Accumulation of defective viral genomes in peripheral blood mononuclear cells of human immunodeficiency virus type 1-infected individuals

Abstract

Human immunodeficiency virus type 1 (HIV-1) genomes present in peripheral blood mononuclear cells (PBMCs) of infected persons or in lymphocytes infected in vitro were studied by long-distance PCR (LD-PCR) using primers localized in the HIV-1 long terminal repeats. The full-length 9-kb DNA was the only LD-PCR product obtained in peripheral and cord blood lymphocytes from seronegative donors infected in vitro. However, a high proportion (27% to 66%) of distinct populations of extensively deleted HIV-1 genomes of variable size was detected in PBMCs of 15 of 16 HIV-1-infected persons. Physical mapping of defective genomes showed that the frequency of deletions is proportional to their proximity to the central part of HIV-1 genome, which is consistent with a deletion mechanism involving a single polymerase jump during reverse transcription. Sequencing of deletion junctions revealed the presence of short direct repeats of three or four nucleotides. The number of defective HIV-1 genomes decreased after in vitro activation of PBMCs. Persistence of full-length and deleted genomes in in vitro activated PBMCs correlated with isolation of an infectious virus. Our results represent the first quantitative assessment of intragenomic rearrangements in HIV-1 genomes in PBMCs of infected persons and demonstrate that, in contrast to in vitro infection, defective genomes accumulate in PBMCs of infected persons.

Longitudinal study reveals HIV-1–infected CD4+ T cell dynamics during long-term antiretroviral therapy

“…These deletions vary widely in length but typically encompass multiple viral genes (13)(14)(15)(16)22). Deletions likely arise during reverse transcription when the polymerase jumps to a homologous region on the same template during negative-strand synthesis, but other mechanisms may contribute (16,(22)(23)(24). Other less common defects include frameshift mutations and inverted segments of the HIV-1 genome inserted within a large multigenic deletion (14).…”

Conservation of the central proline‐rich (PxxP) motifs of human immunodeficiency virus type 1 Nef protein during the disease progression in two hemophiliac patients

“…In spite of extensive investigations of the nef sequence heterogeneity, selective conservation of these regions during the disease progression in vivo has not been previously noted [33–35,39]. Since in these studies the nef gene was amplified directly from the patient plasma or the infected cells, it is likely that these nef sequences did not represent the replication‐competent virus [40–43]. Although these defective viruses may kill the infected cells, they do not produce progeny virions and thus do not contribute to the disease progression.…”

Reservoirs of Human Immunodeficiency Virus Type 1: The Main Obstacles to Viral Eradication

“…It is of note that a study has demonstrated that, in some patients who are receiving virally suppressive HAART with transient spikes of virus of 150 copies/mL in plasma, resistance mutations may develop [21]. Data from the aforementioned studies demonstrated that, although defective proviruses accumulate in CD4 ϩ T lymphocytes in vivo (i.e., "viral graveyard sequences") [22], replication-competent provirus still exists in resting CD4 ϩ T lymphocytes, which may hinder attempts at reducing the viral reservoir and may reseed the body with virus if HAART is discontinued. These possibilities, therefore, confound our attempts toward pharmacologically mediated virus eradication.…”