Molecular basis for nuclear accumulation and targeting of the inhibitor of apoptosis BIRC2.

The inhibitor of apoptosis protein BIRC2 regulates fundamental cell death and survival signaling pathways. Here we show that BIRC2 accumulates in the nucleus via binding of its second and third BIR domains, BIRC2BIR2 and BIRC2BIR3, to the histone H3 tail and report the structure of the BIRC2BIR3-H3 complex. RNA-seq analysis reveals that the genes involved in interferon and defense response signaling and cell-cycle regulation are most affected by depletion of BIRC2. Overexpression of BIRC2 delays DNA damage repair and recovery of the cell-cycle progression. We describe the structural mechanism for targeting of BIRC2BIR3 by a potent but biochemically uncharacterized small molecule inhibitor LCL161 and demonstrate that LCL161 disrupts the association of endogenous BIRC2 with H3 and stimulates cell death in cancer cells. We further show that LCL161 mediates degradation of BIRC2 in human immunodeficiency virus type 1-infected human CD4+ T cells. Our findings provide mechanistic insights into the nuclear accumulation of and blocking BIRC2.

HIV-1 Tat Upregulates TREM1 Expression in Human Microglia.

Because microglia are a reservoir for HIV and are resistant to the cytopathic effects of HIV infection, they are a roadblock for any HIV cure strategy. We have previously identified that triggering receptor expressed on myeloid cells 1 (TREM1) plays a key role in human macrophage resistance to HIV-mediated cytopathogenesis. In this article, we show that HIV-infected human microglia express increased levels of TREM1 and are resistant to HIV-induced apoptosis. Moreover, upon genetic inhibition of TREM1, HIV-infected microglia undergo cell death in the absence of increased viral or proinflammatory cytokine expression or the targeting of uninfected cells. We also show that the expression of TREM1 is mediated by HIV Tat through a TLR4, TICAM1, PG-endoperoxide synthase 2, PGE synthase, and PGE2-dependent manner. These findings highlight the potential of TREM1 as a therapeutic target to eradicate HIV-infected microglia without inducing a proinflammatory response.

IRAK1 inhibition blocks the HIV-1 RNA mediated pro-inflammatory cytokine response from microglia.

Human immunodeficiency virus (HIV)-associated neurocognitive disorders (HAND) are a common source of morbidity in people living with HIV (PLWH). Although antiretroviral therapy (ART) has lessened the severity of neurocognitive disorders, cognitive impairment still occurs in PLWH receiving ART. The pathogenesis of HAND is likely multifaceted, but common factors include the persistence of HIV transcription within the central nervous system, higher levels of pro-inflammatory cytokines in the cerebrospinal fluid, and the presence of activated microglia. Toll-like receptor (TLR) 7 and TLR8 are innate pathogen recognition receptors located in microglia and other immune and non-immune cells that can recognise HIV RNA and trigger pro-inflammatory responses. IL-1 receptor-associated kinase (IRAK) 1 is key to these signalling pathways. Here, we show that IRAK1 inhibition inhibits the TLR7 and TLR8-dependent pro-inflammatory response to HIV RNA. Using genetic and pharmacological inhibition, we demonstrate that inhibition of IRAK1 prevents IRAK1 phosphorylation and ubiquitination, and the subsequent recruitment of TRAF6 and the TAK1 complex to IRAK1, resulting in the inhibition of downstream signalling and the suppression of pro-inflammatory cytokine and chemokine release.

Development of Bread Dough by Sheeting: Effects of Sheeting Regime, Bran Level and Bran Particle Size.

The effects of sheeting on bread dough development and baked loaf quality were investigated, using Dynamic Dough Density and springback to quantify development, and examining effects of the sheeting regime on bread quality in terms of loaf volume and crumb structure. Bread doughs, with and without bran at different levels and particle sizes, were formed through a short mixing period, then sheeted through a benchtop manual sheeter at roll gaps of 6, 9 and 12 mm for different numbers of sheeting passes. The sheeting of doughs without bran increased dough expansion and baked loaf volume up to 12 sheeting passes. Loaves were larger after sheeting at a 6 mm roll gap, reflecting the greater gluten development at the smaller gap, although the crumb structure was less fine, with fewer gas cells and larger average gas cell diameters. The addition of bran decreased dough expansion and loaf volumes, with Fine bran and Coarse bran both more damaging than Medium bran, indicating the opportunity to optimise bran particle size to maximise bread quality. Sheeting was effective in alleviating the damaging effects of bran, with sheeting for 8 passes giving more dough expansion, larger loaf volumes and finer crumb structures than sheeting for 12 passes, indicating an even more damaging effect of bran when gluten is overstretched by sheeting. The work demonstrates the opportunity to enhance bread quality, particularly of healthy high-fibre breads, by employing sheeting to enhance gluten development and to offset the damage to gluten caused by the presence of bran.

Pacritinib Inhibition of IRAK1 Blocks Aberrant TLR8 Signalling by SARS-CoV-2 and HIV-1-Derived RNA.

Macrophages promote an early host response to infection by releasing pro-inflammatory cytokines such as interleukin (IL) 1ß (IL-1ß), tumour necrosis factor (TNF), and IL-6. One of the mechanisms through which cells sense pathogenic microorganisms is through Toll-like receptors (TLRs). IL-1 receptor-associated kinase (IRAK) 1, IRAK2, IRAK3, and IRAK4 are integral to TLR and IL-1 receptor signalling pathways. Recent studies suggest a role for aberrant TLR8 and NLRP3 inflammasome activation during both COVID-19 and HIV-1 infection. Here, we show that pacritinib inhibits the TLR8-dependent pro-inflammatory cytokine response elicited by GU-rich single-stranded RNA derived from SARS-CoV-2 and HIV-1. Using genetic and pharmacologic inhibition, we demonstrate that pacritinib inhibits IRAK1 phosphorylation and ubiquitination which then inhibits the recruitment of the TAK1 complex to IRAK1, thus inhibiting the activation of downstream signalling and the production of pro-inflammatory cytokines.

Current strategies to induce selective killing of HIV-1-infected cells.

Although combination antiretroviral therapy (ART) has led to significant HIV-1 suppression and improvement in immune function, persistent viral reservoirs remain that are refractory to intensified ART. ART poses many challenges such as adherence to drug regimens, the emergence of resistant virus, and cumulative toxicity resulting from long-term therapy. Moreover, latent HIV-1 reservoir cells can be stochastically activated to produce viral particles despite effective ART and contribute to the rapid viral rebound that typically occurs within 2 weeks of ART interruption; thus, lifelong ART is required for continued viral suppression. Several strategies have been proposed to address the HIV-1 reservoir such as reactivation of HIV-1 transcription using latency reactivating agents with a combination of ART, host immune clearance and HIV-1-cytotoxicity to purge the infected cells-a "shock and kill" strategy. However, these approaches do not take into account the multiple transcriptional and translational blocks that contribute to HIV-1 latency or the complex heterogeneity of the HIV-1 reservoir, and clinical trials have thus far failed to produce the desired results. Here, we describe alternative strategies being pursued that are designed to kill selectively HIV-1-infected cells while sparing uninfected cells in the absence of enhanced humoral or adaptive immune responses.

Amphibian mucus triggers a developmental transition in the frog-killing chytrid fungus.

The frog-killing chytrid fungus Batrachochytrium dendrobatidis (Bd) is decimating amphibian populations around the world.1-4Bd has a biphasic life cycle, alternating between motile zoospores that disperse within aquatic environments and sessile sporangia that grow within the mucus-coated skin of amphibians.5,6 Zoospores lack cell walls and swim rapidly through aquatic environments using a posterior flagellum and crawl across solid surfaces using actin structures similar to those of human cells.7,8Bd transitions from this motile dispersal form to its reproductive form by absorbing its flagellum, rearranging its actin cytoskeleton, and rapidly building a chitin-based cell wall-a process called "encystation."5-7 The resulting sporangium increases in volume by two or three orders of magnitude while undergoing rounds of mitosis without cytokinesis to form a large ceonocyte. The sporangium then cellurizes by dividing its cytoplasm into dozens of new zoospores. After exiting the sporangium through a discharge tube onto the amphibian skin, daughter zoospores can then reinfect the same individual or find a new host.5 Although encystation is critical to Bd growth, whether and how this developmental transition is triggered by external signals was previously unknown. We discovered that exposure to amphibian mucus triggers rapid and reproducible encystation within minutes. This response can be recapitulated with purified mucin, the bulk component of mucus, but not by similarly viscous methylcellulose or simple sugars. Mucin-induced encystation does not require gene expression but does require surface adhesion, calcium signaling, and modulation of the actin cytoskeleton. Mucus-induced encystation may represent a key mechanism for synchronizing Bd development with the arrival at the host.

A comparison of monitoring designs to assess wildlife community parameters across spatial scales.

Dedicated long-term monitoring at appropriate spatial and temporal scales is necessary to understand biodiversity losses and develop effective conservation plans. Wildlife monitoring is often achieved by obtaining data at a combination of spatial scales, ranging from local to broad, to understand the status, trends, and drivers of individual species or whole communities and their dynamics. However, limited resources for monitoring necessitates tradeoffs in the scope and scale of data collection. Careful consideration of the spatial and temporal allocation of finite sampling effort is crucial for monitoring programs that span multiple spatial scales. Here we evaluate the ability of five monitoring designs-stratified random, weighted effort, indicator unit, rotating panel, and split panel-to recover parameter values that describe the status (occupancy), trends (change in occupancy), and drivers (spatially varying covariate and an autologistic term) of wildlife communities at two spatial scales. Using an amphibian monitoring program that spans a network of US national parks as a motivating example, we conducted a simulation study for a regional community occupancy sampling program to compare the monitoring designs across varying levels of sampling effort (ranging from 10% to 50%). We found that the stratified random design outperformed the other designs for most parameters of interest at both scales and was thus generally preferable in balancing the estimation of status, trends, and drivers across scales. However, we found that other designs had improved performance in specific situations. For example, the rotating panel design performed best at estimating spatial drivers at a regional level. Thus, our results highlight the nuanced scenarios in which various design strategies may be preferred and offer guidance as to how managers can balance common tradeoffs in large-scale and long-term monitoring programs in terms of the specific knowledge gained. Monitoring designs that improve accuracy in parameter estimates are needed to guide conservation policy and management decisions in the face of broad-scale environmental challenges, but the preferred design is sensitive to the specific objectives of a monitoring program.

Geographic variation and thermal plasticity shape salamander metabolic rates under current and future climates.

Predicted changes in global temperature are expected to increase extinction risk for ectotherms, primarily through increased metabolic rates. Higher metabolic rates generate increased maintenance energy costs which are a major component of energy budgets. Organisms often employ plastic or evolutionary (e.g., local adaptation) mechanisms to optimize metabolic rate with respect to their environment. We examined relationships between temperature and standard metabolic rate across four populations of a widespread amphibian species to determine if populations vary in metabolic response and if their metabolic rates are plastic to seasonal thermal cues. Populations from warmer climates lowered metabolic rates when acclimating to summer temperatures as compared to spring temperatures. This may act as an energy saving mechanism during the warmest time of the year. No such plasticity was evident in populations from cooler climates. Both juvenile and adult salamanders exhibited metabolic plasticity. Although some populations responded to historic climate thermal cues, no populations showed plastic metabolic rate responses to future climate temperatures, indicating there are constraints on plastic responses. We postulate that impacts of warming will likely impact the energy budgets of salamanders, potentially affecting key demographic rates, such as individual growth and investment in reproduction.

Evaluating the effect of expert elicitation techniques on population status assessment in the face of large uncertainty.

Population projection models are important tools for conservation and management. They are often used for population status assessments, for threat analyses, and to predict the consequences of conservation actions. Although conservation decisions should be informed by science, critical decisions are often made with very little information to support decision-making. Conversely, postponing decisions until better information is available may reduce the benefit of a conservation decision. When empirical data are limited or lacking, expert elicitation can be used to supplement existing data and inform model parameter estimates. The use of rigorous techniques for expert elicitation that account for uncertainty can improve the quality of the expert elicited values and therefore the accuracy of the projection models. One recurring challenge for summarizing expert elicited values is how to aggregate them. Here, we illustrate a process for population status assessment using a combination of expert elicitation and data from the ecological literature. We discuss the importance of considering various aggregation techniques, and illustrate this process using matrix population models for the wood turtle (Glyptemys insculpta) to assist U.S. Fish and Wildlife Service decision-makers with their Species Status Assessment. We compare estimates of population growth using data from the ecological literature and four alternative aggregation techniques for the expert-elicited values. The estimate of population growth rate based on estimates from the literature (λmean = 0.952, 95% CI: 0.87-1.01) could not be used to unequivocally reject the hypotheses of a rapidly declining population nor the hypothesis of a stable, or even slightly growing population, whereas our results for the expert-elicited estimates supported the hypothesis that the wood turtle population will decline over time. Our results showed that the aggregation techniques used had an impact on model estimates, suggesting that the choice of techniques should be carefully considered. We discuss the benefits and limitations associated with each method and their relevance to the population status assessment. We note a difference in the temporal scope or inference between the literature-based estimates that provided insights about historical changes, whereas the expert-based estimates were forward looking. Therefore, conducting an expert-elicitation in addition to using parameter estimates from the literature improved our understanding of our species of interest.

Site- and Individual-Level Contaminations Affect Infection Prevalence of an Emerging Infectious Disease of Amphibians.

Emerging infectious disease outbreaks are one of multiple stressors responsible for amphibian declines globally. In the northeastern United States, ranaviral diseases are prevalent in amphibians and other ectothermic species, but there is still uncertainty as to whether their presence is leading to population-level effects. Further, there is also uncertainty surrounding the potential interactions among disease infection prevalence in free-ranging animals and habitat degradation (co-occurrence of chemical stressors). The present study was designed to provide field-based estimates of the relationship between amphibian disease and chemical stressors. We visited 40 wetlands across three protected areas, estimated the prevalence of ranavirus among populations of larval wood frogs and spotted salamanders, and assessed chemical and biological stressors in wetland habitats and larval amphibians using a suite of selected bioassays, screening tools, and chemical analyses. Ranavirus was detected on larval amphibians from each protected area with an estimated occupancy ranging from 0.27 to 0.55. Considerable variation in ranavirus occupancy was also observed within and among each protected area. Of the stressors evaluated, ranavirus prevalence was strongly and positively related to concentrations of metalloestrogens (metals with the potential to bind to estrogen receptors) and total metals in wetland sediments and weakly and negatively related to total pesticide concentrations in larval amphibians. These results can be used by land managers to refine habitat assessments to include such environmental factors with the potential to influence disease susceptibility. Environ Toxicol Chem 2022;41:781-791. © 2022 SETAC. This article has been contributed to by US Government employees and their work is in the public domain in the USA.

Enzyme pre-milling treatments improved milling performance of chickpeas by targeting mechanisms of seed coat and cotyledon adhesion with various effects on dhal quality.

BACKGROUND: Dehulling and splitting are important elements of the milling process to produce dhal from pulses. However, grain that is difficult-to-mill because of tightly adhered seed coats or cotyledons that resist separation makes it difficult to achieve high quality dhal. Milling yields are reduced, energy inputs into the milling process are increased, and the resulting dhal can be of poorer quality, chipped or abraded. RESULTS: Eight enzyme pre-treatments were chosen based on the hypothesised mechanisms of seed coat and cotyledon adhesion established previously. Using a difficult-to-mill chickpea (Cicer arietinum L.) genotype, we examined the effects of these pre-treatments, over time, on laboratory-scale milling performance and dhal quality. We pioneered a texture analyser method to measure the flex of the cotyledons and the force required to cleave the cotyledons. The enzyme-induced changes ranged from negative (tough seed coat, weight loss, deleterious colour and texture, increased visual damage to cotyledons and increased kibble loss, concave cotyledons, increased flex, and changes in taste) to positive (brittle seed coat, increased seed volume, improved dehulling efficiency and splitting yield, reduced cotyledon cleavage force, and acceptable dhal quality and taste). CONCLUSION: All pre-treatments improved milling performance compared to milling the raw seed, although there was considerable variation between them. Two pre-treatments showed no improvement in milling yields compared to the water control, and several pre-treatments resulted in unacceptable qualities. Three pre-treatments, endo-polygalacturonanase, α-galactosidase and cellulase, show potential for commercial milling applications and could assist pulse millers globally to achieve high quality dhal at the same time as minimising milling effort. © 2021 Society of Chemical Industry.

CD4+ T cell-mimicking nanoparticles encapsulating DIABLO/SMAC mimetics broadly neutralize HIV-1 and selectively kill HIV-1-infected cells.

HIV-1 is a major global health challenge. The development of an effective vaccine and a therapeutic cure are top priorities. The creation of vaccines that focus an antibody response toward a particular epitope of a protein has shown promise, but the genetic diversity of HIV-1 stymies this progress. Therapeutic strategies that provide effective and broad-spectrum neutralization against HIV-1 infection are highly desirable. Methods: We investigated the potential of nanoengineered CD4+ T cell membrane-coated nanoparticles (TNP) encapsulating the DIABLO/SMAC mimetics LCL-161 or AT-406 (also known as SM-406 or Debio 1143) to both neutralize HIV-1 and selectively kill HIV-1-infected resting CD4+ T cells and macrophages. Results: DIABLO/SMAC mimetic-loaded TNP displayed outstanding neutralizing breadth and potency, and selectively kill HIV-1-infected cells via autophagy-dependent apoptosis while having no drug-induced off-target or cytotoxic effects on bystander cells. Genetic inhibition of early stages of autophagy abolishes this effect. Conclusion: DIABLO/SMAC mimetic loaded TNP have the potential to be used as therapeutic agents to neutralize cell-free HIV-1 and to kill specifically HIV-1-infected cells as part of an HIV-1 cure strategy.

Induction of Autophagy to Achieve a Human Immunodeficiency Virus Type 1 Cure.

Effective antiretroviral therapy has led to significant human immunodeficiency virus type 1 (HIV-1) suppression and improvement in immune function. However, the persistence of integrated proviral DNA in latently infected reservoir cells, which drive viral rebound post-interruption of antiretroviral therapy, remains the major roadblock to a cure. Therefore, the targeted elimination or permanent silencing of this latently infected reservoir is a major focus of HIV-1 research. The most studied approach in the development of a cure is the activation of HIV-1 expression to expose latently infected cells for immune clearance while inducing HIV-1 cytotoxicity-the "kick and kill" approach. However, the complex and highly heterogeneous nature of the latent reservoir, combined with the failure of clinical trials to reduce the reservoir size casts doubt on the feasibility of this approach. This concern that total elimination of HIV-1 from the body may not be possible has led to increased emphasis on a "functional cure" where the virus remains but is unable to reactivate which presents the challenge of permanently silencing transcription of HIV-1 for prolonged drug-free remission-a "block and lock" approach. In this review, we discuss the interaction of HIV-1 and autophagy, and the exploitation of autophagy to kill selectively HIV-1 latently infected cells as part of a cure strategy. The cure strategy proposed has the advantage of significantly decreasing the size of the HIV-1 reservoir that can contribute to a functional cure and when optimised has the potential to eradicate completely HIV-1.

Metal accumulation varies with life history, size, and development of larval amphibians.

Amphibian larvae are commonly used as indicators of aquatic ecosystem health because they are susceptible to contaminants. However, there is limited information on how species characteristics and trophic position influence contaminant loads in larval amphibians. Importantly, there remains a need to understand whether grazers (frogs and toads [anurans]) and predators (salamanders) provide comparable information on contaminant accumulation or if they are each indicative of unique environmental processes and risks. To better understand the role of trophic position in contaminant accumulation, we analyzed composite tissues for 10 metals from larvae of multiple co-occurring anuran and salamander species from 20 wetlands across the United States. We examined how metal concentrations varied with body size (anurans and salamanders) and developmental stage (anurans) and how the digestive tract (gut) influenced observed metal concentrations. Across all wetlands, metal concentrations were greater in anurans than salamanders for all metals tested except mercury (Hg), selenium (Se), and zinc (Zn). Concentrations of individual metals in anurans decreased with increasing weight and developmental stage. In salamanders, metal concentrations were less correlated with weight, indicating diet played a role in contaminant accumulation. Based on batches of similarly sized whole-body larvae compared to larvae with their digestive tracts removed, our results indicated that tissue type strongly affected perceived concentrations, especially for anurans (gut represented an estimated 46-97% of all metals except Se and Zn). This suggests the reliability of results based on whole-body sampling could be biased by metal, larval size, and development. Overall, our data shows that metal concentrations differs between anurans and salamanders, which suggests that metal accumulation is unique to feeding behavior and potentially trophic position. To truly characterize exposure risk in wetlands, species of different life histories, sizes and developmental stages should be included in biomonitoring efforts.

SARS-CoV-2, SARS-CoV-1, and HIV-1 derived ssRNA sequences activate the NLRP3 inflammasome in human macrophages through a non-classical pathway.

Macrophages promote an early host response to infection by releasing pro-inflammatory cytokines such as interleukin-1ß (IL-1ß), TNF, and IL-6. The bioactivity of IL-1ß is classically dependent on NLRP3 inflammasome activation, which culminates in caspase-1 activation and pyroptosis. Recent studies suggest a role for NLRP3 inflammasome activation in lung inflammation and fibrosis in both COVID-19 and SARS, and there is evidence of NLRP3 involvement in HIV-1 disease. Here, we show that GU-rich single-stranded RNA (GU-rich RNA) derived from SARS-CoV-2, SARS-CoV-1, and HIV-1 trigger a TLR8-dependent pro-inflammatory cytokine response from human macrophages in the absence of pyroptosis, with GU-rich RNA from the SARS-CoV-2 spike protein triggering the greatest inflammatory response. Using genetic and pharmacological inhibition, we show that the induction of mature IL-1ß is through a non-classical pathway dependent on caspase-1, caspase-8, the NLRP3 inflammasome, potassium efflux, and autophagy while being independent of TRIF (TICAM1), vitamin D3, and pyroptosis.

Isoproterenol Causing Coronary Vasospasm and ST Elevations During Tilt Table Testing.

Syncope is a sudden but reversible brief loss of consciousness secondary to an acute reduction of cerebral perfusion. Reflex syncope denotes neurologically mediated syncope, which includes vasovagal, carotid sinus syndrome, and other situational syncope. The most frequent form of syncope is vasovagal, which is triggered by emotional stress or prolonged standing, and may be diagnosed with the tilt table test. A thorough investigation of syncope is necessary as serious cardiovascular disorders may also be a cause. A tilt table test is a widely used tool utilized by clinicians to diagnose vasovagal syncope and is sometimes augmented with isoproterenol, a ß-sympathomimetic that acts on the heart. This report seeks to explain a case of a 48-year-old previously healthy woman who experienced inferior wall ST elevations during tilt table test supplemented with isoproterenol. There is reason to believe that the results of this patient's tilt table test were due to vasovagal syncope in conjunction with right coronary artery vasospasm.

CD4+ T Cell-Mimicking Nanoparticles Broadly Neutralize HIV-1 and Suppress Viral Replication through Autophagy.

Therapeutic strategies that provide effective and broad-spectrum neutralization against HIV-1 infection are highly desirable. Here, we investigate the potential of nanoengineered CD4+ T cell membrane-coated nanoparticles (TNP) to neutralize a broad range of HIV-1 strains. TNP displayed outstanding neutralizing breadth and potency; they neutralized all 125 HIV-1-pseudotyped viruses tested, including global subtypes/recombinant forms, and transmitted/founder viruses, with a geometric mean 80% inhibitory concentration (IC80) of 819 µg ml-1 (range, 72 to 8,570 µg ml-1). TNP also selectively bound to and induced autophagy in HIV-1-infected CD4+ T cells and macrophages, while having no effect on uninfected cells. This TNP-mediated autophagy inhibited viral release and reduced cell-associated HIV-1 in a dose- and phospholipase D1-dependent manner. Genetic or pharmacological inhibition of autophagy ablated this effect. Thus, we can use TNP as therapeutic agents to neutralize cell-free HIV-1 and to target HIV-1 gp120-expressing cells to decrease the HIV-1 reservoir.IMPORTANCE HIV-1 is a major global health challenge. The development of an effective vaccine and/or a therapeutic cure is a top priority. The creation of vaccines that focus an antibody response toward a particular epitope of a protein has shown promise, but the genetic diversity of HIV-1 hinders this progress. Here we developed an approach using nanoengineered CD4+ T cell membrane-coated nanoparticles (TNP). Not only do TNP effectively neutralize all strains of HIV-1, but they also selectively bind to infected cells and decrease the release of HIV-1 particles through an autophagy-dependent mechanism with no drug-induced off-target or cytotoxic effects on bystander cells.

SMAC mimetics induce autophagy-dependent apoptosis of HIV-1-infected macrophages.

Human immunodeficiency type 1 (HIV)-infected macrophages (HIV-Mφ) are a reservoir for latent HIV infection and a barrier to HIV eradication. In contrast to CD4+ T cells, HIV-Mφ are resistant to the cytopathic effects of acute HIV infection and have increased expression of cell survival factors, including X-linked inhibitor of apoptosis (XIAP), baculoviral IAP repeat containing (BIRC) 2/cIAP1, beclin-1, BCL2, BCL-xl, triggering receptor expressed on myeloid cells 1, mitofusin (MFN) 1, and MFN2. DIABLO/SMAC mimetics are therapeutic agents that affect cancer cell survival and induce cell death. We found that DIABLO/SMAC mimetics (LCL-161, AT-406 (also known as SM-406 or Debio 1143), and birinapant) selectively kill HIV-Mφ without increasing bystander cell death. DIABLO/SMAC mimetic treatment of HIV-Mφ-induced XIAP and BIRC2 degradation, leading to the induction of autophagy and the formation of a death-inducing signaling complex on phagophore membranes that includes both pro-apoptotic or necroptotic (FADD, receptor-interacting protein kinase (RIPK) 1, RIPK3, caspase 8, and MLKL) and autophagy (ATG5, ATG7, and SQSTM1) proteins. Genetic or pharmacologic inhibition of early stages of autophagy, but not late stages of autophagy, ablated this interaction and inhibited apoptosis. Furthermore, DIABLO/SMAC mimetic-mediated apoptosis of HIV-Mφ is dependent upon tumor necrosis factor signaling. Our findings thus demonstrate that DIABLO/SMAC mimetics selectively induce autophagy-dependent apoptosis in HIV-Mφ.

TREM-1 Protects HIV-1-Infected Macrophages from Apoptosis through Maintenance of Mitochondrial Function.

Macrophages are a reservoir for latent human immunodeficiency type 1 (HIV) infection and a barrier to HIV eradication. In contrast to CD4+ T cells, macrophages are resistant to the cytopathic effects of acute HIV infection. Emerging data suggest a role for TREM1 (triggering receptor expressed on myeloid cells 1) in this resistance to HIV-mediated cytopathogenesis. Here, we show that upon HIV infection, macrophages increase the expression of BCL2, BCLXL, TREM1, mitofusin 1 (MFN1), and MFN2 and the translocation of BCL2L11 (BIM) to the mitochondria and decrease the expression of BCL2-associated agonist of cell death (BAD) and BAX while maintaining a 95% survival rate over 28 days. The HIV proteins Tat and gp120 and the GU-rich single-stranded RNA (ssRNA) (RNA40) from the HIV long terminal repeat region (and a natural Toll-like receptor 8 [TLR8] agonist) induced similar effects. TREM1 silencing in HIV-infected macrophages led to decreased expression of BCL2, BCLXL, MFN1, and MFN2 and increased expression of BAD and BAX. This correlated with a significant increase in apoptosis mediated by a disruption of the mitochondrial membrane potential (Δψm), leading to the release of cytochrome c and caspase 9 cleavage. Exposure of TREM1-silenced macrophages to Tat, gp120, or RNA40 similarly resulted in the disruption of Δψm, cytochrome c release, caspase 9 cleavage, and apoptosis. Thus, our findings identify a mechanism whereby HIV promotes macrophage survival through TREM1-dependent upregulation of BCL2 family proteins and mitofusins that inhibits BCL2L11-mediated disruption of Δψm and subsequent apoptosis. These findings indicate that TREM1 can be a useful target for elimination of the HIV reservoir in macrophages.IMPORTANCE The major challenge to human immunodeficiency virus (HIV) treatment is the development of strategies that lead to viral eradication. A roadblock to accomplishing this goal is the lack of an approach that would safely eliminate HIV from all resting/latent reservoirs, including macrophages. Macrophages are a key part of the innate immune system and are responsible for recognizing invading microbes and sending appropriate signals to other immune cells. Here, we found that HIV induces the upregulation of the protein TREM1 (triggering receptor expressed on myeloid cells 1), which signals an increase in the expression of antiapoptotic proteins, thus promoting survival of HIV-infected macrophages.