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Receptor fusions as a method to quantify signaling bias

Receptor fusions as a method to quantify signaling bias

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中国生物化学与分子生物学会第十一次会员代表大会暨2014年全国学术会议

作者：孙金鹏山东大学

The concept of biased agonism arises from the recognition that the ability of an agonist toinduce a receptor-mediated response(i.e.,efficacy)can differ across the multiple signaltransduction pathways(e.g.,G protein,GRKs andβ-arrestin(βarr))emanating from a *** the therapeutic promise of biased agonism,the molecular mechanism(s)whereby biased agonists selectively engage signaling pathways remain *** is due,in large part,to the challenges associated with quantifying ligand efficacy in *** addressthis,we developed a cell-free approach to directly quantify the transducer-specificmolecular efficacies of balanced and biased ligands for the Beta adrenergic receptor,aprototypic ***,we defined efficacy in allosteric terms,equating shifts in ligandaffinity(i.e.,KLo/KHi)at B2AR-Gs and B2AR-βarr2 fusion proteins with their respectivemolecular efficacies for activating Gs andβ*** with ternary complex modelpredictions,transducer-specific molecular efficacies were strongly correlated with cellularefficacies for activating Gs andβ*** comparisons across transducers revealedthat biased B2ARR agonists possess biased molecular efficacies which were in strongagreement with the signaling bias observed in cellular *** findings not only representthe first measurements of the thermodynamic driving forces underlying differencesin ligand efficacy between transducers,but also support a molecular mechanism wherebydivergent transducer-specific molecular efficacies generate biased agonism at a GPCR.

关键词： GPCR biased signaling

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Deregulation of H2AK119 Monoubiquitylation in Cancer

Deregulation of H2AK119 Monoubiquitylation in Cancer

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中国生物化学与分子生物学会第十一次会员代表大会暨2014年全国学术会议

作者： Xudong Wu Tianjin Medical University

The importance of genetic mutations and deletions in cancer has long been ***, mutations of chromatin modifier genes are frequently identified by widespreadapplication of genome sequencing in tumor samples. Recently, ASXL1 mutationshave been reported in patients with diverse tumors especially hematological *** ASXL1 mutations are significantly correlated with oncogenic RAS mutations. Othersand we found that ASXL1 interacts with BAP1, a deubiquitylase specifically targeting PRC1-mediated H2 A monoubiquitylation on lysine 119(H2AK119ub1). Given that oncogenic RASalone causes cell senescence, we tried to investigate whether ASXL1 depletion bypassesthe oncogeneinduced senescence and therefore cooperate in tumorigenesis. Here wedemonstrate that ASXL1 acts as a tumor suppressor whose downregulation attenuates theanti-proliferative effect in response to the activation of RAS-BRAF pathways in either epithelialor myeloid cells. In the meeting, we will present some mechanistic data showing how ASXL1-containing H2 A deubiquitylation complex affects the essential down stream targets of RAS pathway. It will provide a novel insight on the dynamic H2AK119 monoubiquitylationin the regulation of cell fates.

关键词： H2AK119ub1 ASXL1 Tumor suppressor

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In vivo substrate diversity and preference of small heat shock protein IbpB as revealed by using a genetically incorporated photocrosslinker

In vivo substrate diversity and preference of small heat sho...

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中国生物化学与分子生物学会第十一次会员代表大会暨2014年全国学术会议

作者：付新苗石小东昌增益北京大学生命科学学院徐州医学院

Small heat shock proteins(sHSPs),as ubiquitous molecular chaperones,are known to beable to protect cells against stresses and suppress the aggregation of a variety of modelsubstrate proteins under in vitro ***,it is poorly understood whatnatural substrate proteins sHSPs act upon in living *** by using a genetically incorporatedphotocrosslinker(Bpa),we identified a total of 95 and 54 natural substrate proteinsof IbpB(a sHSP from Escherichia coli)in living cells with and without heat shock,*** profiling of these proteins(110 in total)suggests that IbpB,althoughbinding to a wide range of cellular proteins,has a remarkable substrate preference fortranslation-related proteins(e.g.,ribosomal proteins and amino-acyl tRNA synthetases)and moderate preference for metabolic ***,these two classes of proteinswere found to be more prone to aggregation and/or inactivation in cells lacking IbpB understress conditions(e.g.,heat shock).Together,our in vivo data offer novel insights intothe chaperone function of IbpB,or sHSPs in general,and suggest that the preferential protectionon protein synthesis machine and metabolic enzymes may dominantly contributeto the well-known protective effect of sHSPs on cell survival against stresses.

关键词： molecular chaperone small heat shock protein unnatural amino acid photoc rosslinking

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Discovery of PHGPx allosteric activators

Discovery of PHGPx allosteric activators

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中国生物化学与分子生物学会第十一次会员代表大会暨2014年全国学术会议

作者： Cong Li Xiaowen Xie Luhua Lai College of Chemistry and Molecular Engineering StateKey Laboratory for Structural Chemistry of Unstable and Stable Species Peking University Center for Quantitative Biology Peking University College of Chemistry and Molecular Engineering State Key Laboratory for Structural Chemistry of Unstable and Stable SpeciesPeking University

Phospholipid hydroperoxide glutathione peroxidases(PHGPx) is capable of reducing oxidativedamaged biomembranes and reactive oxygen species(ROS) in cells. It plays irreplaceablerole in embryonic development, antioxidation, and male fertility. Activating theprotective effect of PHGPx might be used to cure inflammation, cancer, and other ***, activating target protein by allosteric modulation is a novel strategy in drugdesign. Until now, no compound has been reported to activate the enzymatic activity ofPHGPx. Here, we predicted the allosteric site of PHGPx based on a crystal structure withenzymatic activity, and used structure-based virtual screen for PHGPx activators *** the in vitro activity assay, we got 7 compounds which can activate PHGPx. The bestone has AC50 of 58 ± 5 μM, and can increase the enzymatic activity to about two foldcompared to wild type PHGPx. Moreover, it also exhibits PHGPx activity activation in livingcells. The discovery of these activators verified the possibility to design drugs specific tothe allosteric site, especially allosteric activators. Furthermore, these activators provide anew method to inhibit inflammation more effectively and with less side effects.

关键词： PHGPx allosteric modulation activator

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Tanshinone suppresses AURKA through up-regulating theexpression of miR-32 in NSCLC

Tanshinone suppresses AURKA through up-regulating theexpress...

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中国生物化学与分子生物学会第十一次会员代表大会暨2014年全国学术会议

作者： zhongliang ma bingjie zhang detao wang xue li jiali wei botao zhao youxin jin School of life sciences Shanghai University School of Life Sciences Shanghai University State Key Laboratory of Molecular Biology Institute of Biochemistry and Cell Biology Institutes for Biological SciencesChinese Academy of Sciences

Tanshinone, including tanshinone I(T1), tanshinone IIA(T2A), cryptotanshinone(CT), is theliposoluble constituent of Salia miltiorrhiza which has favourable medicinal value. Tanshinonemolecules can inhibit the growth of lung cancer cells in vitro, but its mechanism isstill unclear. AURKA, an oncogene, encodes a serine-threonine kinase which regulates mitoticprocesses in mammalian cells. Here, we report that tanshinone mediates AURKA suppressionpartly through up-regulating the expression of miR-32. We found that tanshinonecould inhibit cell proliferation, promote apoptosis and impede cell-cycle progression thusperforming an antineoplastic function in non-small cell lung cancer(NSCLC). As reported,tanshinone might attain these effects partly by down-regulating AURKA and such was thefact demonstrated by us. Results showed that AURKA gene knocked-down by siRNA couldmake similar effects in NSCLC. We also verified that AURKA was the target of miR-32. Inconclusion, our results demonstrated that tanshinone could inhibit NSCLC by upregulating the expressions of miR-32 and other related miRNAs to suppress AURKA.

关键词： Tanshinone AURKA miRNA NSCLC

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A primary study on mutation of HPRT exposure to γ ray andbenzo-pyrene

A primary study on mutation of HPRT exposure to γ ray andbe...

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中国生物化学与分子生物学会第十一次会员代表大会暨2014年全国学术会议

作者：刘建功 Hong-yan LIU Xu-hong DANG China Institute for Radiation Protection

Objective To investigate the changes of mutation of HPRT exposure to γ ray and benzopyrene,in order to try to find new differential diagnosis methods for cell injury induced byγ ray and benzo-pyrene. Methods Human peripheral blood samples from three personswere exposed to 60 Co γ-rays at 10 different doses(0-5Gy) and benzo-pyrene at 6 differentconcentrations(0-10μg/ml), its DNA was isolated from white cell. The changes of mutationof HPRT exposure to γ ray and benzo-pyrene were detected by real-time quantitative *** The relationship between mutation frequency of HPRT and irradiation dose represen tscertainline correlation from 0～5Gy, alinearregressionequa-tion(y=0.4298+0.0823 x,P<0.01,R2=0.93) has been obtained using statistics regressionanalysis software, while, the relationship between mutation frequency of HPRT and differentconcentrations(0-10μg/ml) represents no statistical significance,which was obtainedusing statistics software as(y=0.4200+0.0019 x, P>0.05, R2=0.450). There is an obvious influenceon mutation frequency of exon 5 of HPRT induced byγray,while there is no obvious influence on mutation frequency of exon 5 of HPRT induced by benzo-pyrene,which may be a new differential diagnosis index in the future.

关键词： γ ray benzo-pyrene mutation frequency HPRT

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Small heat shock protein IbpB acts as a robust chaperone in living cells by hierarchically activating its multi-type substrate-binding residues

Small heat shock protein IbpB acts as a robust chaperone in ...

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中国生物化学与分子生物学会第十一次会员代表大会暨2014年全国学术会议

作者：付新苗石小东昌增益北京大学生命科学学院徐州医学院

Small heat shock proteins(sHSPs),as ubiquitous molecular chaperones,are crucial for *** is not clear why sHSPs are able to bind a wide spectrum of non-nativesubstrate proteins and how such binding is enhanced by heat ***,by utilizing agenetically incorporated photo-crosslinker(Bpa),we systematically characterized the substrate-binding residues in IbpB(a sHSP from Escherichia coli)in living cells over a widespectrum of temperatures(from 20oC to 50oC).A total of 20 and 48 residues were respectivelyidentified at normal and heat shock *** are not necessarily hydrophobicand can be classified into three types:type I and II are respectively activated atlow and normal temperatures;type III mediate oligomerization at low temperature butswitch to substrate-binding at heat shock *** addition,substrate-binding ofIbpB in living cells begins at such a low temperature as 25oC and is further enhanced upontemperature ***,these in vivo data provide novel structural insights intothe wide substrate spectrum of sHSP and suggest that sHSP is able to hierarchically activateits multi-type substrate-binding residues and thus act as a robust chaperone in cellsunder fluctuating growth conditionsReference.

关键词： molecular chaperone small heat shock protein unnatural amino acid photocrosslinking

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Characterize the interactions essential for SIVrcm Vpx-mediated DCAF1/DDB1/Cul4-dependent degradation of red-capped mangabey SAMHD1

Characterize the interactions essential for SIVrcm Vpx-media...

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中国生物化学与分子生物学会第十一次会员代表大会暨2014年全国学术会议

作者： Jian Li Fei Guo Chen Liang Institute of Pathogen Biology Chinese Academy of Medical Sciences and Peking Union Medical College

SAMHD1 inhibits HIV-1 infection of myeloid cells and resting CD4+T ***-2 andsome SIV strains,but not HIV-1,carry a SAMHD1 antagonist called *** recognizesSAMHD1 and recruits the ubiquitin E3 ligase complex CUL4/DDB1/DCAF1 that polyubiquitinatesSAMHD1 followed by proteasomal *** lineages of primate lentivirusescarry Vpx,they are the sooty mangabey SIV(SIVsm)/macaque SIV(SIVmac)/HIV-2lineage and the red-capped mangabey SIV(SIVrcm)*** opposed to the wellcharacterizedinteraction of human SAMHD1,SIVmac Vpx and CUL4/DDB1/DCAF1,SIVrcmVpx adopts a different mode of interaction with SAMHD1 of red-capped *** determined that the 23-WLHR-26 peptide of SIVrcm Vpx is essential for recognizingrcm *** larmodelofthercm SAM-HD1-NtD/Vpxrcm/DCAF1-CtD complex,which reveals a likely much greater interface andmore complex interactions between rcmSAMHD1-NtD and Vpxrcm,as compared to humanSAMHD1-CtD and Vpxsm.

关键词： SAMHD1 SIV Vpx

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MicroRNA-346 Mediates Tumor Necrosis Factor-■-induced Down-Regulation of Vitamin D Receptor in Gut Epithelial Cells

MicroRNA-346 Mediates Tumor Necrosis Factor-■-induced Down-...

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中国生物化学与分子生物学会第十一次会员代表大会暨2014年全国学术会议

作者： yunzi chen 南京医科大学

We recently reported that the gut epithelial vitamin D receptor(VDR) signaling inhibitscolitis through inhibition of intestinal epithelial cell apoptosis, and the level of colonic epithelialVDR is markedly down-regulated in patients with inflammatory bowel diseases(IBD), butthemechanism underlying VDR down-regulation in IBD is unknown. Here we showed thatTNF-αsuppresses VDR expression while simultaneously up-regulating miR-346 in human coloncancercells. Further studies demonstrated that miR-346 inhibits VDR by a specific target sequenceinthe 3' untranslated region of the human VDR transcript, and blockade of miR-346 with ahairpininhibitor abrogates the ability of TNF-α to inhibit VDR. These results indicate that TNF-αdownregulates VDR by inducing miR-346. Consistently, in human IBD biopsies the reductionofepithelial VDR is associated with increased immune cell infiltration and elevation of TNF-αandmiR-346. In an experimental model of colitis mucosal VDR expression is reduced over timewiththe progression of colitis, inversely correlated with the induction of TNF-α and miR-346 inthemucosa. Together these data suggest that during mucosal inflammation TNF-α inducesmiR-346, which down-regulates epithelial VDR. Mucosal VDR reduction in turn compromisestheintegrity of the mucosal epithelial barrier, further driving mucosal inflammation and colitisdevelopment.

关键词： microRNA-346 vitamin Dreceptor

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Benefits of being heteromers: γ-crystallin heteromer is more stable than the homomers during thermal denaturation

Benefits of being heteromers: γ-crystallin heteromer is mor...

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中国生物化学与分子生物学会第十一次会员代表大会暨2014年全国学术会议

作者：冷筱瑶闫永彬 State Key Laboratory of Biomembrane and Membrane Biotechnology School of Life Sciences Tsinghua University

α-,β-andγ-crystallins are the major protein components in human lens.α-Crystallinserves as a molecular chaperone,whileβ-andγ-crystallins are the main structural proteinsin the *** solubility and stability of crystallins are important in maintaining the transparencyand refraction index of *** subject to the persistent environmentalstresses during aging,crystallins may lose their solubility and *** aggregation ofcrystallins will interfere with the vision by scattering of the light,which is generally recognizedas the main cause of aging *** the lens,β-crystallins exist as heteromers with 2-8 subunits using the basic and acidicβ-crystallins as the building *** basicβ-crystallin is generally more stable than acidicβ-*** previous research has shown that the basicβ-crystallin could protect theacidicβ-crystallin against aggregation during refolding and *** this research,we further investigated the sequential events in the thermal denaturation ofβA3-crystallinhomomer,βB1-crystallin homomer andβA3/βB1-crystallin heteromer by a combination ofspectroscopic *** results showed thatβA3/βB1 heteromer was the most stableamong the three proteins,whileβA3-crystallin is the most *** experimentsconfirmed that the Tm ofβA3-crystallin is much lower than those of the other *** results suggested that the formation ofβ-crystallin heteromers not onlystabilized the unstable acidicβ-crystallins,but also improve the stability of the basicβ-*** findings provide insight into the molecular basis of whyβ-crystallins existas heteromers in vivo,and may be important to the understanding of the onset of cataractcaused byβ-crystallin aggregation.

关键词： β-crystallin thermal denaturation heteromer

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