381 Molecular Mechanisms for HIV-1 Restriction in Human Astroglia Roger Pomerantz*, J Fang, E Acheampong, and M Mukhtar Thomas Jefferson Univ, Philadelphia, PA, USA

Background:  Latent and restricted HIV-1 replication has been demonstrated in CD4⁺ T lymphocytes and monocyte/macrophages in patients on virally suppressive HAART. Nevertheless, new data suggest that other cell types may also function as residual disease sites in patients. Astrocytes in the central nervous system have been shown to harbor low levels of HIV-1 replication, with many studies, both in vitro and in vivo, suggesting that multiply spliced out-of-proportion to unspliced HIV-1 RNA is present in these cells, as a transcriptionally active but translationally restricted or Rev-deficient phenotype. 

Methods:  Our analyses were conducted using primary human fetal astrocytes, Western blotting and RT-PCR assays for Rev and DDX1, and siRNA knock-outs.

Results:  Recent studies from our laboratories have demonstrated that a DEAD box protein, DDX1, has been shown to be an important cellular co-factor for the HIV-1 Rev:RRE axis. Our laboratories have also demonstrated that HIV-1 Rev has an unusual subcellular compartmentalization in human fetal astrocytes—cytoplasmic out-of-proportion to nuclear dominance. This is a similar phenotype to DDX1 knockout cells using RNA interference. New studies have now suggested that DDX1 may be the critical co-factor in the aberrant replication pattern of HIV-1 in human fetal astrocytes. DDX1 was demonstrated to be present in the nucleus of human fetal astrocytes and co-localized with HIV-1 Rev. Over-expression of DDX1 in primary human fetal astrocytes directly augmented HIV-1 replication. In addition, Rev function as defined by subgenomic RRE-containing constructs demonstrated the effects of down-regulating DDX1 with small interfering RNA (siRNA) in human fetal astrocytes and this down-regulation of DDX1 also led to a profound further decrease in HIV-1 replication. 

Conclusions:  As such, it is suggested that a Rev-deficient phenotype is demonstrable through complementary techniques in certain human fetal astrocytes and that the newly described Rev binding protein, DDX1, may be critical in explaining the molecular mechanisms behind this novel state of viral restriction and possibly a site for reservoir disease during therapy.

Keywords: Astrocytes; Rev; DDX1