Selective and regulated trapping of nicotinic receptor weak base ligands and relevance to smoking cessation

© The Author(s), 2017. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in eLife 6 (2017): e25651, doi:10.7554/eLife.25651.To better understand smoking cessation, we examined the actions of varenicline (Chantix) during long-term nicotine exposure. Varenicline reduced nicotine upregulation of α4β2-type nicotinic receptors (α4β2Rs) in live cells and neurons, but not for membrane preparations. Effects on upregulation depended on intracellular pH homeostasis and were not observed if acidic pH in intracellular compartments was neutralized. Varenicline was trapped as a weak base in acidic compartments and slowly released, blocking 125I-epibatidine binding and desensitizing α4β2Rs. Epibatidine itself was trapped; 125I-epibatidine slow release from acidic vesicles was directly measured and required the presence of α4β2Rs. Nicotine exposure increased epibatidine trapping by increasing the numbers of acidic vesicles containing α4β2Rs. We conclude that varenicline as a smoking cessation agent differs from nicotine through trapping in α4β2R-containing acidic vesicles that is selective and nicotine-regulated. Our results provide a new paradigm for how smoking cessation occurs and suggest how more effective smoking cessation reagents can be designed.This work was supported by National Institutes of Health RO1DA 035430 and a Pilot Project from the University of Chicago Can- cer Center

Combinations of Griffithsin with Other Carbohydrate-Binding Agents Demonstrate Superior Activity Against HIV Type 1, HIV Type 2, and Selected Carbohydrate-Binding Agent-Resistant HIV Type 1 Strains

Abstract Carbohydrate-binding agents (CBAs) are potential HIV microbicidal agents with a high genetic barrier to resistance. We wanted to evaluate whether two mannose-specific CBAs, recognizing multiple and often distinct glycan structures on the HIV envelope gp120, can interact synergistically against HIV-1, HIV-2, and HIV-1 strains that were selected for resistance against particular CBAs [i.e., 2G12 mAb and microvirin (MVN)]. Paired CBA/CBA combinations mainly showed synergistic activity against both wild-type HIV-1 and HIV-2 but also 2G12 mAb- and MVN-resistant HIV-1 strains as based on the median effect principle with combination indices (CIs) ranging between 0.29 and 0.97. Upon combination, an increase in antiviral potency of griffithsin (GRFT) up to ?12-fold (against HIV-1), ?8-fold (against HIV-2), and ?6-fold (against CBA-resistant HIV-1) was observed. In contrast, HHA/GNA combinations showed additive activity against wild-type HIV-1 and HIV-2 strains, but remarkable synergy with HHA and GNA was observed against 2G12 mAb- and MVN-resistant HIV-1 strains (CI, 0.64 and 0.49, respectively). Overall, combinations of GRFT and other CBAs showed synergistic activity against HIV-1, HIV-2, and even against certain CBA-resistant HIV-1 strains. The CBAs tested appear to have distinct binding patterns on the gp120 envelope and therefore do not necessarily compete with each other's glycan binding sites on gp120. As a result, there might be no steric hindrance between two different CBAs in their competition for glycan binding (except for the HHA/GNA combination). These data are encouraging for the use of paired CBA combinations in topical microbicide applications (e.g., creams, gels, or intravaginal rings) to prevent HIV transmission.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/98459/1/aid%2E2012%2E0026.pd

Phosphorylation of the HCN channel auxiliary subunit TRIP8b is altered in an animal model of temporal lobe epilepsy and modulates channel function

Temporal lobe epilepsy (TLE) is a prevalent neurological disorder with many patients experiencing poor seizure control with existing anti-epileptic drugs. Thus, novel insights into the mechanisms of epileptogenesis and identification of new drug targets can be transformative. Changes in ion channel function have been shown to play a role in generating the aberrant neuronal activity observed in TLE. Previous work demonstrates that hyperpolarization-activated cyclic nucleotide-gated (HCN) channels regulate neuronal excitability and are mislocalized within CA1 pyramidal cells in a rodent model of TLE. The subcellular distribution of HCN channels is regulated by an auxiliary subunit, tetratricopeptide repeat-containing Rab8b-interacting protein (TRIP8b), and disruption of this interaction correlates with channel mislocalization. However, the molecular mechanisms responsible for HCN channel dysregulation in TLE are unclear. Here we investigated whether changes in TRIP8b phosphorylation are sufficient to alter HCN channel function. We identified a phosphorylation site at residue Ser237 of TRIP8b that enhances binding to HCN channels and influences channel gating by altering the affinity of TRIP8b for the HCN cytoplasmic domain. Using a phosphospecific antibody, we demonstrate that TRIP8b phosphorylated at Ser237 is enriched in CA1 distal dendrites and that phosphorylation is reduced in the kainic acid model of TLE. Overall, our findings indicate that the TRIP8b-HCN interaction can be modulated by changes in phosphorylation and suggest that loss of TRIP8b phosphorylation may affect HCN channel properties during epileptogenesis. These results highlight the potential of drugs targeting posttranslational modifications to restore TRIP8b phosphorylation to reduce excitability in TLE

Proteomics Reveals Novel Drosophila Seminal Fluid Proteins Transferred at Mating

Across diverse taxa, seminal fluid proteins (Sfps) transferred at mating affect the reproductive success of both sexes. Such reproductive proteins often evolve under positive selection between species; because of this rapid divergence, Sfps are hypothesized to play a role in speciation by contributing to reproductive isolation between populations. In Drosophila, individual Sfps have been characterized and are known to alter male sperm competitive ability and female post-mating behavior, but a proteomic-scale view of the transferred Sfps has been missing. Here we describe a novel proteomic method that uses whole-organism isotopic labeling to detect transferred Sfps in mated female D. melanogaster. We identified 63 proteins, which were previously unknown to function in reproduction, and confirmed the transfer of dozens of predicted Sfps. Relative quantification of protein abundance revealed that several of these novel Sfps are abundant in seminal fluid. Positive selection and tandem gene duplication are the prevailing forces of Sfp evolution, and comparative proteomics with additional species revealed lineage-specific changes in seminal fluid content. We also report a proteomic-based gene discovery method that uncovered 19 previously unannotated genes in D. melanogaster. Our results demonstrate an experimental method to identify transferred proteins in any system that is amenable to isotopic labeling, and they underscore the power of combining proteomic and evolutionary analyses to shed light on the complex process of Drosophila reproduction

BiblioBouts: A Scalable Online Social Game for the Development of Academic Research Skills

Researchers at the School of Information of the University of Michigan are designing, developing, and evaluating BiblioBouts, an online game that helps students learn academic research skills. Players practice using online library research tools while they work on an in-class assignment and produce a high-quality bibliography, at the same time as they are competing against each other to win the game! While librarians are experts at helping students who want to learn about academic research, most students are reluctant participants because they want just-in-time personal assistance that is tailored to their unique information needs, and faculty are reluctant to cede class time. The BiblioBouts project enlists games to teach undergraduate students information literacy skills and concepts in the classroom. Social gaming reinforces principles of good learning, including getting results by trial and error, self-discovery, following hunches and reinforcement through repetition. BiblioBouts also incorporates collaborative problem solving and participation in a community of learning. The project aims to explore how games can be utilized to achieve information literacy goals and to yield open-source game software that libraries could use immediately to enhance their information literacy programs. The LOEX presentation will incorporate a live interactive demo of the game, as well as videos demonstrating gameplay. We will discuss challenges in situating the game into the classroom and integrating it into existing course syllabi. The presentation will describe how we have adapted the game in response to feedback from students and instructors during the pilot process

The Induction of Autoimmune Arthritis and Sex differences in Mice Impact the Lung Inflammatory Response to Repetitive Inhalant Organic Dust Extract Exposures

Asthma, chronic bronchitis and COPD are common adverse respiratory health effects among persons exposed to agriculture organic dust work environments. Occupational inhalant exposures have been increasingly associated with the risk of rheumatoid arthritis (RA) disease development, particularly among males. Agriculture workers have increased risk of RA and generalized bone disease. Chronic lung disease is associated with production of characteristic autoantibodies associated with RA (e.g.anti-citrullinated antibodies), even in absence of RA disease. The mechanistic link between pulmonary inflammation and arthritis (and vice versa) remains poorly understood. Animal models are lacking.https://digitalcommons.unmc.edu/emet_posters/1004/thumbnail.jp

Activity-dependent Golgi satellite formation in dendrites reshapes the neuronal surface glycoproteome

© The Author(s), 2021. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Govind, A. P., Jeyifous, O., Russell, T. A., Yi, Z., Weigel, A., Ramaprasad, A., Newell, L., Ramos, W., Valbuena, F. M., Casler, J. C., Yan, J.-Z., Glick, B. S., Swanson, G. T., Lippincott-Schwartz, J., & Green, W. N. Activity-dependent Golgi satellite formation in dendrites reshapes the neuronal surface glycoproteome. Elife, 10, (2021): e68910, https://doi.org/10.7554/eLife.68910.Activity-driven changes in the neuronal surface glycoproteome are known to occur with synapse formation, plasticity, and related diseases, but their mechanistic basis and significance are unclear. Here, we observed that N-glycans on surface glycoproteins of dendrites shift from immature to mature forms containing sialic acid in response to increased neuronal activation. In exploring the basis of these N-glycosylation alterations, we discovered that they result from the growth and proliferation of Golgi satellites scattered throughout the dendrite. Golgi satellites that formed during neuronal excitation were in close association with endoplasmic reticulum (ER) exit sites and early endosomes and contained glycosylation machinery without the Golgi structural protein, GM130. They functioned as distal glycosylation stations in dendrites, terminally modifying sugars either on newly synthesized glycoproteins passing through the secretory pathway or on surface glycoproteins taken up from the endocytic pathway. These activities led to major changes in the dendritic surface of excited neurons, impacting binding and uptake of lectins, as well as causing functional changes in neurotransmitter receptors such as nicotinic acetylcholine receptors. Neural activity thus boosts the activity of the dendrite’s satellite micro-secretory system by redistributing Golgi enzymes involved in glycan modifications into peripheral Golgi satellites. This remodeling of the neuronal surface has potential significance for synaptic plasticity, addiction, and disease.This work was financially supported by NIH RO1 DA035430, DA044760, and DA043361 (WNG) R01 GM104010 (BSG), T32 GM007183 (FV), and Peter F McManus Foundation (WNG)

MicroRNA-30c targets cytoskeleton genes involved in breast cancer cell invasion

Metastasis remains a significant challenge in treating cancer. A better understanding of the molecular mechanisms underlying metastasis is needed to develop more effective treatments. Here we show that human breast tumor biomarker miR-30c regulates invasion by targeting the cytoskeleton network genes encoding Twinfilin 1 (TWF1) and Vimentin (VIM). Both VIM and TWF1 have been shown to regulate epithelial-to-mesenchymal transition (EMT). Similar to TWF1, VIM also regulates F-actin formation, a key component of cellular transition to a more invasive mesenchymal phenotype. To further characterize the role of the TWF1 pathway in breast cancer, we found that IL-11 is an important target of TWF1 that regulates breast cancer cell invasion and STAT3 phosphorylation. The miR-30c-VIM/TWF1 signaling cascade is also associated with clinical outcome in breast cancer patients