Kinetics of HTLV-1 reactivation from latency quantified by single-molecule RNA FISH and stochastic modelling

• Published in: PLoS pathogens Vol. 15; no. 11; p. e1008164
• Main Authors: Miura, Michi, Dey, Supravat, Ramanayake, Saumya, Singh, Abhyudai, Rueda, David S., Bangham, Charles R. M.
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 18.11.2019; Public Library of Science (PLoS)
• Subjects: Activation; Analysis; Antisense RNA; Biology and life sciences; Cells, Cultured; Cellular stress response; Cloning; Computer engineering; Computer simulation; Gene Expression Regulation, Viral; Genomes; Health aspects; HTLV-I Infections - genetics; HTLV-I Infections - virology; Human T-lymphotropic virus 1 - physiology; Humans; In Situ Hybridization, Fluorescence; Incubation; Infection; Infectious diseases; Insertion; Kinetics; Latency; Leukemia; Leukocytes (mononuclear); Leukocytes, Mononuclear - virology; Lymphocytes; Lymphocytes T; Medicine; Medicine and Health Sciences; Parameter estimation; Peripheral blood mononuclear cells; Population; Proteins; Reaction kinetics; Research and Analysis Methods; Ribonucleic acid; RNA; Single-Cell Analysis - methods; Software; Stochastic models; Stochastic Processes; Stochasticity; T cells; Transcription; Transcription (Genetics); Viral Proteins - genetics; Virus Activation - genetics; Virus Latency - genetics; Virus Replication; Viruses; United Kingdom > UK; Delaware; United States > US
• ISSN: 1553-7374; 1553-7366; 1553-7374
• DOI: 10.1371/journal.ppat.1008164

The human T cell leukemia virus HTLV-1 establishes a persistent infection in vivo in which the viral sense-strand transcription is usually silent at a given time in each cell. However, cellular stress responses trigger the reactivation of HTLV-1, enabling the virus to transmit to a new host cell. Using single-molecule RNA FISH, we measured the kinetics of the HTLV-1 transcriptional reactivation in peripheral blood mononuclear cells (PBMCs) isolated from HTLV-1+ individuals. The abundance of the HTLV-1 sense and antisense transcripts was quantified hourly during incubation of the HTLV-1-infected PBMCs ex vivo. We found that, in each cell, the sense-strand transcription occurs in two distinct phases: the initial low-rate transcription is followed by a phase of rapid transcription. The onset of transcription peaked between 1 and 3 hours after the start of in vitro incubation. The variance in the transcription intensity was similar in polyclonal HTLV-1+ PBMCs (with tens of thousands of distinct provirus insertion sites), and in samples with a single dominant HTLV-1+ clone. A stochastic simulation model was developed to estimate the parameters of HTLV-1 proviral transcription kinetics. In PBMCs from a leukemic subject with one dominant T-cell clone, the model indicated that the average duration of HTLV-1 sense-strand activation by Tax (i.e. the rapid transcription) was less than one hour. HTLV-1 antisense transcription was stable during reactivation of the sense-strand. The antisense transcript HBZ was produced at an average rate of ~0.1 molecules per hour per HTLV-1+ cell; however, between 20% and 70% of HTLV-1-infected cells were HBZ-negative at a given time, the percentage depending on the individual subject. HTLV-1-infected cells are exposed to a range of stresses when they are drawn from the host, which initiate the viral reactivation. We conclude that whereas antisense-strand transcription is stable throughout the stress response, the HTLV-1 sense-strand reactivation is highly heterogeneous and occurs in short, self-terminating bursts.