Journal of Biological Chemistry最新文献_第8页

Allosteric factors in the calcium/calmodulin-responsive kinase II hub domain determine selectivity of GHB ligands for CaMKIIα. 钙/钙调素响应激酶II中心结构域的变构因子决定了GHB配体对CaMKIIα的选择性。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-04-24 DOI: 10.1016/j.jbc.2025.108543

Stine Juul Gauger, Line B Palmelund, Yongsong Tian, Aleš Marek, Mathias R Namini, Nane Griem-Krey, Madeline Yde Petersen, Stefanie Kickinger, Jonas S Mortensen, Bente Frølund, Petrine Wellendorph

{"title":"Allosteric factors in the calcium/calmodulin-responsive kinase II hub domain determine selectivity of GHB ligands for CaMKIIα.","authors":"Stine Juul Gauger, Line B Palmelund, Yongsong Tian, Aleš Marek, Mathias R Namini, Nane Griem-Krey, Madeline Yde Petersen, Stefanie Kickinger, Jonas S Mortensen, Bente Frølund, Petrine Wellendorph","doi":"10.1016/j.jbc.2025.108543","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.108543","url":null,"abstract":"The Ca2+/CaM-dependent protein kinase II alpha (CaMKIIα) is a highly important synaptic protein, which comprises a unique holoenzyme structure organized via the central hub domain. Recently, a distinct binding pocket in the CaMKIIα hub domain was identified for the endogenous neuromodulator γ-hydroxybutyric acid (GHB) and related synthetic analogues. Intriguingly, of the four native CaMKII isozymes, only CaMKIIα accommodates GHB ligands. Key interacting residues in CaMKIIα were revealed, but their involvement in selectivity towards the alpha variant of CaMKII has remained unresolved. Aimed at elucidating the molecular determinants for this selectivity, we here conducted binding studies to CaMKII-HEK whole-cell homogenates using two different in-house-developed GHB-related radioligands, [3H]HOCPCA and [3H]O-5-HDC, in combination with site-directed mutagenesis. Binding to CaMKIIα with the smaller-type radioligand [3H]HOCPCA validated key involvement of the four known residues (His395, Arg433, Arg453 and Arg469), but also revealed a role for the upper hub flexible loop containing the CaMKIIα-specific residue Trp403 (Leu in all other CaMKII isozymes) previously suggested to be involved in holoenzyme stability. Insertion of the corresponding residues (L467W/C533R) into CaMKIIβ failed to induce [3H]HOCPCA binding. However, with the larger-type radioligand, [3H]O-5-HDC, specific binding in CaMKIIβ (L467W/C533R) was achieved. Thus, the study confirms involvement of central binding residues and identifies the CaMKIIα flexible pocket loop as a distantly located \"allosteric\" factor in determining selectivity of GHB analogues for CaMKIIα. It sheds light on a remarkable interplay of the entire hub cavity for accommodation of ligands, and corroborates GHB analogues as CaMKIIα-selective.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"108543"},"PeriodicalIF":4.0,"publicationDate":"2025-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143986340","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Phase separation as a key mechanism in plant development, environmental adaptation, and abiotic stress response. 相分离是植物发育、环境适应和非生物胁迫响应的关键机制。

IF 4.8 2区生物学

Journal of Biological Chemistry Pub Date : 2025-04-24 DOI: 10.1016/j.jbc.2025.108548

Karina C Pougy,Bruna A Brito,Giovanna S Melo,Anderson S Pinheiro

{"title":"Phase separation as a key mechanism in plant development, environmental adaptation, and abiotic stress response.","authors":"Karina C Pougy,Bruna A Brito,Giovanna S Melo,Anderson S Pinheiro","doi":"10.1016/j.jbc.2025.108548","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.108548","url":null,"abstract":"Liquid-liquid phase separation is a fundamental biophysical process in which biopolymers, such as proteins, nucleic acids, and their complexes, spontaneously demix into distinct coexisting phases. This phenomenon drives the formation of membraneless organelles-cellular subcompartments without a lipid bilayer that perform specialized functions. In plants, phase-separated biomolecular condensates play pivotal roles in regulating gene expression, from genome organization to transcriptional and post-transcriptional processes. In addition, phase separation governs plant-specific traits, such as flowering and photosynthesis. As sessile organisms, plants have evolved to leverage phase separation for rapid sensing and response to environmental fluctuations and stress conditions. Recent studies highlight the critical role of phase separation in plant adaptation, particularly in response to abiotic stress. This review compiles the latest research on biomolecular condensates in plant biology, providing examples of their diverse functions in development, environmental adaptation, and stress responses. We propose that phase separation represents a conserved and dynamic mechanism enabling plants to adapt efficiently to ever-changing environmental conditions. Deciphering the molecular mechanisms underlying phase separation in plant stress responses opens new avenues for biotechnological strategies aimed at engineering stress-resistant crops. These advancements have significant implications for agriculture, particularly in addressing crop productivity in the face of climate change.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":"55 1","pages":"108548"},"PeriodicalIF":4.8,"publicationDate":"2025-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143885534","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Nick sealing of polβ mismatch insertion products by LIG1 and LIG3α during 8-oxoG bypass leads to mutagenic or error-free base excision repair. 在8-oxoG旁路过程中，LIG1和LIG3α对polβ错配插入产物的缺口密封导致突变或无错误的碱基切除修复。

IF 4.8 2区生物学

Journal of Biological Chemistry Pub Date : 2025-04-24 DOI: 10.1016/j.jbc.2025.108540

Kar Men Lee,Erick Castro,Jacob Ratcliffe,Camden Lerner,Melike Çağlayan

{"title":"Nick sealing of polβ mismatch insertion products by LIG1 and LIG3α during 8-oxoG bypass leads to mutagenic or error-free base excision repair.","authors":"Kar Men Lee,Erick Castro,Jacob Ratcliffe,Camden Lerner,Melike Çağlayan","doi":"10.1016/j.jbc.2025.108540","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.108540","url":null,"abstract":"Base excision repair (BER) requires a coordination at the downstream steps involving gap filling by DNA polymerase (pol) β and subsequent nick sealing by DNA ligase (LIG) 1 or 3α. We previously reported that a failure in DNA ligase function, stemming from an impairment in nick sealing of polβ nucleotide insertion products, leads to faulty repair events. Yet, how the fidelity of 8-oxoG bypass by polβ affects the efficiency of ligation remains unclear. Here, we show that LIG1 and LIG3α seal the resulting nick repair product of polβ mutagenic insertion of dATP opposite 8-oxoG, while LIG3α exhibits an inability to ligate polβ dCTP:8-oxoG insertion product, demonstrating that the identity of BER ligase plays a critical role in repair outcomes at the final step. Furthermore, our results show that a lack of ribonucleotide insertion by polβ during 8-oxoG bypass diminishes the repair coordination with both ligases, highlighting the critical role of nucleotide selectivity in maintaining BER accuracy. Finally, our results reveal that AP-Endonuclease 1 (APE1) proofreads nick repair intermediates containing 3'-mismatches or ribonucleotides templating 8-oxoG. Overall, our findings provide a mechanistic insight into how the dual coding potential of the oxidative lesion in -anti versus -syn conformation could govern error-prone versus error-free repair outcomes, leading to deviations in the BER pathway coordination and the formation of deleterious DNA intermediates.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":"25 1","pages":"108540"},"PeriodicalIF":4.8,"publicationDate":"2025-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143885536","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

CpX Hunter web tool allows high-throughput identification of CpG, CpA, CpT, and CpC islands: A case study in Drosophila genome. CpX猎人网络工具允许高通量鉴定CpG， CpA， CpT和CpC岛：果蝇基因组的案例研究。

IF 4.8 2区生物学

Journal of Biological Chemistry Pub Date : 2025-04-24 DOI: 10.1016/j.jbc.2025.108537

Martin Bartas,Michal Petrovič,Václav Brázda,Oldřich Trenz,Aleš Ďurčanský,Jiří Štastný

{"title":"CpX Hunter web tool allows high-throughput identification of CpG, CpA, CpT, and CpC islands: A case study in Drosophila genome.","authors":"Martin Bartas,Michal Petrovič,Václav Brázda,Oldřich Trenz,Aleš Ďurčanský,Jiří Štastný","doi":"10.1016/j.jbc.2025.108537","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.108537","url":null,"abstract":"With continuous advances in DNA sequencing methods, accessibility to high-quality genomic information for all living organisms is ever increasing. However, to interpret this information effectively and formulate hypotheses, users often require higher level programming skills. Therefore, the generation of web-based tools is becoming increasingly popular. CpG island regions in genomes are often found in gene promoters and are prone to DNA methylation; with their methylation status determining if a gene is expressed. Notably, understanding the biological impact of CpX modifications on genomic regulation is becoming increasingly important as these modifications have been associated with diseases such as cancer and neurodegeneration. However, there is currently no easy-to-use scalable tool to detect and quantify CpX islands in full genomes. We have developed a Java-based web server for CpX island analyses that benefits from the DNA Analyzer Web server environment and overcomes several limitations. For a pilot demonstration study, we selected a well-described model organism Drosophila melanogaster. Subsequent analysis of obtained CpX islands revealed several interesting and previously undescribed phenomena. One of them is the fact, that nearly half of long CpG islands were located on chromosome X, and that long CpA and CpT islands were significantly overrepresented at the subcentromeric regions of autosomes (chr2 and chr3) and also on chromosome Y. Wide genome overlays of predicted CpX islands revealed their co-occurrence with various (epi)genomics features comprising cytosine methylations, accessible chromatin, transposable elements, or binding of transcription factors and other proteins. CpX Hunter is freely available as a web tool at: https://bioinformatics.ibp.cz/#/analyse/cpg.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":"36 1","pages":"108537"},"PeriodicalIF":4.8,"publicationDate":"2025-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143885539","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Quantifying small GTPase activation status using a novel fluorescence HPLC-based assay. 定量小GTPase激活状态使用一种新的荧光高效液相色谱为基础的分析。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-04-24 DOI: 10.1016/j.jbc.2025.108545

Makoto Araki, Yukika Kasuya, Kaho Yoshimoto, Toshiaki Katada, Kenji Kontani

{"title":"Quantifying small GTPase activation status using a novel fluorescence HPLC-based assay.","authors":"Makoto Araki, Yukika Kasuya, Kaho Yoshimoto, Toshiaki Katada, Kenji Kontani","doi":"10.1016/j.jbc.2025.108545","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.108545","url":null,"abstract":"Small GTPases play crucial roles in cellular signaling pathways, with their activation states tightly regulated between GDP-bound inactive and GTP-bound active forms. Dysregulation of these nucleotide-binding states, such as in oncogenic RAS, is implicated in diseases like cancer. Accurately quantifying these states in cells is thus crucial for deciphering their functional roles and regulatory mechanisms. However, current methods do not fully meet the necessary sensitivity and versatility, limiting their effectiveness in small GTPase analysis. Here, we present a highly sensitive HPLC-based assay with fluorescence detection (Fluor-HPLC), enabling precise quantification of guanine nucleotide-binding states in small GTPases. Applying this technique, we successfully quantified the guanine nucleotide-binding states of small GTPases at their endogenous expression levels. We demonstrated the utility of Fluor-HPLC by elucidating RHEB and HRAS activation in response to extracellular stimuli. Furthermore, integration of Fluor-HPLC with syngeneic mouse models provided insights into KRAS activation dynamics in tumor tissues and evaluated the effectiveness of targeted therapeutics. Overall, this versatile method paves the way for investigating activation states and regulatory mechanisms of various small GTPases, potentially accelerating our understanding of their roles in cellular processes and disease pathogenesis.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"108545"},"PeriodicalIF":4.0,"publicationDate":"2025-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144005315","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

The Late-Stage Steps of Burkholderia cenocepacia Protein O-Linked Glycan Biosynthesis Are Conditionally Essential. 软心伯克氏菌蛋白o -连接聚糖生物合成的后期步骤是有条件的必要。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-04-24 DOI: 10.1016/j.jbc.2025.108515

Leila Jebeli, Taylor A McDaniels, Duncan T T Ho, Hamza Tahir, Nicholas L Kai-Ming, Molli Mcgaw, Kristian I Karlic, Jessica M Lewis, Nichollas E Scott

{"title":"The Late-Stage Steps of Burkholderia cenocepacia Protein O-Linked Glycan Biosynthesis Are Conditionally Essential.","authors":"Leila Jebeli, Taylor A McDaniels, Duncan T T Ho, Hamza Tahir, Nicholas L Kai-Ming, Molli Mcgaw, Kristian I Karlic, Jessica M Lewis, Nichollas E Scott","doi":"10.1016/j.jbc.2025.108515","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.108515","url":null,"abstract":"Periplasmic O-linked protein glycosylation is a highly conserved process observed across the Burkholderia genus. Within Burkholderia, protein glycosylation requires the five gene cluster known as the O-glycosylation cluster (OGC, ogcXABEI) which facilitates the construction of the O-linked trisaccharide attached to periplasmic proteins. Previous studies have reported conflicting results regarding the essentiality of ogcA, predicted to be responsible for the addition of the final carbohydrate of the O-linked trisaccharide and ogcX, the putative O-linked glycan flippase. Within this work, we aimed to dissect the impact of the loss of ogcA and ogcX on Burkholderia cenocepacia viability. We demonstrate that the loss of either ogcA or ogcX are detrimental if glycosylation is initiated leading to marked phenotypic effects. Proteomic analysis supports that the loss of ogcA/ogcX both blocks glycosylation and drives pleotropic effects in the membrane proteome, resulting in the loss of membrane integrity. Consistent with this, strains lacking ogcA and ogcX exhibit increased sensitivity to membrane stressors including antibiotics and demonstrate marked changes in membrane permeability. These effects are consistent with fouling of the undecaprenyl pool due to dead-end O-linked glycan intermediates, and consistent with this, we show that modulation of the undecaprenyl pool through the overexpression of undecaprenyl pyrophosphate synthase (UppS) or the OGC flippase (OgcX) restores viability while expression of early-stage OGC biosynthesis genes (ogcI and ogcB) reduce B. cenocepacia viability. These findings demonstrate disrupting O-linked glycan biosynthesis or transport appears to dramatically impact B. cenocepacia viability, supporting the assignment of ogcA and ogcX as conditionally essential.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"108515"},"PeriodicalIF":4.0,"publicationDate":"2025-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143993861","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Discovery and biochemical characterization of the D-aspartyl endopeptidase activity of the serine protease LACTB. 丝氨酸蛋白酶LACTB d -天冬氨酸内肽酶活性的发现及生化表征。

IF 4.8 2区生物学

Journal of Biological Chemistry Pub Date : 2025-04-24 DOI: 10.1016/j.jbc.2025.108549

Genta Ito,Naoko Utsunomiya-Tate

{"title":"Discovery and biochemical characterization of the D-aspartyl endopeptidase activity of the serine protease LACTB.","authors":"Genta Ito,Naoko Utsunomiya-Tate","doi":"10.1016/j.jbc.2025.108549","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.108549","url":null,"abstract":"Non-enzymatic D-isomerization of aspartic acid in proteins has been observed in lesions associated with age-related diseases, including cataracts and Alzheimer's disease. Given that D-isomerization of Asp disrupts the physiological conformation of proteins, it has been postulated that D-isomerization of Asp in proteins is a key factor in the pathogenesis of age-related diseases. D-Aspartyl endopeptidase (DAEP) activity, which cleaves proteins at the carboxy terminus of D-Asp and potentially induces degradation of abnormal proteins with D-isomerized Asp, has been observed in mitochondrial fractions of mammalian tissues. However, the specific proteins responsible for mammalian DAEP activity remain unknown. In this study, we identified mitochondrial serine β-lactamase-like protein (LACTB) as the first mammalian protein with DAEP activity by structural comparison with paenidase, a bacterial DAEP. LACTB exhibited DAEP activity similar to paenidase in an in vitro assay. In addition, LACTB cleaved a 10-residue peptide derived from amyloid β1-10 containing D-Asp at position 7, which was also observed with mammalian DAEP. LACTB has previously been characterized as a tumor suppressor and as a protein whose increased expression is associated with an increased risk of Alzheimer's disease. Therefore, our findings suggest that disruption of the proteostasis of D-Asp-containing proteins may underlie the pathogenesis of these diseases.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":"1 1","pages":"108549"},"PeriodicalIF":4.8,"publicationDate":"2025-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143885532","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Allosteric Communication Mechanism in the Glucagon Receptor. 胰高血糖素受体的变构通讯机制。

IF 4.8 2区生物学

Journal of Biological Chemistry Pub Date : 2025-04-23 DOI: 10.1016/j.jbc.2025.108530

Wijnand J C van der Velden,Elizaveta Mukhaleva,Nagarajan Vaidehi

{"title":"Allosteric Communication Mechanism in the Glucagon Receptor.","authors":"Wijnand J C van der Velden,Elizaveta Mukhaleva,Nagarajan Vaidehi","doi":"10.1016/j.jbc.2025.108530","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.108530","url":null,"abstract":"The glucagon receptor is dysregulated in metabolic disorders. Recent drug discovery has shown that agonists for the glucagon receptor might be more promising as therapeutics. Allosteric modulation may pave an alternative way to initiate responses that are required to target these metabolic disorders. Here, we investigated the allosteric communication mechanisms within the glucagon receptor using molecular dynamics simulations on five glucagon receptor states. Results highlighted that the extracellular domain is dynamic in the absence of an orthosteric agonist. In the presence of a partial agonist, we observed increased flexibility in the N-terminus of the receptor compared to the full agonist bound receptor. Class B GPCR microswitches showed repacking going from the inactive to the active state, allowing for G protein coupling. In the full agonist and G protein-bound state, Gαs showed both translational and rotational movement in the N-terminus, core and α5-helix, thereby forming key interactions between the core of the G protein and the glucagon receptor. Lastly, the allosteric communication from the extracellular region to the G protein coupling region of the glucagon receptor was the strongest in the intracellular negative allosteric modulator-bound state, the full agonist and G protein-bound state, and the full agonist-bound G protein-free state. The residue positions predicted to play a significant role in the allosteric communication mechanism showed overlap with disease associated mutations. Overall, our study provides insights into the allosteric communication mechanism in a class B GPCR which sets the foundation for future design of potential allosteric modulators targeting the glucagon receptor.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":"7 1","pages":"108530"},"PeriodicalIF":4.8,"publicationDate":"2025-04-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143885103","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

CRAC channel activity pulsates during cytosolic Ca2+ oscillations. CRAC通道活动脉动在胞质Ca2+振荡。

IF 4.8 2区生物学

Journal of Biological Chemistry Pub Date : 2025-04-23 DOI: 10.1016/j.jbc.2025.108519

Yu-Ping Lin,Erica Scappini,Gary Mirams,Charles J Tucker,Anant B Parekh

{"title":"CRAC channel activity pulsates during cytosolic Ca2+ oscillations.","authors":"Yu-Ping Lin,Erica Scappini,Gary Mirams,Charles J Tucker,Anant B Parekh","doi":"10.1016/j.jbc.2025.108519","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.108519","url":null,"abstract":"Intracellular Ca2+ ions are used as second messengers throughout the phylogenetic tree. They are indispensable for diverse biological processes ranging from fertilization to cell death. In Metazoans, signaling information is conveyed via the amplitude, frequency and spatial profile of cytosolic Ca2+ oscillations. In non-excitable cells, these oscillations generally arise from regenerative release of Ca2+ from inositol 1,4,5-trisphosphate (InsP3)-sensitive intracellular stores, which are refilled by entry of Ca2+ through Ca2+ release-activated Ca2+ (CRAC) channels in the plasma membrane. However, the precise contribution of these store-operated CRAC channels to Ca2+ oscillations has remained controversial for decades. One view proposes that CRAC channels remain open throughout stimulation, functioning as the pacemaker in setting Ca2+ oscillation frequency. An alternative hypothesis is that channel activity oscillates in parallel with InsP3-driven regenerative Ca2+ release. Here, by tethering a genetically encoded Ca2+ indicator to the pore- forming subunit of the CRAC channel, Orai1, we distinguish between these hypotheses and demonstrate that CRAC channel activity fluctuates in phase with cytosolic Ca2+ oscillations during physiological levels of stimulation. We also find that spatially distinct CRAC channel clusters fire in a coordinated manner, revealing that CRAC channels are not independent units but might function in a synchronized manner to provide pulses of Ca2+ signal at the same time.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":"17 1","pages":"108519"},"PeriodicalIF":4.8,"publicationDate":"2025-04-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143885106","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A cobalamin-dependent pathway of choline demethylation from the human gut acetogen Eubacterium limosum. 人类肠道醋酸真杆菌胆碱去甲基化的钴胺依赖途径。

IF 4.8 2区生物学

Journal of Biological Chemistry Pub Date : 2025-04-23 DOI: 10.1016/j.jbc.2025.108524

Ruisheng Jiang,Duncan J Kountz,Liwen Zhang,Joseph A Krzycki

{"title":"A cobalamin-dependent pathway of choline demethylation from the human gut acetogen Eubacterium limosum.","authors":"Ruisheng Jiang,Duncan J Kountz,Liwen Zhang,Joseph A Krzycki","doi":"10.1016/j.jbc.2025.108524","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.108524","url":null,"abstract":"Elevated serum levels of trimethylamine N-oxide (TMAO) are reported to promote the development of atherosclerosis. TMAO is produced by hepatic oxidation of trimethylamine (TMA) produced by the gut microbiome from dietary quaternary amines such as choline. Net TMA production in the gut depends on microbial enzymes that either produce or consume TMA and its precursors. Here we report the elucidation of a novel microbial pathway consuming choline without TMA production. The human gut acetogen Eubacterium limosum grows by demethylating choline to N-N-dimethylaminoethanol. Quantitative mass spectral analysis of the proteome revealed a multi-protein choline to tetrahydrofolate (THF) methyltransferase system present only in choline-grown cells. The components are encoded in a gene cluster on the genome and include MthB, an MttB superfamily member; MthC, homologous to methylotrophic cobalamin-binding proteins; MthA, homologous to cobalamin:THF methyltransferases; and MthK, a protein related to serine kinases. Together, MthB, MthC, and MthA methylate THF with phosphocholine, but not choline or other quaternary amines. MthB specifically methylates Co(I)-MthC with phosphocholine. MthK acts as a bifunctional choline kinase which can utilize ATP or the MthB demethylation product, N,N-dimethylaminoethanol phosphate, to phosphorylate choline. Together, MthK, MthB, MthC, and MthA are proposed to carry out the methylation of THF with choline. These results outline a THF methylation pathway in which choline is first activated with ATP to phosphocholine prior to demethylation to form N,N-dimethylaminoethanol phosphate. The latter can be recycled by MthK to form more phosphocholine without expending additional ATP, thus minimizing energy utilization during choline-dependent acetogenesis.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":"37 1","pages":"108524"},"PeriodicalIF":4.8,"publicationDate":"2025-04-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143885104","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0