The Journal of Biological Chemistry最新文献_第7页

Identification of hydrogen bonding network for proton transfer at the quinol oxidation site of Rhodobacter capsulatus cytochrome bc₁. 荚膜红杆菌细胞色素bc1喹啉氧化位点质子转移氢键网络的鉴定。

The Journal of Biological Chemistry Pub Date : 2023-10-01 Epub Date: 2023-09-14 DOI: 10.1016/j.jbc.2023.105249

Arkadiusz Borek, Anna Wójcik-Augustyn, Patryk Kuleta, Robert Ekiert, Artur Osyczka

{"title":"Identification of hydrogen bonding network for proton transfer at the quinol oxidation site of Rhodobacter capsulatus cytochrome bc1.","authors":"Arkadiusz Borek, Anna Wójcik-Augustyn, Patryk Kuleta, Robert Ekiert, Artur Osyczka","doi":"10.1016/j.jbc.2023.105249","DOIUrl":"10.1016/j.jbc.2023.105249","url":null,"abstract":"Cytochrome bc1 catalyzes electron transfer from quinol (QH2) to cytochrome c in reactions coupled to proton translocation across the energy-conserving membrane. Energetic efficiency of the catalytic cycle is secured by a two-electron and two-proton bifurcation reaction leading to oxidation of QH2 and reduction of the Rieske cluster and heme bL. The proton paths associated with this reaction remain elusive. Here, we used site-directed mutagenesis and quantum mechanical calculations to analyze the contribution of protonable side chains located at the heme bL side of the QH2 oxidation site in Rhodobacter capsulatus cytochrome bc1. We observe that the proton path is effectively switched off when H276 and E295 are simultaneously mutated to the nonprotonable residues in the H276F/E295V double mutant. The two single mutants, H276F or E295V, are less efficient but still transfer protons at functionally relevant rates. Natural selection exposed two single mutations, N279S and M154T, that restored the functional proton transfers in H276F/E295V. Quantum mechanical calculations indicated that H276F/E295V traps the side chain of Y147 in a position distant from QH2, whereas either N279S or M154T induce local changes releasing Y147 from that position. This shortens the distance between the protonable groups of Y147 and D278 and/or increases mobility of the Y147 side chain, which makes Y147 efficient in transferring protons from QH2 toward D278 in H276F/E295V. Overall, our study identified an extended hydrogen bonding network, build up by E295, H276, D278, and Y147, involved in efficient proton removal from QH2 at the heme bL side of QH2 oxidation site.","PeriodicalId":22621,"journal":{"name":"The Journal of Biological Chemistry","volume":" ","pages":"105249"},"PeriodicalIF":0.0,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10583091/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10610445","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

G protein-receptor kinases 5/6 are the key regulators of G protein-coupled receptor 35-arrestin interactions. G蛋白受体激酶5/6是G蛋白偶联受体35抑制蛋白相互作用的关键调节因子。

The Journal of Biological Chemistry Pub Date : 2023-10-01 Epub Date: 2023-09-01 DOI: 10.1016/j.jbc.2023.105218

Amlan Ganguly, Tezz Quon, Laura Jenkins, Babu Joseph, Rima Al-Awar, Andy Chevigne, Andrew B Tobin, David E Uehling, Carsten Hoffmann, Julia Drube, Graeme Milligan

{"title":"G protein-receptor kinases 5/6 are the key regulators of G protein-coupled receptor 35-arrestin interactions.","authors":"Amlan Ganguly, Tezz Quon, Laura Jenkins, Babu Joseph, Rima Al-Awar, Andy Chevigne, Andrew B Tobin, David E Uehling, Carsten Hoffmann, Julia Drube, Graeme Milligan","doi":"10.1016/j.jbc.2023.105218","DOIUrl":"10.1016/j.jbc.2023.105218","url":null,"abstract":"Human G protein-coupled receptor 35 is regulated by agonist-mediated phosphorylation of a set of five phospho-acceptor amino acids within its C-terminal tail. Alteration of both Ser300 and Ser303 to alanine in the GPR35a isoform greatly reduces the ability of receptor agonists to promote interactions with arrestin adapter proteins. Here, we have integrated the use of cell lines genome edited to lack expression of combinations of G protein receptor kinases (GRKs), selective small molecule inhibitors of subsets of these kinases, and antisera able to specifically identify either human GPR35a or mouse GPR35 only when Ser300 and Ser303 (orce; the equivalent residues in mouse GPR35) have become phosphorylated to demonstrate that GRK5 and GRK6 cause agonist-dependent phosphorylation of these residues. Extensions of these studies demonstrated the importance of the GRK5/6-mediated phosphorylation of these amino acids for agonist-induced internalization of the receptor. Homology and predictive modeling of the interaction of human GPR35 with GRKs showed that the N terminus of GRK5 is likely to dock in the same methionine pocket on the intracellular face of GPR35 as the C terminus of the α5 helix of Gα13 and, that while this is also the case for GRK6, GRK2 and GRK3 are unable to do so effectively. These studies provide unique and wide-ranging insights into modes of regulation of GPR35, a receptor that is currently attracting considerable interest as a novel therapeutic target in diseases including ulcerative colitis.","PeriodicalId":22621,"journal":{"name":"The Journal of Biological Chemistry","volume":" ","pages":"105218"},"PeriodicalIF":0.0,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10520886/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10518909","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

The autoactivation of human single-chain urokinase-type plasminogen activator (uPA). 人单链尿激酶型纤溶酶原激活剂（uPA）的自动激活。

The Journal of Biological Chemistry Pub Date : 2023-10-01 Epub Date: 2023-08-20 DOI: 10.1016/j.jbc.2023.105179

Constanza Torres-Paris, Yueyi Chen, Lufan Xiao, Harriet J Song, Pingyu Chen, Elizabeth A Komives

{"title":"The autoactivation of human single-chain urokinase-type plasminogen activator (uPA).","authors":"Constanza Torres-Paris, Yueyi Chen, Lufan Xiao, Harriet J Song, Pingyu Chen, Elizabeth A Komives","doi":"10.1016/j.jbc.2023.105179","DOIUrl":"10.1016/j.jbc.2023.105179","url":null,"abstract":"Most serine proteases are synthesized as inactive zymogens that are activated by cleavage by another protease in a tightly regulated mechanism. The urokinase-type plasminogen activator (uPA) and plasmin cleave and activate each other, constituting a positive feedback loop. How this mutual activation cycle begins has remained a mystery. We used hydrogen deuterium exchange mass spectrometry to characterize the dynamic differences between the inactive single-chain uPA (scuPA) and its active form two-chain uPA (tcuPA). The results show that the C-terminal β-barrel and the area around the new N terminus have significantly reduced dynamics in tcuPA as compared with scuPA. We also show that the zymogen scuPA is inactive but can, upon storage, become active in the absence of external proteases. In addition to plasmin, the tcuPA can activate scuPA by cleavage at K158, a process called autoactivation. Unexpectedly, tcuPA can cleave at position 158 even when this site is mutated. TcuPA can also cleave scuPA after K135 or K136 in the disordered linker, which generates the soluble protease domain of uPA. Plasmin cleaves scuPA exclusively after K158 and at a faster rate than tcuPA. We propose a mechanism by which the uPA receptor dimerization could promote autoactivation of scuPA on cell surfaces. These results resolve long-standing controversies in the literature surrounding the mechanism of uPA activation.","PeriodicalId":22621,"journal":{"name":"The Journal of Biological Chemistry","volume":" ","pages":"105179"},"PeriodicalIF":0.0,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10520878/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10041911","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Oocytes orchestrate protein prenylation for mitochondrial function through selective inactivation of cholesterol biosynthesis in murine species. 卵母细胞通过选择性失活小鼠体内的胆固醇生物合成来协调线粒体功能的蛋白质异戊二烯化。

The Journal of Biological Chemistry Pub Date : 2023-10-01 Epub Date: 2023-08-21 DOI: 10.1016/j.jbc.2023.105183

Yongjuan Sang, Qiwen Yang, Yueshuai Guo, Xiaofei Liu, Di Shen, Chen Jiang, Xinying Wang, Kang Li, Haiquan Wang, Chaofan Yang, Lijun Ding, Haixiang Sun, Xuejiang Guo, Chaojun Li

{"title":"Oocytes orchestrate protein prenylation for mitochondrial function through selective inactivation of cholesterol biosynthesis in murine species.","authors":"Yongjuan Sang, Qiwen Yang, Yueshuai Guo, Xiaofei Liu, Di Shen, Chen Jiang, Xinying Wang, Kang Li, Haiquan Wang, Chaofan Yang, Lijun Ding, Haixiang Sun, Xuejiang Guo, Chaojun Li","doi":"10.1016/j.jbc.2023.105183","DOIUrl":"10.1016/j.jbc.2023.105183","url":null,"abstract":"Emerging research and clinical evidence suggest that the metabolic activity of oocytes may play a pivotal role in reproductive anomalies. However, the intrinsic mechanisms governing oocyte development regulated by metabolic enzymes remain largely unknown. Our investigation demonstrates that geranylgeranyl diphosphate synthase1 (Ggps1), the crucial enzyme in the mevalonate pathway responsible for synthesizing isoprenoid metabolite geranylgeranyl pyrophosphate from farnesyl pyrophosphate, is essential for oocyte maturation in mice. Our findings reveal that the deletion of Ggps1 that prevents protein prenylation in fully grown oocytes leads to subfertility and offspring metabolic defects without affecting follicle development. Oocytes that lack Ggps1 exhibit disrupted mitochondrial homeostasis and the mitochondrial defects arising from oocytes are inherited by the fetal offspring. Mechanistically, the excessive farnesylation of mitochondrial ribosome protein, Dap3, and decreased levels of small G proteins mediate the mitochondrial dysfunction induced by Ggps1 deficiency. Additionally, a significant reduction in Ggps1 levels in oocytes is accompanied by offspring defects when females are exposed to a high-cholesterol diet. Collectively, this study establishes that mevalonate pathway-protein prenylation is vital for mitochondrial function in oocyte maturation and provides evidence that the disrupted protein prenylation resulting from an imbalance between farnesyl pyrophosphate and geranylgeranyl pyrophosphate is the major mechanism underlying impairment of oocyte quality induced by high cholesterol.","PeriodicalId":22621,"journal":{"name":"The Journal of Biological Chemistry","volume":" ","pages":"105183"},"PeriodicalIF":0.0,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10534227/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10061487","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Structural and functional insights into the Pseudomonas aeruginosa glycosyltransferase WaaG and the implications for lipopolysaccharide biosynthesis. 铜绿假单胞菌糖基转移酶WaaG的结构和功能见解及其对脂多糖生物合成的影响。

The Journal of Biological Chemistry Pub Date : 2023-10-01 Epub Date: 2023-09-15 DOI: 10.1016/j.jbc.2023.105256

Emma R Scaletti, Pontus Pettersson, Joan Patrick, Patrick J Shilling, Robert Gustafsson Westergren, Daniel O Daley, Lena Mäler, Göran Widmalm, Pål Stenmark

{"title":"Structural and functional insights into the Pseudomonas aeruginosa glycosyltransferase WaaG and the implications for lipopolysaccharide biosynthesis.","authors":"Emma R Scaletti, Pontus Pettersson, Joan Patrick, Patrick J Shilling, Robert Gustafsson Westergren, Daniel O Daley, Lena Mäler, Göran Widmalm, Pål Stenmark","doi":"10.1016/j.jbc.2023.105256","DOIUrl":"10.1016/j.jbc.2023.105256","url":null,"abstract":"The glycosyltransferase WaaG in Pseudomonas aeruginosa (PaWaaG) is involved in the synthesis of the core region of lipopolysaccharides. It is a promising target for developing adjuvants that could help in the uptake of antibiotics. Herein, we have determined structures of PaWaaG in complex with the nucleotide-sugars UDP-glucose, UDP-galactose, and UDP-GalNAc. Structural comparison with the homolog from Escherichia coli (EcWaaG) revealed five key differences in the sugar-binding pocket. Solution-state NMR analysis showed that WT PaWaaG specifically hydrolyzes UDP-GalNAc and unlike EcWaaG, does not hydrolyze UDP-glucose. Furthermore, we found that a PaWaaG mutant (Y97F/T208R/N282A/T283A/T285I) designed to resemble the EcWaaG sugar binding site, only hydrolyzed UDP-glucose, underscoring the importance of the identified amino acids in substrate specificity. However, neither WT PaWaaG nor the PaWaaG mutant capable of hydrolyzing UDP-glucose was able to complement an E. coli ΔwaaG strain, indicating that more remains to be uncovered about the function of PaWaaG in vivo. This structural and biochemical information will guide future structure-based drug design efforts targeting PaWaaG.","PeriodicalId":22621,"journal":{"name":"The Journal of Biological Chemistry","volume":" ","pages":"105256"},"PeriodicalIF":0.0,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10579960/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10269215","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

The biased M3 mAChR ligand PD 102807 mediates qualitatively distinct signaling to regulate airway smooth muscle phenotype. 偏向性M3 mAChR配体PD 102807介导性质上不同的信号传导以调节气道平滑肌表型。

The Journal of Biological Chemistry Pub Date : 2023-10-01 Epub Date: 2023-09-01 DOI: 10.1016/j.jbc.2023.105209

Eric Tompkins, Bogdana Mimic, Raymond B Penn, Tonio Pera

{"title":"The biased M3 mAChR ligand PD 102807 mediates qualitatively distinct signaling to regulate airway smooth muscle phenotype.","authors":"Eric Tompkins, Bogdana Mimic, Raymond B Penn, Tonio Pera","doi":"10.1016/j.jbc.2023.105209","DOIUrl":"10.1016/j.jbc.2023.105209","url":null,"abstract":"Airway smooth muscle (ASM) cells attain a hypercontractile phenotype during obstructive airway diseases. We recently identified a biased M3 muscarinic acetylcholine receptor (mAChR) ligand, PD 102807, that induces GRK-/arrestin-dependent AMP-activated protein kinase (AMPK) activation to inhibit transforming growth factor-β-induced hypercontractile ASM phenotype. Conversely, the balanced mAChR agonist, methacholine (MCh), activates AMPK yet does not regulate ASM phenotype. In the current study, we demonstrate that PD 102807- and MCh-induced AMPK activation both depend on Ca2+/calmodulin-dependent kinase kinases (CaMKKs). However, MCh-induced AMPK activation is calcium-dependent and mediated by CaMKK1 and CaMKK2 isoforms. In contrast, PD 102807-induced signaling is calcium-independent and mediated by the atypical subtype protein kinase C-iota and the CaMKK1 (but not CaMKK2) isoform. Both MCh- and PD 102807-induced AMPK activation involve the AMPK α1 isoform. PD 102807-induced AMPK α1 (but not AMPK α2) isoform activation mediates inhibition of the mammalian target of rapamycin complex 1 (mTORC1) in ASM cells, as demonstrated by increased Raptor (regulatory-associated protein of mTOR) phosphorylation as well as inhibition of phospho-S6 protein and serum response element-luciferase activity. The mTORC1 inhibitor rapamycin and the AMPK activator metformin both mimic the ability of PD 102807 to attenuate transforming growth factor-β-induced α-smooth muscle actin expression (a marker of hypercontractile ASM). These data indicate that PD 102807 transduces a signaling pathway (AMPK-mediated mTORC1 inhibition) qualitatively distinct from canonical M3 mAChR signaling to prevent pathogenic remodeling of ASM, thus demonstrating PD 102807 is a biased M3 mAChR ligand with therapeutic potential for the management of obstructive airway disease.","PeriodicalId":22621,"journal":{"name":"The Journal of Biological Chemistry","volume":" ","pages":"105209"},"PeriodicalIF":0.0,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10520882/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10499782","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Sialylation of EGFR by ST6GAL1 induces receptor activation and modulates trafficking dynamics. ST6GAL1对EGFR的唾液酸化诱导受体活化并调节运输动力学。

The Journal of Biological Chemistry Pub Date : 2023-10-01 Epub Date: 2023-09-01 DOI: 10.1016/j.jbc.2023.105217

Katherine E Ankenbauer, Tejeshwar C Rao, Alexa L Mattheyses, Susan L Bellis

{"title":"Sialylation of EGFR by ST6GAL1 induces receptor activation and modulates trafficking dynamics.","authors":"Katherine E Ankenbauer, Tejeshwar C Rao, Alexa L Mattheyses, Susan L Bellis","doi":"10.1016/j.jbc.2023.105217","DOIUrl":"10.1016/j.jbc.2023.105217","url":null,"abstract":"Aberrant glycosylation is a hallmark of a cancer cell. One prevalent alteration is an enrichment in α2,6-linked sialylation of N-glycosylated proteins, a modification directed by the ST6GAL1 sialyltransferase. ST6GAL1 is upregulated in many malignancies including ovarian cancer. Prior studies have shown that the addition of α2,6 sialic acid to the epidermal growth factor receptor (EGFR) activates this receptor, although the mechanism was largely unknown. To investigate the role of ST6GAL1 in EGFR activation, ST6GAL1 was overexpressed in the OV4 ovarian cancer line, which lacks endogenous ST6GAL1, or knocked-down in the OVCAR-3 and OVCAR-5 ovarian cancer lines, which have robust ST6GAL1 expression. Cells with high expression of ST6GAL1 displayed increased activation of EGFR and its downstream signaling targets, AKT and NFκB. Using biochemical and microscopy approaches, including total internal reflection fluorescence microscopy, we determined that the α2,6 sialylation of EGFR promoted its dimerization and higher order oligomerization. Additionally, ST6GAL1 activity was found to modulate EGFR trafficking dynamics following EGF-induced receptor activation. Specifically, EGFR sialylation enhanced receptor recycling to the cell surface following activation while simultaneously inhibiting lysosomal degradation. 3D widefield deconvolution microscopy confirmed that in cells with high ST6GAL1 expression, EGFR exhibited greater colocalization with Rab11 recycling endosomes and reduced colocalization with LAMP1-positive lysosomes. Collectively, our findings highlight a novel mechanism by which α2,6 sialylation promotes EGFR signaling by facilitating receptor oligomerization and recycling.","PeriodicalId":22621,"journal":{"name":"The Journal of Biological Chemistry","volume":" ","pages":"105217"},"PeriodicalIF":0.0,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10520885/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10235659","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Knockout of c-Cbl/Cbl-b slows c-Met trafficking resulting in enhanced signaling in corneal epithelial cells. c-Cbl/Cbl-b的敲除减缓了c-Met的运输，从而增强了角膜上皮细胞中的信号传导。

The Journal of Biological Chemistry Pub Date : 2023-10-01 Epub Date: 2023-09-09 DOI: 10.1016/j.jbc.2023.105233

Kate Tarvestad-Laise, Brian P Ceresa

{"title":"Knockout of c-Cbl/Cbl-b slows c-Met trafficking resulting in enhanced signaling in corneal epithelial cells.","authors":"Kate Tarvestad-Laise, Brian P Ceresa","doi":"10.1016/j.jbc.2023.105233","DOIUrl":"10.1016/j.jbc.2023.105233","url":null,"abstract":"In many cell types, the E3 ubiquitin ligases c-Cbl and Cbl-b induce ligand-dependent ubiquitylation of the hepatocyte growth factor (HGF)-stimulated c-Met receptor and target it for lysosomal degradation. This study determines whether c-Cbl/Cbl-b are negative regulators of c-Met in the corneal epithelium (CE) and if their inhibition can augment c-Met-mediated CE homeostasis. Immortalized human corneal epithelial cells were transfected with Cas9 only (Cas9, control cells) or with Cas9 and c-Cbl/Cbl-b guide RNAs to knockout each gene singularly (-c-Cbl or -Cbl-b cells) or both genes (double KO [DKO] cells) and monitored for their responses to HGF. Cells were assessed for ligand-dependent c-Met ubiquitylation via immunoprecipitation, magnitude, and duration of c-Met receptor signaling via immunoblot and receptor trafficking by immunofluorescence. Single KO cells displayed a decrease in receptor ubiquitylation and an increase in phosphorylation compared to control. DKO cells had no detectable ubiquitylation, had delayed receptor trafficking, and a 2.3-fold increase in c-Met phosphorylation. Based on the observed changes in receptor trafficking and signaling, we examined HGF-dependent in vitro wound healing via live-cell time-lapse microscopy in control and DKO cells. HGF-treated DKO cells healed at approximately twice the rate of untreated cells. From these data, we have generated a model in which c-Cbl/Cbl-b mediate the ubiquitylation of c-Met, which targets the receptor through the endocytic pathway toward lysosomal degradation. In the absence of ubiquitylation, the stimulated receptor stays phosphorylated longer and enhances in vitro wound healing. We propose that c-Cbl and Cbl-b are promising pharmacologic targets for enhancing c-Met-mediated CE re-epithelialization.","PeriodicalId":22621,"journal":{"name":"The Journal of Biological Chemistry","volume":" ","pages":"105233"},"PeriodicalIF":0.0,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10622846/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10202585","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Methylation of the transcription factor E2F1 by SETD6 regulates SETD6 expression via a positive feedback mechanism. SETD6对转录因子E2F1的甲基化通过正反馈机制调节SETD6的表达。

The Journal of Biological Chemistry Pub Date : 2023-10-01 Epub Date: 2023-09-09 DOI: 10.1016/j.jbc.2023.105236

Margarita Kublanovsky, Gizem T Ulu, Sara Weirich, Nurit Levy, Michal Feldman, Albert Jeltsch, Dan Levy

{"title":"Methylation of the transcription factor E2F1 by SETD6 regulates SETD6 expression via a positive feedback mechanism.","authors":"Margarita Kublanovsky, Gizem T Ulu, Sara Weirich, Nurit Levy, Michal Feldman, Albert Jeltsch, Dan Levy","doi":"10.1016/j.jbc.2023.105236","DOIUrl":"10.1016/j.jbc.2023.105236","url":null,"abstract":"The protein lysine methyltransferase SET domain-containing protein 6 (SETD6) has been shown to influence different cellular activities and to be critically involved in the regulation of diverse developmental and pathological processes. However, the upstream signals that regulate the mRNA expression of SETD6 are not known. Bioinformatic analysis revealed that the SETD6 promoter has a binding site for the transcription factor E2F1. Using various experimental approaches, we show that E2F1 binds to the SETD6 promoter and regulates SETD6 mRNA expression. Our further observation that this phenomenon is SETD6 dependent suggested that SETD6 and E2F1 are linked. We next demonstrate that SETD6 monomethylates E2F1 specifically at K117 in vitro and in cells. Finally, we show that E2F1 methylation at K117 positively regulates the expression level of SETD6 mRNA. Depletion of SETD6 or overexpression of E2F1 K117R mutant, which cannot be methylated by SETD6, reverses the effect. Taken together, our data provide evidence for a positive feedback mechanism, which regulates the expression of SETD6 by E2F1 in a SETD6 methylation-dependent manner, and highlight the importance of protein lysine methyltransferases and lysine methylation signaling in the regulation of gene transcription.","PeriodicalId":22621,"journal":{"name":"The Journal of Biological Chemistry","volume":" ","pages":"105236"},"PeriodicalIF":0.0,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10551896/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10204909","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Multiple mechanisms of self-association of chemokine receptors CXCR4 and CCR5 demonstrated by deep mutagenesis. 通过深度诱变证明了趋化因子受体CXCR4和CCR5的多种自缔合机制。

The Journal of Biological Chemistry Pub Date : 2023-10-01 Epub Date: 2023-09-09 DOI: 10.1016/j.jbc.2023.105229

Kevin S Gill, Kritika Mehta, Jeremiah D Heredia, Vishnu V Krishnamurthy, Kai Zhang, Erik Procko

{"title":"Multiple mechanisms of self-association of chemokine receptors CXCR4 and CCR5 demonstrated by deep mutagenesis.","authors":"Kevin S Gill, Kritika Mehta, Jeremiah D Heredia, Vishnu V Krishnamurthy, Kai Zhang, Erik Procko","doi":"10.1016/j.jbc.2023.105229","DOIUrl":"10.1016/j.jbc.2023.105229","url":null,"abstract":"Chemokine receptors are members of the rhodopsin-like class A GPCRs whose signaling through G proteins drives the directional movement of cells in response to a chemokine gradient. Chemokine receptors CXCR4 and CCR5 have been extensively studied due to their roles in leukocyte development and inflammation and their status as coreceptors for HIV-1 infection, among other roles. Both receptors form dimers or oligomers of unclear function. While CXCR4 has been crystallized in a dimeric arrangement, available atomic resolution structures of CCR5 are monomeric. To investigate their dimerization interfaces, we used a bimolecular fluorescence complementation (BiFC)-based screen and deep mutational scanning to find mutations that change how the receptors self-associate, either via specific oligomer assembly or alternative mechanisms of clustering in close proximity. Many disruptive mutations promoted self-associations nonspecifically, suggesting they aggregated in the membrane. A mutationally intolerant region was found on CXCR4 that matched the crystallographic dimer interface, supporting this dimeric arrangement in living cells. A mutationally intolerant region was also observed on the surface of CCR5 by transmembrane helices 3 and 4. Mutations predicted from the scan to reduce BiFC were validated and were localized in the transmembrane domains as well as the C-terminal cytoplasmic tails where they reduced lipid microdomain localization. A mutation in the dimer interface of CXCR4 had increased binding to the ligand CXCL12 and yet diminished calcium signaling. There was no change in syncytia formation with cells expressing HIV-1 Env. The data highlight that multiple mechanisms are involved in self-association of chemokine receptor chains.","PeriodicalId":22621,"journal":{"name":"The Journal of Biological Chemistry","volume":" ","pages":"105229"},"PeriodicalIF":0.0,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10551899/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10596318","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0