1. A New Generation of Functional Tagged Proteins for HIV Fluorescence Imaging
- Author
-
Joseph Griffin, Thomas J. Hope, Stéphanie Gambut, and João I. Mamede
- Subjects
0301 basic medicine ,Fluorescence-lifetime imaging microscopy ,viruses ,030106 microbiology ,lcsh:QR1-502 ,Human immunodeficiency virus (HIV) ,HIV Infections ,Computational biology ,medicine.disease_cause ,Virus Replication ,lcsh:Microbiology ,Virus ,Fluorescence ,Article ,Cell Line ,03 medical and health sciences ,Viral Proteins ,Virology ,Cell Line, Tumor ,Fluorescence microscope ,medicine ,Humans ,Infectivity ,Expression vector ,fluorescent HIV ,HIV early-steps ,biology ,Integrases ,Chemistry ,integration competent tagged viruses ,Optical Imaging ,Virion ,Phenotype ,Integrase ,030104 developmental biology ,Infectious Diseases ,HEK293 Cells ,biology.protein ,HIV-1 ,HeLa Cells - Abstract
During the last decade, there was a marked increase in the development of tools and techniques to study the molecular mechanisms of the HIV replication cycle by using fluorescence microscopy. Researchers often apply the fusion of tags and fluorophores to viral proteins, surrogate proteins, or dyes to follow individual virus particles while they progress throughout infection. The inclusion of such fusion motifs or surrogates frequently disrupts viral infectivity or results in a change of the wild-type phenotype. Here, we detail the construction and functional characterization of two new constructs where we fused fluorescent proteins to the N-terminus of HIV-1 Integrase. In the first, IN is recruited into assembling particles via a codon optimized Gag to complement other viral constructs, while the second is fused to a Gag-Pol expression vector fully capable of integration. Our data shows that N-terminal tagged IN is functional for integration by both recovery of integration of catalytically inactive IN and by the successful infectivity of viruses carrying only labeled IN. These tools will be important to study the individual behavior of viral particles and associate such behavior to infectivity.
- Published
- 2021