Turning up the volume on mutational pressure: Is more of a good thing always better? (A case study of HIV-1 Vif and APOBEC3)

APOBEC3G and APOBEC3F are human cytidine deaminases that serve as innate antiviral defense mechanisms primarily by introducing C-to-U changes in the minus strand DNA of retroviruses during replication (resulting in G-to-A mutations in the genomic sense strand sequence). The HIV-1 Vif protein counteracts this defense by promoting the proteolytic degradation of APOBEC3G and APOBEC3F in the host cell. In the absence of Vif expression, APOBEC3 is incorporated into HIV-1 virions and the viral genome undergoes extensive G-to-A mutation, or "hypermutation", typically rendering it non-viable within a single replicative cycle. Consequently, Vif is emerging as an attractive target for pharmacological intervention and therapeutic vaccination. Although a highly effective Vif inhibitor may result in mutational meltdown of the viral quasispecies, a partially effective Vif inhibitor may accelerate the evolution of drug resistance and immune escape due to the codon structure and recombinogenic nature of HIV-1. This hypothesis rests on two principal assumptions which are supported by experimental evidence: a) there is a dose response between intracellular APOBEC concentration and degree of viral hypermutation, and, b) HIV-1 can tolerate an elevated mutation rate, and a true error or extinction threshold is as yet undetermined. Rigorous testing of this hypothesis will have timely and critical implications for the therapeutic management of HIV/AIDS, and delve into the complexities underlying the induction of lethal mutagenesis in a viral pathogen

QT peak prolongation predicts cardiac death following stroke

Cardiac death has been linked in many populations to prolongation of the QT interval (QTe). However, basic science research suggested that the best estimate of the time point when repolarisation begins is near the T-wave peak. We found QT peak (QTp) was longer in hypertensive subjects with LVH. A prolonged “depolarisation” phase, rather than “repolarisation” (T peak to T end) might therefore account for the higher incidence of cardiac death linked to long QT. Hypothesis: We have tested the hypothesis that QT peak (QTp) prolongation predicts cardiac death in stroke survivors. Methods and Results: ECGs were recorded from 296 stroke survivors (152 male), mean age 67.2 (SD 11.6) approximately 1 year after the event. Their mean blood pressure was 152/88 mmHg (SD 29/15mmHg). These ECGs were digitised by one observer who was blinded to patient outcome. The patients were followed up for a median of 3.3 years. The primary endpoint was cardiac death. A prolonged heart rate corrected QT peak (QTpc) of lead I carried the highest relative risk of death from all cause as well as cardiac death, when compared with the other more conventional QT indices. In multivariate analyses, when adjusted for conventional risk factors of atherosclerosis, a prolonged QTpc of lead I was still associated with a 3-fold increased risk of cardiac death. (adjusted relative risk 3.0 [95% CI 1.1 - 8.5], p=0.037). Conclusion: QT peak prolongation in lead I predicts cardiac death after strok

HIV Exploits Antiviral Host Innate GCN2-ATF4 Signaling for Establishing Viral Replication Early in Infection.

Antiviral innate host defenses against acute viral infections include suppression of host protein synthesis to restrict viral protein production. Less is known about mechanisms by which viral pathogens subvert host antiviral innate responses for establishing their replication and dissemination. We investigated early innate defense against human immunodeficiency virus (HIV) infection and viral evasion by utilizing human CD4+ T cell cultures in vitro and a simian immunodeficiency virus (SIV) model of AIDS in vivo Our data showed that early host innate defense against the viral infection involves GCN2-ATF4 signaling-mediated suppression of global protein synthesis, which is exploited by the virus for supporting its own replication during early viral infection and dissemination in the gut mucosa. Suppression of protein synthesis and induction of protein kinase GCN2-ATF4 signaling were detected in the gut during acute SIV infection. These changes diminished during chronic viral infection. HIV replication induced by serum deprivation in CD4+ T cells was linked to the induction of ATF4 that was recruited to the HIV long terminal repeat (LTR) to promote viral transcription. Experimental inhibition of GCN2-ATF4 signaling either by a specific inhibitor or by amino acid supplementation suppressed the induction of HIV expression. Enhancing ATF4 expression through selenium administration resulted in reactivation of latent HIV in vitro as well as ex vivo in the primary CD4+ T cells isolated from patients receiving suppressive antiretroviral therapy (ART). In summary, HIV/SIV exploits the early host antiviral response through GCN2-ATF4 signaling by utilizing ATF4 for activating the viral LTR transcription to establish initial viral replication and is a potential target for HIV prevention and therapy.IMPORTANCE Understanding how HIV overcomes host antiviral innate defense response in order to establish infection and dissemination is critical for developing prevention and treatment strategies. Most investigations focused on the viral pathogenic mechanisms leading to immune dysfunction following robust viral infection and dissemination. Less is known about mechanisms that enable HIV to establish its presence despite rapid onset of host antiviral innate response. Our novel findings provide insights into the viral strategy that hijacks the host innate response of the suppression of protein biosynthesis to restrict the virus production. The virus leverages transcription factor ATF4 expression during the GCN2-ATF4 signaling response and utilizes it to activate viral transcription through the LTR to support viral transcription and production in both HIV and SIV infections. This unique viral strategy is exploiting the innate response and is distinct from the mechanisms of immune dysfunction after the critical mass of viral loads is generated

Heterogeneity in HIV and cellular transcription profiles in cell line models of latent and productive infection: implications for HIV latency.

BackgroundHIV-infected cell lines are widely used to study latent HIV infection, which is considered the main barrier to HIV cure. We hypothesized that these cell lines differ from each other and from cells from HIV-infected individuals in the mechanisms underlying latency.ResultsTo quantify the degree to which HIV expression is inhibited by blocks at different stages of HIV transcription, we employed a recently-described panel of RT-ddPCR assays to measure levels of 7 HIV transcripts ("read-through," initiated, 5' elongated, mid-transcribed/unspliced [Pol], distal-transcribed [Nef], polyadenylated, and multiply-sliced [Tat-Rev]) in bulk populations of latently-infected (U1, ACH-2, J-Lat) and productively-infected (8E5, activated J-Lat) cell lines. To assess single-cell variation and investigate cellular genes associated with HIV transcriptional blocks, we developed a novel multiplex qPCR panel and quantified single cell levels of 7 HIV targets and 89 cellular transcripts in latently- and productively-infected cell lines. The bulk cell HIV transcription profile differed dramatically between cell lines and cells from ART-suppressed individuals. Compared to cells from ART-suppressed individuals, latent cell lines showed lower levels of HIV transcriptional initiation and higher levels of polyadenylation and splicing. ACH-2 and J-Lat cells showed different forms of transcriptional interference, while U1 cells showed a block to elongation. Single-cell studies revealed marked variation between/within cell lines in expression of HIV transcripts, T cell phenotypic markers, antiviral factors, and genes implicated in latency. Expression of multiply-spliced HIV Tat-Rev was associated with expression of cellular genes involved in activation, tissue retention, T cell transcription, and apoptosis/survival.ConclusionsHIV-infected cell lines differ from each other and from cells from ART-treated individuals in the mechanisms governing latent HIV infection. These differences in viral and cellular gene expression must be considered when gauging the suitability of a given cell line for future research on HIV. At the same time, some features were shared across cell lines, such as low expression of antiviral defense genes and a relationship between productive infection and genes involved in survival. These features may contribute to HIV latency or persistence in vivo, and deserve further study using novel single cell assays such as those described in this manuscript

Presumed Diffuse Unilateral Subacute Neuroretinitis: A Review Supported by a Unique Case

Diffuse unilateral subacute neuroretinitis (DUSN) is a rare, sight-threatening inflammatory condition caused by a nematode. It typically affects healthy children and young adults. Various nematodes have been implicated, including those that originate from dogs and raccoons. Late stage findings include optic nerve atrophy, retinal arterial attenuation, “retinitis pigmentosa-like” retinal pigmentary changes, and central and peripheral vision loss. We present a case study of long-standing DUSN. The combination of optical coherence tomography (OCT) and visual electrophysiology testing proved invaluable in establishing a presumptive diagnosis of this rare condition

The maize INDETERMINATE1 flowering time regulator defines a highly conserved zinc finger protein family in higher plants

BACKGROUND: The maize INDETERMINATE1 gene, ID1, is a key regulator of the transition to flowering and the founding member of a transcription factor gene family that encodes a protein with a distinct arrangement of zinc finger motifs. The zinc fingers and surrounding sequence make up the signature ID domain (IDD), which appears to be found in all higher plant genomes. The presence of zinc finger domains and previous biochemical studies showing that ID1 binds to DNA suggests that members of this gene family are involved in transcriptional regulation. RESULTS: Comparison of IDD genes identified in Arabidopsis and rice genomes, and all IDD genes discovered in maize EST and genomic databases, suggest that ID1 is a unique member of this gene family. High levels of sequence similarity amongst all IDD genes from maize, rice and Arabidopsis suggest that they are derived from a common ancestor. Several unique features of ID1 suggest that it is a divergent member of the maize IDD family. Although no clear ID1 ortholog was identified in the Arabidopsis genome, highly similar genes that encode proteins with identity extending beyond the ID domain were isolated from rice and sorghum. Phylogenetic comparisons show that these putative orthologs, along with maize ID1, form a group separate from other IDD genes. In contrast to ID1 mRNA, which is detected exclusively in immature leaves, several maize IDD genes showed a broad range of expression in various tissues. Further, Western analysis with an antibody that cross-reacts with ID1 protein and potential orthologs from rice and sorghum shows that all three proteins are detected in immature leaves only. CONCLUSION: Comparative genomic analysis shows that the IDD zinc finger family is highly conserved among both monocots and dicots. The leaf-specific ID1 expression pattern distinguishes it from other maize IDD genes examined. A similar leaf-specific localization pattern was observed for the putative ID1 protein orthologs from rice and sorghum. These similarities between ID1 and closely related genes in other grasses point to possible similarities in function

Attenuated and Nonproductive Viral Transcription in the Lymphatic Tissue of HIV-1-Infected Patients Receiving Potent Antiretroviral Therapy

Human immunodeficiency virus type 1 (HIV-1) RNA that persists in the lymphoid tissue of patients despite treatment with highly active antiretroviral therapy (HAART) may represent extracellular virions or intracellular RNAs residing within HIV-infected cells. To further characterize residual viral transcription, tonsil biopsy specimens from patients receiving long-term HAART, untreated patients, and patients undergoing 2 weeks of structured treatment interruption were analyzed by polymerase chain reaction quantification of virion-encapsidated RNA, intracellular unspliced HIV RNA (HIV UsRNA), multiply spliced HIV RNA encoding tat and rev (HIV MsRNA), and HIV DNA. Tonsil biopsy specimens from viremic patients harbored high amounts of virions, which primarily stemmed from local production, as indicated by a strong correlation of extracellular tonsillar RNA with intracellular HIV-1 nucleic acid levels but not with plasma viremia, and as shown by phylogenetic analysis of clonal env sequences from lymphoid tissue and plasma. In patients receiving HAART, intracellular HIV UsRNA persisted at significantly decreased levels, whereas HIV MsRNA and lymphoid virion levels were depleted. Thus, residual lymphoid HIV-1 RNA in patients receiving HAART indicates attenuated viral transcription in HIV-1-infected cells that lack virion productio

Loss of H2A.Z Is Not Sufficient to Determine Transcriptional Activity of Snf2-Related CBP Activator Protein or p400 Complexes

The p400 and SRCAP (Snf2-related CBP activator protein) complexes remodel chromatin by catalyzing deposition of histone H2A.Z into nucleosomes. This remodeling activity has been proposed as a basis for regulation of transcription by these complexes. Transcript levels of p21 or Sp1 mRNAs after knockdown of p400 or SRCAP reveals that each regulates transcription of these promoters differently. In this study, we asked whether deposition of H2A.Z within specific nucleosomes by p400 or SRCAP dictates transcriptional activity. Our data indicates that nucleosome density at specific p21 or Sp1 promoter positions is not altered by the loss of either remodeling complex. However, knockdown of SRCAP or p400 reduces deposition of H2A.Z∼50% into all p21 and Sp1 promoter nucleosomes. Thus, H2A.Z deposition is not targeted to specific nucleosomes. These results indicate that the deposition of H2A.Z by the p400 or SRCAP complexes is not sufficient to determine how each regulates transcription. This conclusion is further supported by studies that demonstrate a SRCAPΔATP mutant unable to deposit H2A.Z has similar transcriptional activity as wild-type SRCAP