Biology subjects

Luban, J.

Publications and source records attributed to Luban, J..

5 recordsLinked to original sources

bioRxiv2018

Constrained mutational sampling of amino acids in HIV-1 protease evolution

The evolution of HIV-1 protein sequences should be governed by a combination of factors including nucleotide mutational probabilities, the genetic code, and fitness. The impact of these factors on protein sequence evolution are interdependent, making it challenging to infer the individual contribution of each factor from phylogenetic analyses alone. We investigated the protein sequence evolution of HIV-1 by determining an experimental fitness landscape of all individual amino acid changes in protease. We compared our experimental results to the frequency of protease variants in a publicly available dataset of 32,163 sequenced isolates from drug-naive individuals. The most common amino acids in sequenced isolates supported robust experimental fitness, indicating that the experimental fitness landscape captured key features of selection acting on protease during viral infections of hosts. Amino acid changes requiring multiple mutations from the likely ancestor were slightly less likely to support robust experimental fitness than single mutations, consistent with the genetic code favoring chemically conservative amino acid changes. Amino acids that were common in sequenced isolates were predominantly accessible by single mutations from the likely protease ancestor. Multiple mutations commonly observed in isolates were accessible by mutational walks with highly fit single mutation intermediates. Our results indicate that the prevalence of multiple base mutations in HIV-1 protease is strongly influenced by mutational sampling.

evolutionary biology↗

bioRxiv2018

Primate immunodeficiency virus Vpx and Vpr counteract transcriptional repression of proviruses by the HUSH complex

Drugs that inhibit HIV-1 replication and prevent progression to AIDS do not eliminate HIV-1 proviruses from the chromosomes of long-lived CD4+ memory T cells. To escape eradication by these antiviral drugs, or by the host immune system, HIV-1 exploits poorly defined host factors that silence provirus transcription. These same factors, though, must be overcome by all retroviruses, including HIV-1 and other primate immunodeficiency viruses, in order to activate provirus transcription and produce new virus. Here we show that Vpx and Vpr, proteins from a wide range of primate immunodeficiency viruses, activate provirus transcription in human CD4+ T cells. Provirus activation required the DCAF1 adaptor that links Vpx and Vpr to the CUL4A/B ubiquitin ligase complex, but did not require degradation of SAMHD1, a well-characterized target of Vpx and Vpr. A loss-of-function screen for transcription silencing factors that mimic the effect of Vpx on provirus silencing identified all components of the Human Silencing Hub (HUSH) complex, FAM208A (TASOR/RAP140), MPHOSPH8 (MPP8), PPHLN1 (PERIPHILIN), and MORC2. Vpx associated with the HUSH complex components and decreased steady-state levels of these proteins in a DCAF-dependent manner. Finally, vpx and FAM208A knockdown accelerated HIV-1 and SIVMAC replication kinetics in CD4+ T cells to a similar extent, and HIV-2 replication required either vpx or FAM208A disruption. These results demonstrate that the HUSH complex restricts transcription of primate immunodeficiency viruses and thereby contributes to provirus latency. To counteract this restriction and activate provirus expression, primate immunodeficiency viruses encode Vpx and Vpr proteins that degrade HUSH complex components.

molecular biology↗

bioRxiv2018

OMEGA: a software tool for the management, analysis, and dissemination of intracellular trafficking data that incorporates motion type classification and quality control

MOTIVATIONParticle tracking coupled with time-lapse microscopy is critical for understanding the dynamics of intracellular processes of clinical importance. Spurred on by advances in the spatiotemporal resolution of microscopy and automated computational methods, this field is increasingly amenable to multi-dimensional high-throughput data collection schemes (Snijder et al., 2012). Typically, complex particle tracking datasets generated by individual laboratories are produced with incompatible methodologies that preclude comparison to each other. There is therefore an unmet need for data management systems that facilitate data standardization, meta-analysis, and structured data dissemination. The integration of analysis, visualization, and quality control capabilities into such systems would eliminate the need for manual transfer of data to diverse downstream analysis tools. At the same time, it would lay the foundation for shared trajectory data, particle tracking, and motion analysis standards. RESULTSHere, we present Open Microscopy Environment inteGrated Analysis (OMEGA), a cross-platform data management, analysis, and visualization system, for particle tracking data, with particular emphasis on results from viral and vesicular trafficking experiments. OMEGA provides intuitive graphical interfaces to implement integrated particle tracking and motion analysis workflows while providing easy to use facilities to automatically keep track of error propagation, harvest data provenance and ensure the persistence of analysis results and metadata. Specifically, OMEGA: 1) imports image data and metadata from data management tools such as the Open Microscopy Environment Remote Objects (OMERO; Allan et al., 2012); 2) tracks intracellular particles movement; 3) facilitates parameter optimization and trajectory results inspection and validation; 4) performs downstream trajectory analysis and motion type classification; 5) estimates the uncertainty propagating through the motion analysis pipeline; and, 6) facilitates storage and dissemination of analysis results, and analysis definition metadata, on the basis of our newly proposed FAIRsharing.org complainant Minimum Information About Particle Tracking Experiments (MIAPTE; Rigano and Strambio-De-Castillia, 2016; 2017) guidelines in combination with the OME-XML data model (Goldberg et al., 2005). In so doing, OMEGA maintains a persistent link between raw image data, intermediate analysis steps, the overall analysis output, and all necessary metadata to repeat the analysis process and reproduce its results. Availability and implementationOMEGA is a cross-platform, open-source software developed in Java. Source code and cross-platform binaries are freely available on GitHub at https://github.com/OmegaProject/Omega (doi: 10.5281/zenodo.2535523), under the GNU General Public License v.3. Contactcaterina.strambio@umassmed.edu and alex.rigano@umassmed.edu Supplementary informationSupplementary Material is available at BioRxiv.org

bioinformatics↗

bioRxiv2017

HIV-1 unmasks the plasticity of innate lymphoid cells

HIV-1-infected people who take medications that suppress viremia, preserve CD4+ T cells, and prevent AIDS, have chronic inflammation with increased cardiovascular mortality. To investigate the etiology of this inflammation, the effect of HIV-1 on innate lymphoid cells (ILCs) and NK cells was examined. Homeostatic ILCs in blood and intestine were depleted permanently. NK cells were skewed towards a memory subset. Cytokines that are elevated during HIV-1 infection reproduced both abnormalities ex vivo. Pseudotime analysis of single NK cell transcriptomes revealed a developmental trajectory towards a subset with expression profile, chromatin state, and biological function like memory T lymphocytes. Expression of TCF7, a WNT transcription factor, increased over the course of the trajectory. TCF7 disruption, or WNT inhibition, prevented memory NK cell induction by inflammatory cytokines. These results demonstrate that inflammatory cytokines associated with HIV-1 infection irreversibly disrupt homeostatic ILCs and cause developmental shift towards TCF7+ memory NK cells.\n\nHighlightsO_LIHIV-1 infection depletes homeostatic ILCs in blood and intestine and shifts NK cells towards a memory cell phenotype, irrespective of viremia or CD4 count\nC_LIO_LIInflammatory cytokines recapitulate ILC and NK cell abnormalities ex vivo\nC_LIO_LITCF7 expression correlates with a developmental trajectory that culminates in memory NK cells\nC_LIO_LITCF7/WNT signaling is required for establishment of memory NK cells\nC_LI\n\n\n\nO_FIG O_LINKSMALLFIG WIDTH=172 HEIGHT=200 SRC=\"FIGDIR/small/221010v3_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (32K):\norg.highwire.dtl.DTLVardef@ae82a7org.highwire.dtl.DTLVardef@103b2b4org.highwire.dtl.DTLVardef@1c9585aorg.highwire.dtl.DTLVardef@1c7bb2d_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO C_FIG

immunology↗

bioRxiv2017

Intron-containing RNA from the HIV-1 provirus activates type I interferon and inflammatory cytokines

HIV-1-infected people who take drugs that suppress viremia to undetectable levels are protected from developing AIDS. Nonetheless, these individuals have chronic inflammation associated with heightened risk of cardiovascular pathology. HIV-1 establishes proviruses in long-lived CD4+ memory T cells, and perhaps other cell types, that preclude elimination of the virus even after years of continuous antiviral therapy. Though the majority of proviruses that persist during antiviral therapy are defective for production of infectious virions, many are expressed, raising the possibility that the HIV-1provirus or its transcripts contribute to ongoing inflammation. Here we found that the HIV-1 provirus activated innate immune signaling in isolated dendritic cells, macrophages, and CD4+ T cells. Immune activation required transcription from the HIV-1 provirus and expression of CRM1-dependent, Rev-dependent, RRE-containing, unspliced HIV-1 RNA. If rev was provided in trans, all HIV-1 coding sequences were dispensable for activation except those cis-acting sequences required for replication or splicing. These results indicate that the complex, post-transcriptional regulation intrinsic to HIV-1 RNA is detected by the innate immune system as a danger signal, and that drugs which disrupt HIV-1 transcription or HIV-1 RNA metabolism would add qualitative benefit to current antiviral drug regimens.

microbiology↗