Chongguang Chen
Temple University
Network
Latest external collaboration on country level. Dive into details by clicking on the dots.
Publication
Featured researches published by Chongguang Chen.
Journal of Pharmacology and Experimental Therapeutics | 2006
Wei Xu; Su-In Yoon; Peng Huang; Yulin Wang; Chongguang Chen; Parkson Lee-Gau Chong; Lee-Yuan Liu-Chen
Lipid rafts are microdomains of plasma membranes enriched in cholesterol and sphingolipids in the outer layer. We determined whether κ opioid receptors (KOR) in human placenta and FLAG (DYKDDDDK)-tagged human KOR (FLAG-hKOR) expressed in Chinese hamster ovary (CHO) cells are localized in lipid rafts and whether changes in cholesterol contents affect hKOR properties and signaling. Lipid rafts were prepared from placenta membranes and CHO cells expressing FLAG-hKOR using the Na2CO3 method and fractionation through a sucrose density gradient. The majority of the KOR in the placenta and FLAG-hKOR in CHO cells, determined by [3H]diprenorphine binding and/or immunoblotting with an anti-FLAG antibody, was present in low-density fractions, coinciding with high levels of caveolin-1 and cholesterol, markers of lipid rafts, which indicated that the KOR is localized in lipid rafts. Pretreatment with 2% methyl β-cyclodextrin (MCD) reduced cholesterol content by ∼48% and changed the cells from spindle-shaped to spherical. MCD treatment disrupted lipid rafts, shifted caveolin-1 and FLAG-hKOR to higher density fractions, increased the affinity of (–)-(trans)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)cyclohexyl]benzeneacetamide (U50,488H) for the hKOR, and greatly increased U50,488H-induced [35S]guanosine 5′-O-(3-thio)triphosphate binding and p42/44 mitogen-activated protein kinase phosphorylation. Cholesterol replenishment reversed all the MCD effects. Caveolin-1 immunoprecipitated with Gαi proteins and MCD treatment reduced caveolin-1 associated with Gαi proteins, which may contribute to the enhanced agonist-induced G protein activation. Caveolin-1 also immunoprecipitated with FLAG-hKOR, but MCD treatment had no effect on the association. Thus, the KOR is located in lipid rafts and its localization in the microdomains greatly affects coupling to G proteins.
Journal of Biological Chemistry | 2006
Chongguang Chen; Jian-Guo Li; Yong Chen; Peng Huang; Yulin Wang; Lee-Yuan Liu-Chen
We identified a truncated form (38–117) of GEC1 that interacts with the C-tail of the human κ opioid receptor (hKOR) by yeast two-hybrid screening. GEC1-(38–117) did not interact with the C-tail of the μ or δ opioid receptors. GEC1, a 117-amino acid protein (Pellerin, I., Vuillermoz, C., Jouvenot, M., Ordener, C., Royez, M., and Adessi, G. L. (1993) Mol. Cell Endocrinol. 90, R17–R21), is highly homologous to GABARAP, GATE-16, and Apg8/aut7, all members of the microtubule associated protein (MAP) family. In pull-down assays, GST-GEC1 interacted directly with the hKOR C-tail, full-length hKOR, and tubulin. When expressed in Chinese hamster ovary (CHO) cells, GEC1 co-immunoprecipitated with FLAG-hKOR. Expression of GEC1 greatly increased total and cell-surface KOR but not μ or δ opioid receptors. GEC1 expression slightly reduced U50,488H-promoted down-regulation, without affecting ligand binding affinity, receptor-G protein coupling, or U50,488H-induced desensitization and internalization. HA-GEC1 expressed in CHO cells was localized in the Golgi apparatus and endoplasmic reticulum (ER). When cells were pulsed with [35S]Met/Cys, GEC1 expression enhanced the level of the mature form (55-kDa band) of FLAG-hKOR at 4, 8, and 22 h after pulse without affecting the precursors (39- and 45-kDa bands), indicating that GEC1 facilitates trafficking of FLAG-hKOR from the ER/Golgi to plasma membranes. GEC1 interacted with N-ethylmaleimide-sensitive factor (NSF) in pull-down assays and co-immunoprecipitated with NSF in rat brain extracts. The interaction with NSF may contribute to GEC1 effects. This is the first report on biological functions of GEC1 and the first demonstration that a GPCR interacts with a protein of the MAP family. The interaction is important for trafficking of the receptor in the biosynthesis pathway.
Journal of Leukocyte Biology | 2003
Imre Szabo; Michele A. Wetzel; Ning Zhang; Amber D. Steele; David E. Kaminsky; Chongguang Chen; Lee Yuan Liu-Chen; Filip Bednar; Earl E. Henderson; O. M. Zack Howard; Joost J. Oppenheim; Thomas J. Rogers
The opiates are well‐established immunomodulatory factors, and recent evidence suggests that μ‐ and δ‐opioid receptor ligands alter chemokine‐driven chemotactic responses through the process of heterologous desensitization. In the present report, we sought to examine the capacity of μ‐ and δ‐opioids to modulate the function of chemokine receptors CCR5 and CXCR4, the two major human immunodeficiency virus (HIV) coreceptors. We found that the chemotactic responses to the CCR1/5 ligand CCL5/regulated on activation, normal T expressed and secreted, but not the CXCR4 ligand stromal cell‐derived factor‐1α/CXCL12 were inhibited following opioid pretreatment. Studies were performed with primary monocytes and Chinese hamster ovary cells transfected with CCR5 and the μ‐opioid receptor to determine whether cross‐desensitization of CCR5 was a result of receptor internalization. Using radiolabeled‐binding analysis, flow cytometry, and confocal microscopy, we found that the heterologous desensitization of CCR5 was not associated with a significant degree of receptor internalization. Despite this, we found that the cross‐desensitization of CCR5 by opioids was associated with a decrease in susceptibility to R5 but not X4 strains of HIV‐1. Our findings are consistent with the notion that impairment of the normal signaling activity of CCR5 inhibits HIV‐1 coreceptor function. These results have significant implications for our understanding of the effect of opioids on the regulation of leukocyte trafficking in inflammatory disease states and the process of coreceptor‐dependent HIV‐1 infection. The interference with HIV‐1 uptake by heterologous desensitization of CCR5 suggests that HIV‐1 interaction with this receptor is not passive but involves a signal transduction process.
Life Sciences | 1999
Jian-Guo Li; Chongguang Chen; Jinling Yin; Kenner C. Rice; Ying Zhang; Dorota Matecka; J. Kim de Riel; Renee L. Desjarlais; Lee-Yuan Liu-Chen
We tested the hypotheses that the carboxylate side chain of Asp147 of the mu opioid receptor interacts with the protonated nitrogen of naltrexone and morphine and that this interaction is important for pharmacological properties of the two compounds. Mutation of Asp147 to Ala or Asn substantially reduced the affinity of naltrexone and the affinity, potency and efficacy of morphine, while the Glu mutant had similar properties as the wildtype, indicating the significant role of the carboxylate group of Asp147 in receptor binding and activation. This role could be due to its direct interaction with ligands or involvement in interhelical interactions. The unprotonated analogs of naltrexone and morphine, cyclopropylcarbonyl noroxymorphone (CPCNOM) and N-formylnormorphine (NFNM), respectively, were used to discriminate between these mechanisms. CPCNOM was much less potent as an antagonist and had substantially lower affinity for the mu receptor than naltrexone. Similarly, NFNM was unable to activate the mu receptor and had much lower affinity than morphine. These results indicate the importance of the protonated nitrogen. Notably, the D147A and D147N mutations did not appreciably affect the binding affinities of CPCNOM and NFNM. In addition, the D147E mutant had similar affinities for CPCNOM and NFNM as the D147A and D147N mu receptors. Thus, the carboxylate group of Asp147 is not important for binding of the two unprotonated compounds. These results indicate that the carboxylate group of Asp147 of the mu receptor interacts directly with the protonated nitrogen of naltrexone and morphine and this interaction is important for binding and receptor activation.
Journal of Pharmacology and Experimental Therapeutics | 2006
Yong Chen; Chongguang Chen; Yulin Wang; Lee-Yuan Liu-Chen
Two peptide agonists, eight nonpeptide agonists, and five nonpeptide antagonists were evaluated for their capacity to regulate FLAG (DYKDDDDK)-tagged human κ opioid receptors (hKORs) stably expressed in Chinese hamster ovary cells after incubation for 4 h with a ligand at a concentration ∼1000-fold of its EC50 (agonist) or Ki (antagonist) value. Dynorphins A and B decreased the fully glycosylated mature form (55-kDa) of FLAG-hKOR by 70%, whereas nonpeptide full agonists [2-(3,4-dichlorophenyl)-N-methyl-N-[(2R)-2-pyrrolidin-1-ylcyclohexyl-]acetamide (U50,488H), 17-cyclopropylmethyl-3,14-dihydroxy-4,5-epoxy-6-[N-methyl-trans-3-(3-furyl) acrylamido] morphinan hydrochloride (TRK-820), ethylketocyclazocine, bremazocine, asimadoline, and (RS)-[3-[1-[[(3,4-dichlorophenyl)acetyl]-methylamino]-2-(1-pyrrolidinyl)ethyl]phenoxy] acetic acid hydrochloride (ICI 204,448) caused 10–30% decreases. In contrast, pentazocine (partial agonist) and etorphine (full agonist) up-regulated by ∼15 and 25%, respectively. The antagonists naloxone and norbinaltorphimine also significantly increased the 55-kDa receptor, whereas selective μ, δ, and D1 receptor antagonists had no effect. Naloxone up-regulated the receptor concentration- and time-dependently and enhanced the receptor maturation extent, without affecting its turnover. Treatment with brefeldin A (BFA), which disrupts Golgi, resulted in generation of a 51-kDa form that resided intracellularly. Naloxone up-regulated the new species, indicating that its action site is in the endoplasmic reticulum as a pharmacological chaperone. After treatment with BFA, all nonpeptide agonists up-regulated the 51-kDa form, whereas dynorphins A and B did not, indicating that nonpeptide agonists act as pharmacological chaperones, but peptide agonists do not. BFA treatment enhanced down-regulation of the cell surface receptor induced by nonpeptide agonists, but not that by peptide agonists, and unmasked etorphine- and pentazocine-mediated receptor down-regulation. These results demonstrate that ligands have dual effects on receptor levels: enhancement by chaperone-like effects and agonist-promoted down-regulation, and the net effect reflects the algebraic sum of the two.
Brain Research | 2007
Peng Huang; Wei Xu; Su-In Yoon; Chongguang Chen; Parkson Lee-Gau Chong; Ellen M. Unterwald; Lee-Yuan Liu-Chen
Lipid rafts are small cholesterol- and glycosphingolipid-enriched membrane subdomains. Here we compared the mu opioid receptor (MOR)-lipid rafts relationship in the rat brain, where neurons have non-caveolae rafts, and in CHO cells stably transfected with HA-rat MOR (CHO-HA-rMOR), which are enriched in caveolae. Membranes of rat caudate putamen (CPu) and thalamus or CHO-HA-rMOR cells were homogenized, sonicated in a detergent-free 0.5 M Na(2)CO(3) buffer and fractionated through sucrose density gradients. Western blot and [(3)H]diprenorphine binding showed that approximately 70% of MOR in CHO-HA-rMOR was present in low-density (5-20% sucrose) fractions enriched in cholesterol and/or ganglioside M1 (GM1) (lipid rafts) in plasma membranes, whereas about 70% and 45% of MOR in CPu and thalamus, respectively, were associated with lipid rafts. Incubation with a saturating concentration of etorphine or morphine at 37 degrees C for 30 min failed to change the MOR location in rafts in CHO-HA-rMOR, indicating that the internalized MOR does not move out of rafts, in contrast to the delta opioid receptor. In vivo, rafts association of MOR in CPu and thalamus was not affected significantly in rats implanted with two 75-mg morphine pellets for 72 h. In addition, cholesterol reduction by methyl-beta-cyclodextrin (MCD) disrupted rafts and shifted MOR to higher density fractions in both CHO-HA-rMOR and CPu membranes. However, MCD treatment had opposite impacts on MOR signaling in the two tissues: it attenuated MOR-mediated [(35)S]GTPgammaS binding in CPu but enhanced it in CHO-HA-rMOR.
FEBS Letters | 1998
Chongguang Chen; Vafa Shahabi; Wei Xu; Lee-Yuan Liu-Chen
We examined whether the μ opioid receptor was palmitoylated and attempted to determine sites of palmitoylation. Following metabolic labeling with [3H]palmitic acid and immunoaffinity purification of the μ opioid receptor, SDS‐PAGE and fluorography revealed a broad labeled band with M r of ∼80 kDa in CHO cells stably expressing the rat μ receptor, but not in CHO cells transfected with the vector alone, indicating that the μ receptor is palmitoylated. Activation of the receptor with morphine did not affect the extent of palmitoylation. Hydroxylamine or dithiothreitol treatment removed most of the radioactivity, demonstrating that [3H]palmitic acid is incorporated into Cys residue(s) via thioester bond(s). Surprisingly, mutations of the only two Cys residues in the C‐terminal domain did not reduce [3H]palmitic acid incorporation significantly. Thus, unlike many G‐protein coupled receptors, the palmitoylation site(s) of the rat μ opioid receptor do(es) not reside in the C‐terminal domain.
Journal of Biological Chemistry | 1996
Jinmin Zhu; Jinling Yin; Ping Yee Law; Patricia A. Claude; Kenner C. Rice; Christopher J. Evans; Chongguang Chen; Lei Yu; Lee Yuan Liu-Chen
Binding of cis-(+)-3-methylfentanyl isothiocyanate (SUPERFIT) to cloned opioid receptors stably expressed in Chinese hamster ovary cells was characterized. SUPERFIT inhibited [3H]diprenorphine binding with much higher affinity for the than the μ or κ receptor. Pretreatment with SUPERFIT followed by extensive washing reduced binding with an IC value of 7.1 nM, yet it did not affect μ and κ binding up to 0.1 μM. The reduction in binding by SUPERFIT pretreatment was due to a decrease in B with no change in K. These results indicate that SUPERFIT is a highly selective irreversible ligand. We then determined the region in the receptor that confered binding selectivity for SUPERFIT by examining its binding to six μ/ chimeric receptors. SUPERFIT bound to , μ/1 (amino acids μ1-94/76-372), /μ3 (1-134/μ154-398), and /μ4 (1-187/μ207-398) receptors with high affinity but to μ, /μ1 (1-75/μ95-398), μ/3 (μ1-153/135-372), and μ/4 (μ1-206/188-372) receptors with low affinity. Pretreatment with SUPERFIT potently inhibited [3H]diprenorphine binding to , μ/1, /μ3, and /μ4 but affected binding to μ, /μ1, μ/3, and μ/4 only at much higher concentrations. Thus, the segment from the beginning of the first intracellular loop to the middle of the third transmembrane helix of the receptor is important for selective binding of SUPERFIT.
FEBS Letters | 1996
Jinmin Zhu; Ji Chun Xue; Ping Yee Law; Patricia A. Claude; Lai Yi Luo; Jinling Yin; Chongguang Chen; Lee Yuan Liu-Chen
We determined the binding domains of sufentanil and lofentanil in the μ opioid receptor by comparing their binding affinities to seven μ/δ and six μ/κ chimeric receptors with those to μ, δ and κ opioid receptors. TMHs 6 and 7 and the e3 loop of the μ opioid receptor were important for selective binding of sufentanil and lofentanil to the μ over the κ receptor. TMHs 1–3 and the e1 loop of the μ opioid receptor conferred binding selectivity for sufentanil over the δ receptor. Thus, the region that conferred binding selectivity for sufentanil differs, depending on chimeras used. In addition, the interaction TMHs 1–3 and TMHs 6–7 was crucial for the high affinity binding of these two ligands. These two regions are likely to contain sites of interaction with the ligands or to confer conformations specific to the μ receptor.
Journal of Biological Chemistry | 2011
Chongguang Chen; Yulin Wang; Peng Huang; Lee-Yuan Liu-Chen
We demonstrated previously that GEC1, a member of the microtubule-associated protein (MAP) family, bound to the human κ opioid receptor (hKOPR) and promoted hKOPR cell surface expression by facilitating its trafficking along the secretory pathway. GABAA receptor-associated protein (GABARAP), a GEC1 analog, also enhanced KOPR expression, but to a lesser extent. The MAP family proteins undergo cleavage of their C-terminal residue(s), and the exposed conserved glycine forms conjugates with phosphatidylethanolamine, which associate with membranes. Here, we examined whether such modifications were required for GEC1 and GABARAP to enhance hKOPR expression. When transiently transfected into CHO or Neuro2A cells, GEC1 and GABARAP were cleaved at the C termini. G116A mutation alone or combined with deletion of Lys117 in GEC1 (GEC1-A) or Leu117 in GABARAP (GABARAP-A) blocked their C-terminal cleavage, indicating that the conserved Gly116 is necessary for C-terminal modification. The two GEC1 mutants enhanced hKOPR expression to similar extents as the wild-type GEC1; however, the two GABARAP mutants did not. Immunofluorescence studies showed that HA-GEC1, HA-GEC1-A, and HA-GABARAP were distributed in a punctate manner and co-localized with KOPR-EGFP in the Golgi apparatus, whereas HA-GABARAP-A did not. Pulldown assay of GST-KOPR-C-tail with HA-GEC1 or HA-GABARAP revealed that GEC1 had stronger association with KOPR-C-tail than GABARAP. These results suggest that because of its stronger binding for hKOPR, GEC1 is able to be recruited by hKOPR sufficiently without membrane association via its C-terminal modification; however, due to its weaker affinity for the hKOPR, GABARAP appears to require C-terminal modifications to enhance KOPR expression.