Transcriptional competence of the integrated HIV-1 provirus at the nuclear periphery.

Spatial distribution of genes within the nucleus contributes to transcriptional control, allowing optimal gene expression as well as constitutive or regulated gene repression. Human immunodeficiency virus type 1 (HIV-1) integrates into host chromatin to transcribe and replicate its genome. Lymphocytes harbouring a quiescent but inducible provirus are a challenge to viral eradication in infected patients undergoing antiviral therapy. Therefore, our understanding of the contribution of sub-nuclear positioning to viral transcription may also have far-reaching implications in the pathology of the infection. To gain an insight into the conformation of chromatin at the site of HIV-1 integration, we investigated lymphocytes carrying a single latent provirus. In the silenced state, the provirus was consistently found at the nuclear periphery, associated in trans with a pericentromeric region of chromosome 12 in a significant number of quiescent cells. After induction of the transcription, this association was lost, although the location of the transcribing provirus remained peripheral. These results, extended to several other cell clones, unveil a novel mechanism of transcriptional silencing involved in HIV-1 post-transcriptional latency and reinforce the notion that gene transcription may also occur at the nuclear periphery.

High prevalence of secondary resistance mutations in Venezuelan HIV-1 isolates.

The genetic variability was studied in HIV-1 from Venezuelan patients with and without treatment, in order to evaluate the presence of polymorphisms and drug resistance mutations. Proviral DNA from peripheral blood mononuclear cells or viral RNA from plasma was extracted from the blood of 30 patients. Two regions from the polymerase gene, protease (Pr) and reverse transcriptase (RT) and one genomic fragment from the envelope (Env) gene were amplified and sequenced. All HIV-1 samples analyzed were classified as subtype B, without evidence of recombination. Although no primary protease mutations were detected, a high frequency of secondary mutations (86%, 19/22), associated to restoration of viral replicative fitness, was observed in strains circulating both in treated and non-treated patients. Resistance mutations to nucleoside RT inhibitors (NRTI) and non-nucleoside RT inhibitors (NNRTI) were detected in 35% (6/17) and 12% (2/17) of the viruses circulating in treated patients, respectively. Resistance mutations were also present in the virus infecting one antiretroviral naive individual (7.7%), suggesting that local screening for resistant mutation in naive patient might be important to minimize therapy failure. Future studies are warranted to assess the role of secondary mutation in the success of viral infection.

Nuclear positional control of HIV transcription in 4D.

Retroviruses integrate their genome into the chromatin of the host cell and are subject to the same control mechanisms governing transcription in the nucleus. There is increasing evidence that the spatial position of a gene within the nucleus in time affects its activity. Therefore it becomes important to study the chromatin environment in space and time of the HIV-1 provirus, particularly in cells where a tight transcriptional control allows the virus to hide away from antiviral treatment and immune response. We recently showed that the HIV-1 provirus is found at the nuclear periphery of latently infected lymphocytes associated in trans with centromeric heterochromatin. After induction of transcription, this association was lost, although the location of the transcribing provirus remained peripheral. Our results reveal a novel mechanism of transcriptional silencing involved in HIV-1 post-transcriptional latency and open wider perspectives for the general organization of chromatin in the nucleus.

High prevalence of secondary resistance mutations in Venezuelan HIV-1 isolates

Se estudiaron pacientes seropositivos para el virus de inmunodeficiencia humana tipo 1 (VIH-1) con y sin tratamiento, con el fin de determinar el polimorfismo y la prevalencia de mutaciones de resistencia a la terapia antirretroviral. El material genético viral fue extraído a partir de células mononucleares de sangre periféricas (ADN) y del plasma (ARN) de 30 pacientes. Se amplificaron 2 regiones del gen Pol, Transcriptasa Reversa (TR) y Proteasa (Pr) y el gen de envoltura (Env) por medio de la técnica de PCR y se obtuvo la secuencia genómica de los productos. Todos los aislados analizados pertenecieron al subtipo B. No se observaron mutaciones primarias asociadas a resistencia a inhibidores de Pr pero sí un alto porcentaje (86 por ciento, 19/22) de mutaciones no asociadas con resistencia sino a restitución de la capacidad replicativa de cepas mutantes (mutaciones secundarias). Se observó la presencia de mutaciones asociadas a resistencia a inhibidores nucleósidos de la TR (INTR) en 35 por ciento (6/17) de los pacientes sometidos a tratamiento, mientras que 12 por ciento (2/17) de ellos presentaron mutaciones de resistencia a inhibidores no nucleósidos de la TR (INNTR). Interesantemente, un paciente no tratado estaba infectado con una cepa que presentaba mutaciones primarias (7,7 por ciento); este resultado sugiere que podría ser importante plantearse el estudio local de determinación de resistencia genotípica en pacientes antes del tratamiento, con miras a minimizar fallas terapéuticas. Se requieren estudios adicionales para evaluar el rol de las mutaciones secundarias en el éxito de la infección viral