Toll-like receptors (TLRs) are a class of pattern recognition receptors that play a bridging role in innate immunity and adaptive immunity. The activated TLRs not only induce inflammatory responses, but also elicit the development of antigen specific immunity. TLR7, a member of TLR family, is an intracellular receptor expressed on the membrane of endosomes. TLR7 can be triggered not only by ssRNA during viral infections, but also by immune modifiers that share a similar structure to nucleosides. Its powerful immune stimulatory action can be potentially used in the anti-tumor therapy. This article reviewed the anti-tumor activity and mechanism of TLR7 agonists that are frequently applied in preclinical and clinical investigations, and mainly focused on small synthetic molecules, including imiquimod, resiquimod, gardiquimod, and 852A, etc.
Peripheral and integral membrane proteins can be located in several different subcellular compartments, and it is often necessary to determine the location of such proteins or to track their movement in living cells. Image-based colocalization of labeled membrane proteins and compartment markers is frequently used for this purpose, but this method is limited in terms of throughput and resolution. Here we show that bioluminescence resonance energy transfer (BRET) between membrane proteins of interest and compartment-targeted BRET partners can report subcellular location and movement of membrane proteins in live cells. The sensitivity of the method is sufficent to localize a few hundred protein copies per cell. The spatial resolution can be sufficient to determine membrane topology, and the temporal resolution is sufficient to track changes that occur in less than one second. BRET requires little user intervention, and is thus amenable to large-scale experimental designs with standard instruments.
The molecular mechanisms underlying the transport from the Golgi to the cell surface of G protein‐coupled receptors (GPCRs) remain poorly elucidated. Here we determined the role of Rab26, a Ras‐like small GTPase involved in vesicle‐mediated secretion, in the cell‐surface export of α2‐adrenergic receptors (α2‐ARs). We found that transient expression of Rab26 mutants and siRNA‐mediated depletion of Rab26 significantly attenuated the cell‐surface numbers of α2A‐AR and α2B‐AR as well as ERK1/2 activation by α2B‐AR. Furthermore, the receptors were extensively arrested in the Golgi by Rab26 mutants and siRNA. Moreover, Rab26 directly and activation‐dependently interacted with α2BAR, specifically the third intracellular loop. These data demonstrate that the small GTPase Rab26 regulates the Golgi‐to‐cell surface traffic of α2‐ARs, likely through a physical interaction. These data also provide the first evidence implicating an important function of Rab26 in coordinating plasma membrane protein transport (R01GM076167, R01GM096762 and R01GM078319).
The transport protein particle (TRAPP) was initially identified as a vesicle tethering factor in yeast and as a guanine nucleotide exchange factor (GEF) for Ypt1/Rab1. In mammals, structures and functions of various TRAPP complexes are beginning to be understood. We found that mammalian TRAPPII was a GEF for both Rab18 and Rab1. Inactivation of TRAPPII-specific subunits by various methods including siRNA depletion and CRISPR-Cas9-mediated deletion reduced lipolysis and resulted in aberrantly large lipid droplets. Recruitment of Rab18 onto lipid droplet (LD) surface was defective in TRAPPII-deleted cells, but the localization of Rab1 on Golgi was not affected. COPI regulates LD homeostasis. We found that the previously documented interaction between TRAPPII and COPI was also required for the recruitment of Rab18 to the LD. We hypothesize that the interaction between COPI and TRAPPII helps bring TRAPPII onto LD surface, and TRAPPII, in turn, activates Rab18 and recruits it on the LD surface to facilitate its functions in LD homeostasis.
Background: ERK1/2 activation by G protein-coupled receptors can be mediated through multiple pathways. Results: ␣ 2B -Adrenergic receptor interacts with ADP-ribosylation factor 1 (ARF1) to modulate ERK1/2 activation. Conclusion: ERK1/2 activation by ␣ 2B -adrenergic receptor is mediated through a novel pathway involving ARF1. Significance: The small GTPase ARF1 may function as an important regulator in receptor-mediated ERK1/2 activation.
This article reviews mushrooms with anti-breast cancer activity. The mushrooms covered which are better known include the following: button mushroom Agaricus bisporus, Brazilian mushroom Agaricus blazei, Amauroderma rugosum, stout camphor fungus Antrodia camphorata, Jew's ear (black) fungus or black wood ear fungus Auricularia auricula-judae, reishi mushroom or Lingzhi Ganoderma lucidum, Ganoderma sinense, maitake mushroom or sheep's head mushroom Grifola frondosa, lion's mane mushroom or monkey head mushroom Hericium erinaceum, brown beech mushroom Hypsizigus marmoreus, sulfur polypore mushroom Laetiporus sulphureus, Lentinula edodes (shiitake mushroom), Phellinus linteus (Japanese "meshimakobu," Chinese "song gen," Korean "sanghwang," American "black hoof mushroom"), abalone mushroom Pleurotus abalonus, king oyster mushroom Pleurotus eryngii, oyster mushroom Pleurotus ostreatus, tuckahoe or Fu Ling Poria cocos, and split gill mushroom Schizophyllum commune. Antineoplastic effectiveness in human clinical trials and mechanism of anticancer action have been reported for Antrodia camphorata, Cordyceps sinensis, Coriolus versicolor, Ganoderma lucidum, Grifola frondosa, and Lentinula edodes. Agaricus placomycesThe 68-kDa laccase from Agaricus placomyces belongs to a family of copper-containing oxidases involved in lignin degradation and possessing anticancer activity (Mizerska-Dudka et al. 2015). It potently expressed antiproliferative activity toward MCF-7 human breast cancer cells with an IC50 of 1.8 μM (Sun et al. 2012). Agaricus sylvaticusIn a randomized, double-blind, and placebo-controlled clinical trial, 46 patients with stage II and III breast cancer who received chemotherapy demonstrated an improved appetite and fewer gastrointestinal complaints, nausea, and vomiting following dietary supplementation with A. sylvaticus (2.1 g/day) (Valadares et al. 2013). Amauroderma rude Amauroderma rude had higher tumoricidal activity than Ganoderma lucidum. A. rude lowered cell survival and triggered apoptosis in MDA-MB-231 breast cancer cells. Tumor growth in athymic nude mice bearing MDA-MB-231-xenografts was retarded and tumor cell death was promoted by treatment with A. rude. Expression of the c-Myc oncogene was downregulated . A. rude exhibited a higher content of ergosterol (ergosta-5,7,22-trien-3β-ol) in comparison with 12 other mushrooms examined. Under the influence of ergosterol, the viability of MDA-MB-231 breast cancer cells was reduced, whereas expression of the tumor suppressor Foxo3 and downstream signaling molecules including BimL, BimS, Fas, and FasL was enhanced resulting in apoptosis of the breast cancer cells (Li et al. 2015).Amauroderma rugosum (also known as black Lingzhi, a member of five-color Lingzhi) Organic molecules from A. rugosum inhibited oxidant activity, tumor necrosis factor-α and nitric oxide formation, and MCF7 cell proliferation (Zhang et al. 2017a). A single treatment with pulverized A. rugosum (2000 mg/kg) by mouth did not impair growth, blood parameters, histological structures of t...
SUMMARYG-protein-coupled receptors (GPCRs) constitute the largest superfamily of cell-surface signaling proteins. However, mechanisms underlying their surface targeting and sorting are poorly understood. Here, we screen the Rab family of small GTPases in the surface transport of multiple GPCRs. We find that manipulation of Rab43 function significantly alters the surface presentation and signaling of all GPCRs studied without affecting non-GPCR membrane proteins. Rab43 specifically regulates the transport of nascent GPCRs from the endoplasmic reticulum (ER) to the Golgi. More interestingly, Rab43 directly interacts with GPCRs in an activation-dependent fashion. The Rab43-binding domain identified in the receptors effectively converts non-GPCR membrane protein transport into a Rab43-dependent pathway. These data reveal a crucial role for Rab43 in anterograde ER-Golgi transport of nascent GPCRs, as well as the ER sorting of GPCR members by virtue of its ability to interact directly.
Bioluminescence resonance energy transfer (BRET) is often used to study association of membrane proteins, and in particular oligomerization of G protein-coupled receptors (GPCRs). Oligomerization of class A GPCRs is controversial, in part because the methods used to study this question are not completely understood. Here we reconsider oligomerization of the class A β2 adrenergic receptor (β2AR), and reevaluate BRET titration as a method to study membrane protein association. Using inducible expression of the energy acceptor at multiple levels of donor expression we find that BRET between β2AR protomers is directly proportional to the density of the acceptor up to ~3,000 acceptors μm−2, and does not depend on the density of the donor or on the acceptor:donor (A:D) stoichiometry. In contrast, BRET between tightly-associating control proteins does not depend on the density of the acceptor, but does depend on the density of the donor and on the A:D ratio. We also find that the standard frameworks used to interpret BRET titration experiments rely on simplifying assumptions that are frequently invalid. These results suggest that β2ARs do not oligomerize in cells, and demonstrate a reliable method of assessing membrane protein association with BRET.
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