the infected individuals develop adult T-cell leukemia (ATL) or a neurological disease termed tropical spastic paraparesis (TSP) after a clinical latency of years to decades. Through the use of two promoters and alternative splicing HTLV-1 expresses at least 12 different proteins. HTLV-1 establishes a life-long persistent infection by inducing the clonal expansion of infected cells, a property largely ascribed to the viral genes Tax and HBZ. However, the fact that ATL arises in a minority of infected individuals after a long clinical latency suggests the existence of factors counterbalancing the oncogenic potential of HTLV-1 in the context of natural infection.
To study the role of the different HTLV-1 gene products in the HTLV-1 life cycle, we optimized a transfection protocol for primary T-cells using an approach based on the electroporation of in vitro-transcribed RNA. Results showed that the RNA transfection technique combines a high transfection efficiency with low toxicity, not only in Jurkat T-cells but also in primary T-cells. These findings suggest that RNA electroporation is preferable for experiments aimed at investigating the role of HTLV-1 gene products in the context of primary T-cells, which represent the main target of HTLV-1 in vivo.
Key wordsHTLV-1 ATL In vitro transcription RNA electroporation Peripheral blood mononuclear cells (PBMCs)
The authors are grateful to Prof. Ugur Sahin (Research Center for Immunotherapy (FZI), Mainz, Germany; TRON—Translational Oncology at the University Medical Center of Johannes Gutenberg University, Mainz, Germany; Biopharmaceutical New Technologies (BioNTech) Corporation, Mainz, Germany) for providing the plasmid pST1-eGFPmut-2hBgUTR-A120. We are grateful to Armin Ensser and Benjamin Vogel (Institute of Clinical and Molecular Virology, Erlangen, Germany) for helpful discussions.
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