title: Live-cell coimaging of the genomic RNAs and Gag proteins of two lentiviruses.
abstract: Human immunodeficiency virus type 1 (HIV-1) Gag and genomic RNA determinants required for encapsidation are well established, but where and when encapsidation occurs in the cell is unknown. We constructed MS2 phage coat protein labeling systems to track spatial dynamics of primate and nonprimate lentiviral genomic RNAs (HIV-1 and feline immunodeficiency virus [FIV]) vis-à-vis their Gag proteins in live cells. Genomic RNAs of both lentiviral genera were observed to traffic into the cytoplasm, and this was Rev dependent. In transit, FIV Gag and genomic RNA accumulated independently of each other at the nuclear envelope, and focal colocalizations of genomic RNA with an intact packaging signal (psi) and Gag were observed to extend outward from the cytoplasmic face. In contrast, although HIV-1 genomic RNA was detected at the nuclear envelope, HIV-1 Gag was not. For both lentiviruses, genomic RNAs were seen at the plasma membrane if and only if Gag was present and psi was intact. In addition, HIV-1 and FIV genomes accumulated with Gag in late endosomal foci, again, only psi dependently. Thus, lentiviral genomic RNAs require specific Gag binding to accumulate at the plasma membrane, packaged genomes cointernalize with Gag into the endosomal pathway, and plasma membrane RNA incorporation by Gag does not trigger committed lentiviral particle egress from the cell. Based on the FIV results, we hypothesize that the Gag-genome association may initiate at the nuclear envelope.
authors.affiliation: Department of Molecular Medicine, Guggenheim 18, Mayo Clinic College of Medicine, 200 First Street SW, Rochester, MN 55905, USA.
authors.forename: Iris
authors.initials: I
authors.lastname: Kemler
authors.forename: Anne
authors.initials: A
authors.lastname: Meehan
authors.forename: Eric M
authors.initials: EM
authors.lastname: Poeschla
mesh_terms: D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry; D002467:Cell Nucleus / Q000737:chemistry; D003593:Cytoplasm / Q000737:chemistry; D011992:Endosomes / Q000737:chemistry; D015683:Gene Products, gag / Q000378:metabolism*; D015497:HIV-1 / Q000502:physiology*; D006801:Humans; D007091:Image Processing, Computer-Assisted / Q000379:methods; D016089:Immunodeficiency Virus, Feline / Q000502:physiology*; D012367:RNA, Viral / Q000378:metabolism*; D019065:Virus Assembly
chemical_list: D015683:Gene Products, gag; D012367:RNA, Viral
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