Pub Date : 2025-11-10DOI: 10.1038/s41594-025-01710-6
Zhaoxiang He, Fei Teng, Yanqing Yang, Ruiyang Guo, Mengchen Wu, Fangzhu Han, Hongtao Tian, Jiawei Wang, Yiqi Hu, Yangwei Jiang, Leili Zhang, Chenyang Xu, Fan Yang, Jiancang Zhou, Shan Zhang, James A. Letts, Ruhong Zhou, Long Zhou
Metformin is the only antihyperglycemic biguanide targeting type 2 diabetes mellitus with proven safety. Although a mechanism of action involving tight inhibition of the respiratory complex I has been proposed for hydrophobic biguanides, it remains elusive for the hydrophilic metformin, whose excellent pharmacological tolerance depends on weak complex I inhibition without competitive nature. Here we solved cryo-electron microscopy structures of the metformin-bound porcine respirasome. Our structural and kinetic data are consistent with a model in which metformin enters complex I only in its open state and becomes trapped at the ubiquinone redox site by ubiquinone-induced conformational closing of the enzyme. By contrast, the hydrophobic proguanil alone occupies both the entrance and the redox site of the ubiquinone channel in open and closed complex I and is kinetically consistent with competitive inhibition with conformation-dependent affinities. Our data provide the molecular basis for metformin’s well-known superior properties, such as a wide therapeutic window and positive ubiquinone cooperativity, leading to its clinical success and facilitating future therapeutic developments. He, Teng and Yang et al. report how metformin, the widely used antidiabetic drug, inhibit its target, the respiratory complex I, through a distinct state-dependent, inhibitor trapping mechanism, thus providing the molecular basis for its superior clinical tolerance.
{"title":"Hydrophilic metformin and hydrophobic biguanides inhibit mitochondrial complex I by distinct mechanisms","authors":"Zhaoxiang He, Fei Teng, Yanqing Yang, Ruiyang Guo, Mengchen Wu, Fangzhu Han, Hongtao Tian, Jiawei Wang, Yiqi Hu, Yangwei Jiang, Leili Zhang, Chenyang Xu, Fan Yang, Jiancang Zhou, Shan Zhang, James A. Letts, Ruhong Zhou, Long Zhou","doi":"10.1038/s41594-025-01710-6","DOIUrl":"10.1038/s41594-025-01710-6","url":null,"abstract":"Metformin is the only antihyperglycemic biguanide targeting type 2 diabetes mellitus with proven safety. Although a mechanism of action involving tight inhibition of the respiratory complex I has been proposed for hydrophobic biguanides, it remains elusive for the hydrophilic metformin, whose excellent pharmacological tolerance depends on weak complex I inhibition without competitive nature. Here we solved cryo-electron microscopy structures of the metformin-bound porcine respirasome. Our structural and kinetic data are consistent with a model in which metformin enters complex I only in its open state and becomes trapped at the ubiquinone redox site by ubiquinone-induced conformational closing of the enzyme. By contrast, the hydrophobic proguanil alone occupies both the entrance and the redox site of the ubiquinone channel in open and closed complex I and is kinetically consistent with competitive inhibition with conformation-dependent affinities. Our data provide the molecular basis for metformin’s well-known superior properties, such as a wide therapeutic window and positive ubiquinone cooperativity, leading to its clinical success and facilitating future therapeutic developments. He, Teng and Yang et al. report how metformin, the widely used antidiabetic drug, inhibit its target, the respiratory complex I, through a distinct state-dependent, inhibitor trapping mechanism, thus providing the molecular basis for its superior clinical tolerance.","PeriodicalId":49141,"journal":{"name":"Nature Structural & Molecular Biology","volume":"33 1","pages":"100-111"},"PeriodicalIF":10.1,"publicationDate":"2025-11-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145478131","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-06DOI: 10.1038/s41594-025-01709-z
mTORC1 promotes cell growth by sensing nutrients and driving anabolic processes. When nutrients are scarce, GATOR1 turns off growth signals. Our study describes how KICSTOR links GATOR1 to lysosomes to enable its function, a finding that may help us to understand certain neurological disorders.
{"title":"Caught in the act: how a brake on cell growth is anchored to lysosomes","authors":"","doi":"10.1038/s41594-025-01709-z","DOIUrl":"10.1038/s41594-025-01709-z","url":null,"abstract":"mTORC1 promotes cell growth by sensing nutrients and driving anabolic processes. When nutrients are scarce, GATOR1 turns off growth signals. Our study describes how KICSTOR links GATOR1 to lysosomes to enable its function, a finding that may help us to understand certain neurological disorders.","PeriodicalId":49141,"journal":{"name":"Nature Structural & Molecular Biology","volume":"32 12","pages":"2383-2384"},"PeriodicalIF":10.1,"publicationDate":"2025-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145447669","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The human KICSTOR complex, comprising KPTN, ITFG2, C12orf66 and the scaffolding protein SZT2, anchors the mTORC1 inhibitor GATOR1 to lysosomes. Mutations affecting KICSTOR subunits are associated with severe neurodevelopmental and epileptic disorders. Loss of KICSTOR mimics GATOR1 inactivation, resulting in constitutive mTORC1 activation, highlighting its critical role in nutrient sensing. Here, we used cryo-electron microscopy and computational modeling to determine the architectures of KICSTOR and the GATOR1–KICSTOR supercomplex. We show that SZT2 forms a crescent-shaped scaffold with repetitive tandem units, binding the ITFG2–KPTN heterodimer and C12orf66 at its C terminus. Structural and biochemical analyses revealed that GATOR1 binds the SZT2 N-terminal domain through NPRL3; disruption of this interaction hyperactivates mTORC1 and mislocalizes TFE3 independently of nutrient status. We further demonstrate the membrane-binding ability of KICSTOR, with SZT2 and C12orf66 preferentially interacting with negatively charged lipids—a requirement for lysosomal localization. These findings identify how KICSTOR positions GATOR1 on lysosomes to regulate nutrient-dependent mTORC1 signaling. Teng and Zeng et al. use cryo-electron microscopy to show that the crescent scaffold of KICSTOR anchors GATOR1 to lysosomes and disruption of the interaction causes mTORC1 hyperactivation and TFE3 mislocalization. KICSTOR enables nutrient-dependent mTORC1 regulation by binding anionic lipids for lysosomal targeting.
{"title":"Architecture of the human KICSTOR and GATOR1–KICSTOR complexes","authors":"Fei Teng, Huan Zeng, Xinyi Mai, Shujun Chen, Lulu Wang, Zeming Feng, Shuyun Tian, Shan Wang, Goran Stjepanovic, Chun-Yan Lim, Ming-Yuan Su","doi":"10.1038/s41594-025-01693-4","DOIUrl":"10.1038/s41594-025-01693-4","url":null,"abstract":"The human KICSTOR complex, comprising KPTN, ITFG2, C12orf66 and the scaffolding protein SZT2, anchors the mTORC1 inhibitor GATOR1 to lysosomes. Mutations affecting KICSTOR subunits are associated with severe neurodevelopmental and epileptic disorders. Loss of KICSTOR mimics GATOR1 inactivation, resulting in constitutive mTORC1 activation, highlighting its critical role in nutrient sensing. Here, we used cryo-electron microscopy and computational modeling to determine the architectures of KICSTOR and the GATOR1–KICSTOR supercomplex. We show that SZT2 forms a crescent-shaped scaffold with repetitive tandem units, binding the ITFG2–KPTN heterodimer and C12orf66 at its C terminus. Structural and biochemical analyses revealed that GATOR1 binds the SZT2 N-terminal domain through NPRL3; disruption of this interaction hyperactivates mTORC1 and mislocalizes TFE3 independently of nutrient status. We further demonstrate the membrane-binding ability of KICSTOR, with SZT2 and C12orf66 preferentially interacting with negatively charged lipids—a requirement for lysosomal localization. These findings identify how KICSTOR positions GATOR1 on lysosomes to regulate nutrient-dependent mTORC1 signaling. Teng and Zeng et al. use cryo-electron microscopy to show that the crescent scaffold of KICSTOR anchors GATOR1 to lysosomes and disruption of the interaction causes mTORC1 hyperactivation and TFE3 mislocalization. KICSTOR enables nutrient-dependent mTORC1 regulation by binding anionic lipids for lysosomal targeting.","PeriodicalId":49141,"journal":{"name":"Nature Structural & Molecular Biology","volume":"32 12","pages":"2587-2600"},"PeriodicalIF":10.1,"publicationDate":"2025-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145447663","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-06DOI: 10.1038/s41594-025-01695-2
Connor Arkinson, Christine L. Gee, Zeyuan Zhang, Ken C. Dong, Andreas Martin
The 26S proteasome targets many cellular proteins for degradation during homeostasis and quality control. Proteasome-interacting cofactors modulate these functions and aid in substrate degradation. Here we solve high-resolution structures of the redox active cofactor TXNL1 bound to the human 26S proteasome at saturating and substoichiometric concentrations by time-resolved cryo-electron microscopy (cryo-EM). We identify distinct binding modes of TXNL1 that depend on the proteasome conformation and ATPase motor states. Together with biophysical and biochemical experiments, we show that the resting-state proteasome binds TXNL1 with low affinity and in variable positions on top of the Rpn11 deubiquitinase. In contrast, in the actively degrading proteasome, TXNL1 uses additional interactions for high-affinity binding, whereby its C-terminal tail covers the catalytic groove of Rpn11 and coordinates the active-site Zn2+. Furthermore, these cryo-EM structures of the degrading proteasome capture the ATPase hexamer in several spiral-staircase arrangements that indicate temporally asymmetric hydrolysis and conformational changes in bursts during mechanical substrate unfolding and translocation. Remarkably, we catch the proteasome in the act of unfolding the β-barrel mEos3.2 substrate while the ATPase hexamer is in a particular staircase register. Our findings advance current models for protein translocation through hexameric AAA+ motors and reveal how the proteasome uses its distinct conformational states to coordinate cofactor binding and substrate processing. The authors use time-resolved cryo-electron microscopy to reveal the interactions of the redox-active cofactor TXNL1 with the human 26S proteasome and detect ATPase motor states that indicate burst-like mechanisms for hand-over-hand substrate translocation.
{"title":"Structural landscape of the degrading 26S proteasome reveals conformation-specific binding of TXNL1","authors":"Connor Arkinson, Christine L. Gee, Zeyuan Zhang, Ken C. Dong, Andreas Martin","doi":"10.1038/s41594-025-01695-2","DOIUrl":"10.1038/s41594-025-01695-2","url":null,"abstract":"The 26S proteasome targets many cellular proteins for degradation during homeostasis and quality control. Proteasome-interacting cofactors modulate these functions and aid in substrate degradation. Here we solve high-resolution structures of the redox active cofactor TXNL1 bound to the human 26S proteasome at saturating and substoichiometric concentrations by time-resolved cryo-electron microscopy (cryo-EM). We identify distinct binding modes of TXNL1 that depend on the proteasome conformation and ATPase motor states. Together with biophysical and biochemical experiments, we show that the resting-state proteasome binds TXNL1 with low affinity and in variable positions on top of the Rpn11 deubiquitinase. In contrast, in the actively degrading proteasome, TXNL1 uses additional interactions for high-affinity binding, whereby its C-terminal tail covers the catalytic groove of Rpn11 and coordinates the active-site Zn2+. Furthermore, these cryo-EM structures of the degrading proteasome capture the ATPase hexamer in several spiral-staircase arrangements that indicate temporally asymmetric hydrolysis and conformational changes in bursts during mechanical substrate unfolding and translocation. Remarkably, we catch the proteasome in the act of unfolding the β-barrel mEos3.2 substrate while the ATPase hexamer is in a particular staircase register. Our findings advance current models for protein translocation through hexameric AAA+ motors and reveal how the proteasome uses its distinct conformational states to coordinate cofactor binding and substrate processing. The authors use time-resolved cryo-electron microscopy to reveal the interactions of the redox-active cofactor TXNL1 with the human 26S proteasome and detect ATPase motor states that indicate burst-like mechanisms for hand-over-hand substrate translocation.","PeriodicalId":49141,"journal":{"name":"Nature Structural & Molecular Biology","volume":"32 12","pages":"2403-2415"},"PeriodicalIF":10.1,"publicationDate":"2025-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.nature.comhttps://www.nature.com/articles/s41594-025-01695-2.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145447664","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-05DOI: 10.1038/s41594-025-01699-y
Kejia Li, Clement M. Potel, Isabelle Becher, Nico Hüttmann, Martin Garrido-Rodriguez, Jennifer Schwarz, Mira Lea Burtscher, Mikhail M. Savitski
Systematic mapping of protein–ligand interactions is essential for understanding biological processes and drug mechanisms. Peptide-centric local stability assay (PELSA) is a powerful tool for detecting these interactions and identifying potential binding sites. However, its original workflow is limited in throughput, sample compatibility and accessible protein targets. Here, we introduce a high-throughput adaptation—HT-PELSA—that increases sample processing efficiency by 100-fold while maintaining high sensitivity and reproducibility. HT-PELSA substantially extends the capabilities of the original method by enabling sensitive protein–ligand profiling in crude cell, tissue and bacterial lysates, allowing the identification of membrane protein targets in diverse biological systems. We demonstrate that HT-PELSA can precisely and accurately determine binding affinities of small molecule inhibitors, sensitively detect direct and allosteric ATP binding sites, and reveal off-target interactions of a marketed kinase inhibitor in heart tissue. By enhancing scalability, reducing costs and enabling system-wide drug screening across a wide range of sample types, HT-PELSA—when combined with next-generation mass spectrometry—may offer a powerful platform poised to accelerate both drug discovery and basic biological research. Li et al. further develop a high-throughput peptide-centric local stability assay that speeds up sample preparation 100-fold and extends protein–ligand identification to membrane proteins, tissues and bacteria.
{"title":"High-throughput peptide-centric local stability assay extends protein–ligand identification to membrane proteins, tissues and bacteria","authors":"Kejia Li, Clement M. Potel, Isabelle Becher, Nico Hüttmann, Martin Garrido-Rodriguez, Jennifer Schwarz, Mira Lea Burtscher, Mikhail M. Savitski","doi":"10.1038/s41594-025-01699-y","DOIUrl":"10.1038/s41594-025-01699-y","url":null,"abstract":"Systematic mapping of protein–ligand interactions is essential for understanding biological processes and drug mechanisms. Peptide-centric local stability assay (PELSA) is a powerful tool for detecting these interactions and identifying potential binding sites. However, its original workflow is limited in throughput, sample compatibility and accessible protein targets. Here, we introduce a high-throughput adaptation—HT-PELSA—that increases sample processing efficiency by 100-fold while maintaining high sensitivity and reproducibility. HT-PELSA substantially extends the capabilities of the original method by enabling sensitive protein–ligand profiling in crude cell, tissue and bacterial lysates, allowing the identification of membrane protein targets in diverse biological systems. We demonstrate that HT-PELSA can precisely and accurately determine binding affinities of small molecule inhibitors, sensitively detect direct and allosteric ATP binding sites, and reveal off-target interactions of a marketed kinase inhibitor in heart tissue. By enhancing scalability, reducing costs and enabling system-wide drug screening across a wide range of sample types, HT-PELSA—when combined with next-generation mass spectrometry—may offer a powerful platform poised to accelerate both drug discovery and basic biological research. Li et al. further develop a high-throughput peptide-centric local stability assay that speeds up sample preparation 100-fold and extends protein–ligand identification to membrane proteins, tissues and bacteria.","PeriodicalId":49141,"journal":{"name":"Nature Structural & Molecular Biology","volume":"33 1","pages":"184-192"},"PeriodicalIF":10.1,"publicationDate":"2025-11-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.nature.comhttps://www.nature.com/articles/s41594-025-01699-y.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145440938","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Dicer has RNase activity and is an essential enzyme in microRNA (miRNA) biogenesis. Mutations in the DICER1 gene have been linked to various cancers, notably DICER1 syndrome. To investigate the impact of pathogenic hotspot mutations in DICER1-associated tumors, we introduced a hotspot mutation into the endogenous Dicer1 locus of a mouse embryonic carcinoma cell line using CRISPR. Our findings not only confirm the loss of 5p-miRNAs, as previously reported, but also demonstrate unexpected upregulation of specific 3p-miRNAs. These upregulated 3p-miRNAs, which are usually considered to be passenger strands in wild-type cells, are selectively loaded into the Argonaute protein in mutant cells based on their 5′-end characteristics, resulting in a ‘strand-switch’ phenomenon. Functional assays and transcriptome analyses demonstrate the activity of the passenger 3p-miRNAs. These results suggest that the Dicer hotspot mutation is not merely a loss-of-function mutation for 5p-miRNAs but also a gain-of-function mutation for passenger 3p-miRNAs, potentially contributing to DICER1-associated tumorigenesis. Malagobadan et al. show that a DICER1 hotspot mutation, previously thought to cause partial loss of function, also leads to unexpected gain of function by activating normally silent 3p passenger microRNAs, revealing a strand-switch mechanism in tumorigenesis.
{"title":"DICER1 hotspot mutation induces 3p microRNA gain of function via Argonaute strand switch","authors":"Sharan Malagobadan, Chunmei Shi, Acong Yang, Indranil Mondal, Habikah Baldeh, Karrie Spain, Wilfried Guiblet, Shuo Gu","doi":"10.1038/s41594-025-01671-w","DOIUrl":"10.1038/s41594-025-01671-w","url":null,"abstract":"Dicer has RNase activity and is an essential enzyme in microRNA (miRNA) biogenesis. Mutations in the DICER1 gene have been linked to various cancers, notably DICER1 syndrome. To investigate the impact of pathogenic hotspot mutations in DICER1-associated tumors, we introduced a hotspot mutation into the endogenous Dicer1 locus of a mouse embryonic carcinoma cell line using CRISPR. Our findings not only confirm the loss of 5p-miRNAs, as previously reported, but also demonstrate unexpected upregulation of specific 3p-miRNAs. These upregulated 3p-miRNAs, which are usually considered to be passenger strands in wild-type cells, are selectively loaded into the Argonaute protein in mutant cells based on their 5′-end characteristics, resulting in a ‘strand-switch’ phenomenon. Functional assays and transcriptome analyses demonstrate the activity of the passenger 3p-miRNAs. These results suggest that the Dicer hotspot mutation is not merely a loss-of-function mutation for 5p-miRNAs but also a gain-of-function mutation for passenger 3p-miRNAs, potentially contributing to DICER1-associated tumorigenesis. Malagobadan et al. show that a DICER1 hotspot mutation, previously thought to cause partial loss of function, also leads to unexpected gain of function by activating normally silent 3p passenger microRNAs, revealing a strand-switch mechanism in tumorigenesis.","PeriodicalId":49141,"journal":{"name":"Nature Structural & Molecular Biology","volume":"32 12","pages":"2542-2552"},"PeriodicalIF":10.1,"publicationDate":"2025-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.nature.comhttps://www.nature.com/articles/s41594-025-01671-w.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145434679","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-04DOI: 10.1038/s41594-025-01701-7
David Jee, Seungjae Lee, Dapeng Yang, Robert Rickert, Renfu Shang, Danwei Huangfu, Eric C. Lai
The core miRNA biogenesis enzyme DICER1 sustains recurrent mutations in cancer that compromise its RNase IIIb domain, which cleaves 5p arms of precursor microRNA hairpins. However, the lack of knock-in models has limited fuller understanding. Here, we generated DICER1-knockout and DICER1S1344L (homozygous and hemizygous) human embryonic stem cells; the latter is a noncatalytic substitution in RNase IIIa that impairs RNase IIIb activity. DICER1 knockouts lack canonical miRNAs, while S1344L induces two trends: ablation of miRNA-5p strands and selective changes in miRNA-3p strands. Curiously, we recognized directional upregulation of miRNA-3p passenger strands, indicating a broad strand switch. We used multiple in vitro assays to show 3p-arm-nicked pre-miRNAs preferentially load miRNA-3p species into Argonaute, compared to corresponding duplexes. Moreover, activity assays, RNA-sequencing data and Argonaute mRNA profiling confirm that these confer increased repression capacity. These data expand the molecular consequences of DICER1 hotspot mutations in cancer. Jee et al. study a cancer hotspot allele of DICER1 that disrupts RNaseIIIb activity. Beyond ablating 5p hairpin cleavage, 3p passenger strands are globally upregulated and active. Thus, this setting induces both loss and gain of miRNA function.
{"title":"Human DICER1 hotspot mutation induces both loss and gain of miRNA function","authors":"David Jee, Seungjae Lee, Dapeng Yang, Robert Rickert, Renfu Shang, Danwei Huangfu, Eric C. Lai","doi":"10.1038/s41594-025-01701-7","DOIUrl":"10.1038/s41594-025-01701-7","url":null,"abstract":"The core miRNA biogenesis enzyme DICER1 sustains recurrent mutations in cancer that compromise its RNase IIIb domain, which cleaves 5p arms of precursor microRNA hairpins. However, the lack of knock-in models has limited fuller understanding. Here, we generated DICER1-knockout and DICER1S1344L (homozygous and hemizygous) human embryonic stem cells; the latter is a noncatalytic substitution in RNase IIIa that impairs RNase IIIb activity. DICER1 knockouts lack canonical miRNAs, while S1344L induces two trends: ablation of miRNA-5p strands and selective changes in miRNA-3p strands. Curiously, we recognized directional upregulation of miRNA-3p passenger strands, indicating a broad strand switch. We used multiple in vitro assays to show 3p-arm-nicked pre-miRNAs preferentially load miRNA-3p species into Argonaute, compared to corresponding duplexes. Moreover, activity assays, RNA-sequencing data and Argonaute mRNA profiling confirm that these confer increased repression capacity. These data expand the molecular consequences of DICER1 hotspot mutations in cancer. Jee et al. study a cancer hotspot allele of DICER1 that disrupts RNaseIIIb activity. Beyond ablating 5p hairpin cleavage, 3p passenger strands are globally upregulated and active. Thus, this setting induces both loss and gain of miRNA function.","PeriodicalId":49141,"journal":{"name":"Nature Structural & Molecular Biology","volume":"32 12","pages":"2553-2563"},"PeriodicalIF":10.1,"publicationDate":"2025-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145434678","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-04DOI: 10.1038/s41594-025-01697-0
Qiwei Jiang, Peipei Song, Shuishen Zhang, Qin Ren, Meiyan Zhu, Jiawei Ge, Lang Bu, Wei Chen, Xueji Wu, Shiyao Han, Yaqing Su, Lei Wang, Wei Xie, Chao Cheng, Zhenwei Peng, Jianping Guo
Ubiquitination regulates various physiological and pathological processes. However, the impact of atypical AKT ubiquitination and its potential role in tumorigenesis remain unclear. Here we show that AKT is modified by K27-linked ubiquitination by the E3 ubiquitin ligase TRIM21, a process antagonized by the deubiquitinase OTUD6A. As such, TRIM21 acts as a tumor suppressor by repressing AKT activity, whereas OTUD6A counteracts AKT suppression. Mechanistically, TRIM21-mediated AKT ubiquitination disrupts SKP2-mediated or TRAF6-mediated K63 ubiquitination, thereby blocking AKT membrane localization and its kinase activity. Upon activation in response to amino acids, S6K1 directly phosphorylates and inactivates OTUD6A, enabling a negative feedback loop regulating AKT activity in a deubiquitination-dependent manner. In agreement with this model, Otud6a deficiency reduces lung tumorigenesis in a KrasG12D-driven lung cancer mouse model and TRIM21 induction alleviates hyperactive AKT-induced tumor growth in vivo. Thus, our findings unveil a fine-tuned regulation of AKT through atypical ubiquitination and suggest the strategy for combating AKT-driven cancers by targeting the TRIM21–OTUD6A axis. The authors here show that the TRIM21–OTUD6A axis controls AKT activity through K27-linked ubiquitination, which is further regulated by S6K1-mediated phosphorylation, introducing a putative therapeutic strategy against AKT-driven cancers.
{"title":"TRIM21 and OTUD6A orchestrate AKT K27-linked atypical ubiquitination to modulate cancer chemoresistance","authors":"Qiwei Jiang, Peipei Song, Shuishen Zhang, Qin Ren, Meiyan Zhu, Jiawei Ge, Lang Bu, Wei Chen, Xueji Wu, Shiyao Han, Yaqing Su, Lei Wang, Wei Xie, Chao Cheng, Zhenwei Peng, Jianping Guo","doi":"10.1038/s41594-025-01697-0","DOIUrl":"10.1038/s41594-025-01697-0","url":null,"abstract":"Ubiquitination regulates various physiological and pathological processes. However, the impact of atypical AKT ubiquitination and its potential role in tumorigenesis remain unclear. Here we show that AKT is modified by K27-linked ubiquitination by the E3 ubiquitin ligase TRIM21, a process antagonized by the deubiquitinase OTUD6A. As such, TRIM21 acts as a tumor suppressor by repressing AKT activity, whereas OTUD6A counteracts AKT suppression. Mechanistically, TRIM21-mediated AKT ubiquitination disrupts SKP2-mediated or TRAF6-mediated K63 ubiquitination, thereby blocking AKT membrane localization and its kinase activity. Upon activation in response to amino acids, S6K1 directly phosphorylates and inactivates OTUD6A, enabling a negative feedback loop regulating AKT activity in a deubiquitination-dependent manner. In agreement with this model, Otud6a deficiency reduces lung tumorigenesis in a KrasG12D-driven lung cancer mouse model and TRIM21 induction alleviates hyperactive AKT-induced tumor growth in vivo. Thus, our findings unveil a fine-tuned regulation of AKT through atypical ubiquitination and suggest the strategy for combating AKT-driven cancers by targeting the TRIM21–OTUD6A axis. The authors here show that the TRIM21–OTUD6A axis controls AKT activity through K27-linked ubiquitination, which is further regulated by S6K1-mediated phosphorylation, introducing a putative therapeutic strategy against AKT-driven cancers.","PeriodicalId":49141,"journal":{"name":"Nature Structural & Molecular Biology","volume":"33 1","pages":"84-99"},"PeriodicalIF":10.1,"publicationDate":"2025-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145434683","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-03DOI: 10.1038/s41594-025-01688-1
James A. W. Stowell, Conny W. H. Yu, Zhuo A. Chen, Lily K. DeBell, Giselle Lee, Tomos Morgan, Ludwig Sinn, Sylvie Agnello, Francis J. O’Reilly, Juri Rappsilber, Stefan M. V. Freund, Lori A. Passmore
Shortening of messenger RNA poly(A) tails by the Ccr4–Not complex initiates mRNA decay and is a major determinant of gene regulation. RNA adaptors modulate the specificity of deadenylation by binding to Ccr4–Not through their intrinsically disordered regions (IDRs). However, the determinants of specificity and their regulation are largely unclear. Here we use nuclear magnetic resonance spectroscopy, biochemical reconstitution and structural modeling to show that dispersed segments within the IDR of the fission yeast Puf3 RNA adaptor interact with Ccr4–Not, consistent with multivalency. Binding can be modulated by phosphorylation, altering the deadenylation rate in a continuously tunable manner. Regulation of deadenylation through multivalency and phosphorylation likely occurs in evolutionarily divergent IDRs from additional RNA adaptors, including human Pumilio and Tristetraprolin. Overall, our in vitro data suggest that mRNA decay can be regulated not only as a bistable on–off switch but also by a graded mechanism, rationalizing how post-transcriptional gene expression can be fine-tuned. Stowell et al. show that the intrinsically disordered region (IDR) of Puf3 binds the Ccr4–Not deadenylase complex using an extended interface. Phosphorylation of the IDR regulates the interaction to tune exonucleolytic activity in a graded manner.
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