CG Dinucleotide Removal in Bioluminescent and Fluorescent Reporters Improves HIV-1 Replication and Reporter Gene Expression for Dual Imaging in Humanized Mice.
Chandra N RoyMariana A Benitez MorenoChris KlineZandrea AmbrosePublished in: Journal of virology (2021)
Visualizing the transmission and dissemination of human immunodeficiency virus type 1 (HIV-1) in real time in humanized mouse models is a robust tool to investigate viral replication during treatments and in tissue reservoirs. However, the stability and expression of HIV-1 reporter genes are obstacles for long-term serial imaging in vivo. Two replication-competent CCR5-tropic HIV-1 reporter constructs were created that encode either nanoluciferase (nLuc) or a near-infrared fluorescent protein (iRFP) upstream of nef. HIV-1 reporter virus replication and reporter gene expression was measured in cell culture and in humanized mice. While reporter gene expression in vivo correlated initially with plasma viremia, expression decreased after 4 to 5 weeks despite high plasma viremia. The reporter genes were codon optimized to remove cytosine/guanine (CG) dinucleotides, and new CO-nLuc and CO-iRFP viruses were reconstructed. Removal of CG dinucleotides in HIV-1 reporter viruses improved replication in vitro and reporter expression in vivo and ex vivo. Both codon-optimized reporter viruses could be visualized during coinfection and in vivo reporter gene expression during treatment failure preceded detection of plasma viremia. While the dynamic range of CO-iRFP HIV-1 was lower than that of CO-nLuc HIV-1, both viruses could have utility in studying and visualizing HIV-1 infection in humanized mice. IMPORTANCE Animal models are important for studying HIV-1 pathogenesis and treatments. We developed two viruses each encoding a reporter gene that can be expressed in cells after infection. This study shows that HIV-1 infection can be visualized by noninvasive, whole-body imaging in mice with human immune cells over time by reporter expression. We improved reporter expression to reflect HIV-1 replication and showed that two viral variants can be tracked over time in the same animal and can predict failure of antiretroviral therapy to suppress virus.
Keyphrases
- antiretroviral therapy
- human immunodeficiency virus
- hiv infected
- hiv positive
- crispr cas
- hiv aids
- gene expression
- hiv infected patients
- hepatitis c virus
- hiv testing
- men who have sex with men
- poor prognosis
- dna methylation
- binding protein
- south africa
- insulin resistance
- high fat diet induced
- genome wide
- endothelial cells
- transcription factor
- sensitive detection
- mouse model
- sars cov
- immune response
- adipose tissue
- label free