The subcortical maternal complex (SCMC) is essential for mammalian preimplantation development, yet how SCMCcore (MATER/NLRP5, TLE6, FLOPED/OOEP) engages regulatory partners remains unclear. We determined cryo-EM structures of mouse SCMC bound to ZBED3 and human SCMC bound to NLRP2. Our structure reveals that ZBED3 interacts with all three SCMCcore subunits via its zinc finger domain, with conserved residue Phe73 mediating specific contacts. In contrast, human NLRP2 only binds to the WD40 domain of TLE6 through its leucine-rich repeat (LRR) domain. Similar interactions were also confirmed for NLRP7 with TLE6. These findings were cross-validated by in vivo proximity ligation and in vitro pull-down assays. Our work proposes a paradigmatic "Lego-like" assembly model, where the SCMCcore sequentially recruits different partners through diverse molecular interfaces. These findings provide critical structural insights into the SCMC's architecture and its multifaceted regulatory roles in early mammalian embryogenesis.
{"title":"Structural assembly of the subcortical maternal complex SCMC.","authors":"Guojin Ou,Qingting Liu,Haizhan Jiao,Zhuo Han,Jinhong Li,Ling Min,Pengliang Chi,Sibei Liu,Jialu Li,Qianqian Qi,Zihan Zhang,Li Guo,Xiang Wang,Lei Li,Jing Chen,Hongli Hu,Dong Deng","doi":"10.1016/j.str.2025.09.009","DOIUrl":"https://doi.org/10.1016/j.str.2025.09.009","url":null,"abstract":"The subcortical maternal complex (SCMC) is essential for mammalian preimplantation development, yet how SCMCcore (MATER/NLRP5, TLE6, FLOPED/OOEP) engages regulatory partners remains unclear. We determined cryo-EM structures of mouse SCMC bound to ZBED3 and human SCMC bound to NLRP2. Our structure reveals that ZBED3 interacts with all three SCMCcore subunits via its zinc finger domain, with conserved residue Phe73 mediating specific contacts. In contrast, human NLRP2 only binds to the WD40 domain of TLE6 through its leucine-rich repeat (LRR) domain. Similar interactions were also confirmed for NLRP7 with TLE6. These findings were cross-validated by in vivo proximity ligation and in vitro pull-down assays. Our work proposes a paradigmatic \"Lego-like\" assembly model, where the SCMCcore sequentially recruits different partners through diverse molecular interfaces. These findings provide critical structural insights into the SCMC's architecture and its multifaceted regulatory roles in early mammalian embryogenesis.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"108 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-10-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145338595","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}
Pub Date : 2025-10-16DOI: 10.1016/j.str.2025.09.008
Maria del Carmen Fernandez-Ramirez, Binh A. Nguyen, Shumaila Afrin, Virender Singh, Bret Evers, John M. Shelton, Christian Lopez Escobar, Parker Bassett, Lanie Wang, Maja Pękała, Yasmin Ahmed, Luis O. Cabrera Hernandez, Rose Pedretti, Preeti Singh, Jacob Canepa, Aleksandra Wosztyl, Yang Li, David R. Boyer, Qin Cao, Lorena Saelices
Transthyretin amyloidosis is a systemic protein misfolding disorder with diverse clinical phenotypes, including cardiomyopathy, polyneuropathy, or a combination of both. While structural polymorphism of amyloid fibrils has been linked to disease heterogeneity in neurodegenerative disorders, its role in transthyretin amyloidosis remains unclear. Here, we used cryo-electron microscopy to analyze ex vivo fibrils extracted from the hearts of three patients carrying the T60A mutation, a variant associated with mixed cardiac and neuropathic symptoms. In one patient, we additionally examined fibrils from the thyroid, kidney, and liver. All fibrils across patients and tissues adopted a single morphology previously associated with cardiomyopathy. Complementary molecular analyses revealed high compositional homogeneity. Notably, we extracted fibrils from the liver, an organ considered fibril-free, with seeding capacity in vitro. These findings suggest structural homogeneity as a hallmark of cardiac and mixed phenotypes, and provide a mechanistic rationale for the transmission of amyloidosis following domino liver transplantation.
{"title":"Structural and molecular homogeneity of ATTRv-T60A amyloid fibrils across patients and organs","authors":"Maria del Carmen Fernandez-Ramirez, Binh A. Nguyen, Shumaila Afrin, Virender Singh, Bret Evers, John M. Shelton, Christian Lopez Escobar, Parker Bassett, Lanie Wang, Maja Pękała, Yasmin Ahmed, Luis O. Cabrera Hernandez, Rose Pedretti, Preeti Singh, Jacob Canepa, Aleksandra Wosztyl, Yang Li, David R. Boyer, Qin Cao, Lorena Saelices","doi":"10.1016/j.str.2025.09.008","DOIUrl":"https://doi.org/10.1016/j.str.2025.09.008","url":null,"abstract":"Transthyretin amyloidosis is a systemic protein misfolding disorder with diverse clinical phenotypes, including cardiomyopathy, polyneuropathy, or a combination of both. While structural polymorphism of amyloid fibrils has been linked to disease heterogeneity in neurodegenerative disorders, its role in transthyretin amyloidosis remains unclear. Here, we used cryo-electron microscopy to analyze <em>ex vivo</em> fibrils extracted from the hearts of three patients carrying the T60A mutation, a variant associated with mixed cardiac and neuropathic symptoms. In one patient, we additionally examined fibrils from the thyroid, kidney, and liver. All fibrils across patients and tissues adopted a single morphology previously associated with cardiomyopathy. Complementary molecular analyses revealed high compositional homogeneity. Notably, we extracted fibrils from the liver, an organ considered fibril-free, with seeding capacity <em>in vitro</em>. These findings suggest structural homogeneity as a hallmark of cardiac and mixed phenotypes, and provide a mechanistic rationale for the transmission of amyloidosis following domino liver transplantation.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"9 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-10-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145295468","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}
Pub Date : 2025-10-09DOI: 10.1016/j.str.2025.09.007
William S. Henriques, Jarrett Bowman, Laina N. Hall, Colin C. Gauvin, Hui Wei, Huihui Kuang, Christina M. Zimanyi, Edward T. Eng, Andrew Santiago-Frangos, Blake Wiedenheft
Cas1 and Cas2 are the hallmark proteins of prokaryotic adaptive immunity. However, these two proteins are often fused to other proteins and the functional association of these fusions often remain poorly understood. Here we purify and determine structures of Cas1 and the Cas2/3 fusion proteins from Pseudomonas aeruginosa at distinct stages of CRISPR adaptation. Collectively, these structures reveal a prominent, positively charged channel on one face of the integration complex that captures short fragments of foreign DNA. Foreign DNA binding triggers conformational changes in Cas2/3 that expose new DNA binding surfaces necessary for homing the DNA-bound integrase to specific CRISPR loci. The length of the foreign DNA substrate determines if Cas1-2/3 docks completely onto the CRISPR repeat to successfully catalyze two sequential transesterification reactions required for integration. Together, these structures clarify how the Cas1-2/3 proteins orchestrate foreign DNA capture, site-specific delivery, and integration of new DNA into the bacterial genome.
{"title":"Structures reveal how the Cas1-2/3 integrase captures, delivers, and integrates foreign DNA into CRISPR loci","authors":"William S. Henriques, Jarrett Bowman, Laina N. Hall, Colin C. Gauvin, Hui Wei, Huihui Kuang, Christina M. Zimanyi, Edward T. Eng, Andrew Santiago-Frangos, Blake Wiedenheft","doi":"10.1016/j.str.2025.09.007","DOIUrl":"https://doi.org/10.1016/j.str.2025.09.007","url":null,"abstract":"Cas1 and Cas2 are the hallmark proteins of prokaryotic adaptive immunity. However, these two proteins are often fused to other proteins and the functional association of these fusions often remain poorly understood. Here we purify and determine structures of Cas1 and the Cas2/3 fusion proteins from <em>Pseudomonas aeruginosa</em> at distinct stages of CRISPR adaptation. Collectively, these structures reveal a prominent, positively charged channel on one face of the integration complex that captures short fragments of foreign DNA. Foreign DNA binding triggers conformational changes in Cas2/3 that expose new DNA binding surfaces necessary for homing the DNA-bound integrase to specific CRISPR loci. The length of the foreign DNA substrate determines if Cas1-2/3 docks completely onto the CRISPR repeat to successfully catalyze two sequential transesterification reactions required for integration. Together, these structures clarify how the Cas1-2/3 proteins orchestrate foreign DNA capture, site-specific delivery, and integration of new DNA into the bacterial genome.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"16 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-10-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145247581","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}
Pub Date : 2025-10-03DOI: 10.1016/j.str.2025.09.006
Evan J. McMahon, Alexander G. Cioffi, Patrick R. Visperas, Yueqing Lin, Michael Shaghafi, Courtney M. Daczkowski, Johannes C. Hermann, Robert A. Everley, Richard M. Neve, Daniel A. Erlanson, Kevin R. Webster, Vikram Narayan, Weiru Wang
Targeted protein degradation (TPD) leverages the ubiquitin-proteasome system to eliminate disease-causing proteins via E3 ligases. To date, the field is limited to utilizing a few of the over 600 human E3 ligases. To expand this repertoire, we conducted structural and functional validation of DDB1 (Damage-specific DNA binding protein 1) and Cullin-associated factor (DCAF)2 (DTL/CDT2), a Cullin4-RING ligase substrate adaptor implicated in DNA damage response and cancer, as a novel E3 for TPD. Cryoelectron microscopy (cryo-EM) structures of the DCAF2:DDB1:DDA1 complex (3.3 Å), a ligand bound complex (3.1 Å), and a ternary complex with a covalent proteolysis-targeting chimera (PROTAC) and BRD4 (3.4 Å) reveal PROTAC-mediated substrate recruitment. Using covalent bifunctional tool compounds engaging residue C141 in the WD40 domain, we demonstrate robust ubiquitination in biochemical assays and cellular TPD using the COFFEE (covalent functionalization followed by E3 electroporation) method. These findings position DCAF2 as a promising E3 adaptor for PROTAC strategies and identify C141 as a relevant site for future PROTAC discovery.
{"title":"Structural basis for DCAF2 as a novel E3 ligase for PROTAC-mediated targeted protein degradation","authors":"Evan J. McMahon, Alexander G. Cioffi, Patrick R. Visperas, Yueqing Lin, Michael Shaghafi, Courtney M. Daczkowski, Johannes C. Hermann, Robert A. Everley, Richard M. Neve, Daniel A. Erlanson, Kevin R. Webster, Vikram Narayan, Weiru Wang","doi":"10.1016/j.str.2025.09.006","DOIUrl":"https://doi.org/10.1016/j.str.2025.09.006","url":null,"abstract":"Targeted protein degradation (TPD) leverages the ubiquitin-proteasome system to eliminate disease-causing proteins via E3 ligases. To date, the field is limited to utilizing a few of the over 600 human E3 ligases. To expand this repertoire, we conducted structural and functional validation of DDB1 (Damage-specific DNA binding protein 1) and Cullin-associated factor (DCAF)2 (DTL/CDT2), a Cullin4-RING ligase substrate adaptor implicated in DNA damage response and cancer, as a novel E3 for TPD. Cryoelectron microscopy (cryo-EM) structures of the DCAF2:DDB1:DDA1 complex (3.3 Å), a ligand bound complex (3.1 Å), and a ternary complex with a covalent proteolysis-targeting chimera (PROTAC) and BRD4 (3.4 Å) reveal PROTAC-mediated substrate recruitment. Using covalent bifunctional tool compounds engaging residue C141 in the WD40 domain, we demonstrate robust ubiquitination in biochemical assays and cellular TPD using the COFFEE (covalent functionalization followed by E3 electroporation) method. These findings position DCAF2 as a promising E3 adaptor for PROTAC strategies and identify C141 as a relevant site for future PROTAC discovery.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"52 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-10-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145209661","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}
Pub Date : 2025-10-02DOI: 10.1016/j.str.2025.09.005
S. M. Naimul Hasan, Elnaz Khalili Samani, Alexander F.A. Keszei, Mahtab Heydari, Mohammad T. Mazhab-Jafari
Mycobiota fatty acid synthases (FASs) catalyze iterative cycles of condensation, dehydration, and reduction to produce saturated fatty acids. Although these multienzymes are attractive antifungal drug targets, no clinically approved small-molecule inhibitors exist, and the regulation of de novo fatty acid synthesis remains poorly understood. Here, we identify an allosteric regulation of the FAS ketoacyl reduction reaction by palmitoyl-CoA. The palmitate moiety binds a distal site on the central wheel of fungal FAS from Saccharomyces cerevisiae and Candida albicans. This site also accommodates shorter acyl chains, but only palmitoyl-CoA suppresses ketoacyl reductase (KR) activity. While no major conformational changes occur in the reductase domain, palmitoyl-CoA binding quenches dynamics in the central disk, improving local resolution and stabilizing structured water molecules. This entropic effect underlies allosteric communication to the reductase site. Our findings uncover a regulatory mechanism of fungal FAS exploitable for antifungal drug design.
{"title":"Allosteric regulation of fungal fatty acid synthesis","authors":"S. M. Naimul Hasan, Elnaz Khalili Samani, Alexander F.A. Keszei, Mahtab Heydari, Mohammad T. Mazhab-Jafari","doi":"10.1016/j.str.2025.09.005","DOIUrl":"https://doi.org/10.1016/j.str.2025.09.005","url":null,"abstract":"Mycobiota fatty acid synthases (FASs) catalyze iterative cycles of condensation, dehydration, and reduction to produce saturated fatty acids. Although these multienzymes are attractive antifungal drug targets, no clinically approved small-molecule inhibitors exist, and the regulation of <em>de novo</em> fatty acid synthesis remains poorly understood. Here, we identify an allosteric regulation of the FAS ketoacyl reduction reaction by palmitoyl-CoA. The palmitate moiety binds a distal site on the central wheel of fungal FAS from <em>Saccharomyces cerevisiae</em> and <em>Candida albicans</em>. This site also accommodates shorter acyl chains, but only palmitoyl-CoA suppresses ketoacyl reductase (KR) activity. While no major conformational changes occur in the reductase domain, palmitoyl-CoA binding quenches dynamics in the central disk, improving local resolution and stabilizing structured water molecules. This entropic effect underlies allosteric communication to the reductase site. Our findings uncover a regulatory mechanism of fungal FAS exploitable for antifungal drug design.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"32 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145204016","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}
In this issue of Structure, Connor et al.1 combine topology and residue-level energetics of α-synuclein and tau fibrils to identify isoenergetic fold families. They reveal how conserved hotspot residues, mutations, and, potentially, environmental cues steer polymorph selection, providing a unified framework for comparing amyloid strains and classifying emergent structures.
{"title":"Toward a thermodynamic taxonomy of amyloid fibrils","authors":"Nikolaos Louros, Joost Schymkowitz, Frederic Rousseau","doi":"10.1016/j.str.2025.08.010","DOIUrl":"https://doi.org/10.1016/j.str.2025.08.010","url":null,"abstract":"In this issue of <em>Structure</em>, Connor et al.<span><span><sup>1</sup></span></span> combine topology and residue-level energetics of α-synuclein and tau fibrils to identify isoenergetic fold families. They reveal how conserved hotspot residues, mutations, and, potentially, environmental cues steer polymorph selection, providing a unified framework for comparing amyloid strains and classifying emergent structures.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"76 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145204018","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}
Pub Date : 2025-10-02DOI: 10.1016/j.str.2025.08.014
Fiyaz Mohammed, Carrie R. Willcox, Benjamin E. Willcox
In this issue of Structure, Rashleigh et al.1 used cryoelectron microscopy to assess recognition of the model foreign antigen phycoerythrin (PE) by a γδ TCR, an αβ TCR, and an antibody. With potential relevance for anti-pathogen immunity, this work demonstrates CDR3-dominated contacts to a shared hydrophobic PE epitope, emphasizing similarities between γδ TCR and antibody recognition.
{"title":"A triple take on antigen receptor recognition of PE","authors":"Fiyaz Mohammed, Carrie R. Willcox, Benjamin E. Willcox","doi":"10.1016/j.str.2025.08.014","DOIUrl":"https://doi.org/10.1016/j.str.2025.08.014","url":null,"abstract":"In this issue of <em>Structure</em>, Rashleigh et al.<span><span><sup>1</sup></span></span> used cryoelectron microscopy to assess recognition of the model foreign antigen phycoerythrin (PE) by a γδ TCR, an αβ TCR, and an antibody. With potential relevance for anti-pathogen immunity, this work demonstrates CDR3-dominated contacts to a shared hydrophobic PE epitope, emphasizing similarities between γδ TCR and antibody recognition.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"53 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-10-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145204017","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}
Pub Date : 2025-09-30DOI: 10.1016/j.str.2025.09.004
Eva M. Huber, Wolfgang Heinemeyer, Michael Groll
Single point mutations in proteasome subunits can cause severe autoinflammatory syndromes. By still largely unknown mechanisms, some of these disease-associated mutations impair normal proteasome function and induce the production of pro-inflammatory cytokines, thereby leading to systemic inflammations. In order to obtain more insights on why and how the mutations T3M and G128V in the immunoproteasome subunit β5i trigger such deleterious effects, we created the respective yeast mutants and characterized their phenotypes with special emphasis on proteasome structure and activity. X-ray crystallographic data revealed that the mutation T3M influences structure and flexibility of the proteasomal substrate-binding channel with moderate impairment of proteasome biogenesis, whereas the amino acid substitution G128V causes larger structural rearrangements that severely disturb particle assembly and maturation. The obtained results provide a deeper understanding of how single point mutations can affect proteasome subunit structure as well as particle biogenesis and ultimately cause chronic inflammatory diseases.
{"title":"Proteasome-associated autoinflammatory syndromes: The impact of mutations in proteasome subunits on particle assembly, structure, and activity","authors":"Eva M. Huber, Wolfgang Heinemeyer, Michael Groll","doi":"10.1016/j.str.2025.09.004","DOIUrl":"https://doi.org/10.1016/j.str.2025.09.004","url":null,"abstract":"Single point mutations in proteasome subunits can cause severe autoinflammatory syndromes. By still largely unknown mechanisms, some of these disease-associated mutations impair normal proteasome function and induce the production of pro-inflammatory cytokines, thereby leading to systemic inflammations. In order to obtain more insights on why and how the mutations T3M and G128V in the immunoproteasome subunit β5i trigger such deleterious effects, we created the respective yeast mutants and characterized their phenotypes with special emphasis on proteasome structure and activity. X-ray crystallographic data revealed that the mutation T3M influences structure and flexibility of the proteasomal substrate-binding channel with moderate impairment of proteasome biogenesis, whereas the amino acid substitution G128V causes larger structural rearrangements that severely disturb particle assembly and maturation. The obtained results provide a deeper understanding of how single point mutations can affect proteasome subunit structure as well as particle biogenesis and ultimately cause chronic inflammatory diseases.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"100 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-09-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145189270","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}
Pub Date : 2025-09-29DOI: 10.1016/j.str.2025.09.003
Chenyao Li, Rouslan G. Efremov
Ryanodine receptors (RyRs) are intracellular tetrameric ion channels responsible for Ca2+ release from the sarcoplasmic and endoplasmic reticulum. Ryanodine receptor 1 (RyR1) isoform, critical for muscle contraction, has been studied most extensively. While cryoelectron microscopy (cryo-EM) has been instrumental in revealing near-atomic details of RyR gating mechanisms, the open probability of RyR1 under cryo-EM conditions is notably lower than that observed in electrophysiological studies. Here, we present a cryo-EM study examining the open probability of RyR1 solubilized in CHAPS with varying lipid concentrations. We found that increasing lipid concentration from 0.001% to 0.05% raised the RyR1 open probability from 16% to 84%, whereas RyR1 reconstituted into lipid nanodiscs remained closed. We modeled 72 lipid molecules in the map reconstructed at the highest lipid concentration. These findings demonstrate the important role of lipids in modulating the open fraction of solubilized RyR1 channels under cryo-EM conditions and suggest optimal lipid mimetics for structural studies of RyR1 gating.
{"title":"Lipids modulate the open probability of RyR1 under cryo-EM conditions","authors":"Chenyao Li, Rouslan G. Efremov","doi":"10.1016/j.str.2025.09.003","DOIUrl":"https://doi.org/10.1016/j.str.2025.09.003","url":null,"abstract":"Ryanodine receptors (RyRs) are intracellular tetrameric ion channels responsible for Ca<sup>2+</sup> release from the sarcoplasmic and endoplasmic reticulum. Ryanodine receptor 1 (RyR1) isoform, critical for muscle contraction, has been studied most extensively. While cryoelectron microscopy (cryo-EM) has been instrumental in revealing near-atomic details of RyR gating mechanisms, the open probability of RyR1 under cryo-EM conditions is notably lower than that observed in electrophysiological studies. Here, we present a cryo-EM study examining the open probability of RyR1 solubilized in CHAPS with varying lipid concentrations. We found that increasing lipid concentration from 0.001% to 0.05% raised the RyR1 open probability from 16% to 84%, whereas RyR1 reconstituted into lipid nanodiscs remained closed. We modeled 72 lipid molecules in the map reconstructed at the highest lipid concentration. These findings demonstrate the important role of lipids in modulating the open fraction of solubilized RyR1 channels under cryo-EM conditions and suggest optimal lipid mimetics for structural studies of RyR1 gating.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"67 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-09-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145183026","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}
Interleukin-31 (IL-31) signals through the IL-31 receptor alpha (IL-31RA) and oncostatin M receptor beta (OSMRβ) heterodimer, mediating pruritus, dermatitis, inflammatory responses, neuroimmune interactions, and certain cancers. Here, we present the crystal structure of canine IL-31 (cIL-31) in complex with a neutralizing caninized monoclonal antibody (2D10-2). This antibody competitively inhibited cIL-31 binding to canine OSMRβ (cOSMRβ) but not to canine IL-31RA (cIL-31RA). Moreover, it effectively blocked cIL-31-induced STAT5 phosphorylation in vitro and alleviated cIL-31-induced pruritus in beagle dogs. Structural analysis identified key antibody-binding residues in α-helical A, α-helical D, and the AB loop of cIL-31. Systematic mutagenesis based on the complex structure further defined the conformational epitopes of cIL-31 recognized by cOSMRβ. In summary, this study reports the IL-31 structure, revealing a four-α-helical bundle cytokine, and elucidates 2D10-2’s neutralizing mechanism by targeting the cIL-31-cOSMRβ interaction. These findings advance our understanding of IL-31 and offer insights for developing IL-31-targeted therapeutics.
{"title":"Structural insights into IL-31 signaling inhibition by a neutralizing antibody","authors":"Tianling Guo, Yuxin Zheng, Zheng Fan, Ping Liu, Yan Chai, Xiaoping Liao, Caili Zhang, Xuefei Pang, Delin Li, Feng Gao, Haixia Xiao","doi":"10.1016/j.str.2025.09.002","DOIUrl":"https://doi.org/10.1016/j.str.2025.09.002","url":null,"abstract":"Interleukin-31 (IL-31) signals through the IL-31 receptor alpha (IL-31RA) and oncostatin M receptor beta (OSMRβ) heterodimer, mediating pruritus, dermatitis, inflammatory responses, neuroimmune interactions, and certain cancers. Here, we present the crystal structure of canine IL-31 (cIL-31) in complex with a neutralizing caninized monoclonal antibody (2D10-2). This antibody competitively inhibited cIL-31 binding to canine OSMRβ (cOSMRβ) but not to canine IL-31RA (cIL-31RA). Moreover, it effectively blocked cIL-31-induced STAT5 phosphorylation <em>in vitro</em> and alleviated cIL-31-induced pruritus in beagle dogs. Structural analysis identified key antibody-binding residues in α-helical A, α-helical D, and the AB loop of cIL-31. Systematic mutagenesis based on the complex structure further defined the conformational epitopes of cIL-31 recognized by cOSMRβ. In summary, this study reports the IL-31 structure, revealing a four-α-helical bundle cytokine, and elucidates 2D10-2’s neutralizing mechanism by targeting the cIL-31-cOSMRβ interaction. These findings advance our understanding of IL-31 and offer insights for developing IL-31-targeted therapeutics.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"17 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-09-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145140951","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}
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