Journal of Biological Chemistry最新文献_第9页

Topological isomers of a potent wound healing peptide: structural insights and implications for bioactivity. 一种有效的伤口愈合肽的拓扑异构体：结构的见解和对生物活性的影响。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-06-04 DOI: 10.1016/j.jbc.2025.110340

Tiziano Raffaelli, David T Wilson, Mehdi Mobli, Michael J Smout, Guangzu Zhao, Rozita Takjoo, Paramjit S Bansal, Rilei Yu, Zixuan Zhang, Alex Loukas, Norelle L Daly

{"title":"Topological isomers of a potent wound healing peptide: structural insights and implications for bioactivity.","authors":"Tiziano Raffaelli, David T Wilson, Mehdi Mobli, Michael J Smout, Guangzu Zhao, Rozita Takjoo, Paramjit S Bansal, Rilei Yu, Zixuan Zhang, Alex Loukas, Norelle L Daly","doi":"10.1016/j.jbc.2025.110340","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.110340","url":null,"abstract":"There are numerous examples of topological isomers in organic chemistry, but such isomers are rare in disulfide-rich peptides. Here we characterise two structurally well-defined topological isomers in a peptide (GRN-P4A) containing the mini-granulin fold. The mini-granulin fold is emerging as an important disulfide-rich structural motif with promising implications for the enhancement of wound healing strategies. The two topological isomers of GRN-P4A have well-defined structures that do not interconvert, and although they have the same disulfide bond connectivity and similar overall structures, they have structural differences related to the first inter-cysteine loop. These structural changes influence the bioactivity as the isomers have significant differences in their cell proliferation activity. Prediction of the structure using AlphaFold3 identified the correct disulfide bond connectivity, but the structure of loop 1 was similar to the less abundant isomer of GRN-P4A and did not indicate topological isomerisation. These topological isomers introduce significant complexity to the understanding of folding mechanisms in this class of peptides, and potentially other disulfide-rich peptides, offering valuable insights for protein design and engineering by presenting a novel topological fold-switching mechanism. Additionally, they hold practical implications for the production of GRN-P4A, given its promising potential as a wound-healing agent.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"110340"},"PeriodicalIF":4.0,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144248065","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

Endothelial Protein C Receptor and Thrombomodulin Facilitate Protease-Activated Receptor 1 Cleavage at Arginine-46 by Thrombin and Activated Protein C. 内皮蛋白C受体和凝血调节素促进凝血酶和活化蛋白C在精氨酸-46位点切割蛋白酶激活受体1。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-06-04 DOI: 10.1016/j.jbc.2025.110339

Indranil Biswas, Mariko Kudo, Alireza R Rezaie

{"title":"Endothelial Protein C Receptor and Thrombomodulin Facilitate Protease-Activated Receptor 1 Cleavage at Arginine-46 by Thrombin and Activated Protein C.","authors":"Indranil Biswas, Mariko Kudo, Alireza R Rezaie","doi":"10.1016/j.jbc.2025.110339","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.110339","url":null,"abstract":"Protease-activated receptor 1 (PAR1) has two cleavage sites for activation by coagulation proteases (Arg41 and Arg46). The cleavage of Arg46 by either activated protein C (APC) or thrombin leads to cytoprotective signaling, however, neither protease can cleave this site in the absence of their receptors, endothelial protein C receptor (EPCR) and thrombomodulin (TM), respectively. Arg41 is the preferred cleavage site for both proteases in the absence of receptors. The mechanism by which these receptors function as cofactors to catalyze the cleavage of PAR1-R46 by coagulation proteases is not known. Here, we hypothesized that both receptors alleviate inhibitory interactions of the P2-Leu45 residue on the extracellular domain of PAR1 with protease catalytic pockets. To test this hypothesis, we prepared a PAR1-R41A mutant in which P2-Leu45 of the receptor was substituted with a Pro. Both PAR1-R41A and PAR1-R41A-L45P constructs were transfected to PAR1-knockout EA.hy926 endothelial cells lacking or expressing EPCR or TM followed by monitoring the protease activation of receptors by signaling assays. Furthermore, wildtype or EPCR and TM expressing HEK-293 cells were transfected with PAR1 cleavage reporter constructs carrying N-terminal NanoLuc luciferase and C-terminal enhanced yellow fluorescent protein tags. Signaling and receptor cleavage assays indicated that both APC and thrombin cleave Arg46 in cells expressing PAR1-R41A-L45P, but not PAR1-41A, independent of their receptors. The catalytic activity of thrombin was >10-fold faster than APC in both assays. These results suggest that EPCR and TM function as cofactors to alleviate inhibitory interactions of P2-Leu45 of PAR1 with target proteases.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"110339"},"PeriodicalIF":4.0,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144247986","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

M54 selectively stabilizes the circadian clock component of CRY1 and enhances the period of circadian rhythm at cellular level. M54选择性地稳定CRY1的生物钟成分，并在细胞水平上提高昼夜节律的周期。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-06-04 DOI: 10.1016/j.jbc.2025.110333

Zeynep Melis Gül, Selahattin Aydoğan, Saliha Surme, Seden Nadire Harputluoğlu Efendi, Onur Özcan, Elif Uyanık, Ibrahim Baris, Seref Gul, Ibrahim Halil Kavakli

{"title":"M54 selectively stabilizes the circadian clock component of CRY1 and enhances the period of circadian rhythm at cellular level.","authors":"Zeynep Melis Gül, Selahattin Aydoğan, Saliha Surme, Seden Nadire Harputluoğlu Efendi, Onur Özcan, Elif Uyanık, Ibrahim Baris, Seref Gul, Ibrahim Halil Kavakli","doi":"10.1016/j.jbc.2025.110333","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.110333","url":null,"abstract":"Circadian rhythms are the daily oscillations in biochemical, physiological, and behavioral changes within living organisms, intricately tied to a 24-hour cycle and orchestrated by a molecular clock. This molecular clock operates through transcriptional-translational feedback loops generated by core clock proteins such as BMAL1, CLOCK, PERs, and CRYs. CRY1 and CRY2, along with PERs, repress BMAL1:CLOCK dependent transcriptional activity. In addition, several studies suggested that CRYs have differential functions in molecular clock. Our previous research identified M54 as a modulator of circadian rhythm at the cellular level through CRY1. Here, we demonstrate that M54 specifically binds to CRY1, but not CRY2, reducing the ubiquitination of CRY1 and, in turn, enhancing its stability. Consequently, M54 lengthens the period of the U2-OS circadian rhythm and decreases the transcription of clock-controlled genes in a concentration-dependent manner. This study opens a new avenue for therapeutic approaches targeting circadian clock-related disorders associated with dampened CRY1 levels.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"110333"},"PeriodicalIF":4.0,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144248058","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

Interactome of FMRP-N-tat therapeutic unveils key interactions for cellular function in Fragile X neurons. fmrp - n的相互作用组揭示了脆性X神经元细胞功能的关键相互作用。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-06-04 DOI: 10.1016/j.jbc.2025.110341

Kévin Leguay, Mariana Acevedo, Eva Colic, Preya U Patel, Saeideh Shamsi, Helen Lb Chan, Sharon Sun, Daneck Lang-Ouellette, Benny Chan, Xiaoqin Zhan, Ray W Turner, Joseph Mancini, Oliver A Kent

{"title":"Interactome of FMRP-N-tat therapeutic unveils key interactions for cellular function in Fragile X neurons.","authors":"Kévin Leguay, Mariana Acevedo, Eva Colic, Preya U Patel, Saeideh Shamsi, Helen Lb Chan, Sharon Sun, Daneck Lang-Ouellette, Benny Chan, Xiaoqin Zhan, Ray W Turner, Joseph Mancini, Oliver A Kent","doi":"10.1016/j.jbc.2025.110341","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.110341","url":null,"abstract":"Therapeutic protein replacement has demonstrated pre-clinical and clinical efficacy in neurological disorders but has not been used clinically for Fragile X syndrome (FXS), a genetic neurodevelopmental disorder caused by loss of Fragile X messenger ribonucleoprotein (FMRP). FXS results from a triplet repeat expansion of over 200 CGG repeats in the 5'-UTR of the FMR1 gene leading to epigenetic silencing of FMRP. Currently, no clinically approved disease-modifying treatments for FXS exist. Recently, a tat-conjugated FMRP fragment encompassing residues 1-297 (FMRP N-tat) was shown to restore aspects of neuronal function in a mouse model of FXS. Promising in vivo data hinted to the therapeutic potential of FMRP N-tat. Herein, affinity purification mass spectrometry was used to identify the FMRP N-tat interactome in tsA-201 FMR1 knockout cells and FXS patient iPSC-derived neurons. The FMRP N-tat interactome included RNA binding proteins and constituents of the ribosome, which aligned closely with the known functions of FMRP. Further, the FMRP N-tat associated proteins included FXR2, STAU1, TRIM28, C1QBP, VDAC2, and several ribosomal proteins to regulate mRNA stability, cellular stress responses, mitochondrial function, and translation. The results highlight the potential of FMRP N-tat to orchestrate assembly of factors to correct lost function in FMRP deficient cells.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"110341"},"PeriodicalIF":4.0,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144248057","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

Biophysical and structural analysis of KRAS switch-II pocket inhibitors reveals allele-specific binding constraints. KRAS开关- ii口袋抑制剂的生物物理和结构分析揭示了等位基因特异性结合约束。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-06-03 DOI: 10.1016/j.jbc.2025.110331

Patrick Alexander, Albert H Chan, Dana Rabara, Monalisa Swain, Erik K Larsen, Marcin Dyba, Oleg Chertov, Mariam Ashraf, Alison Champagne, Ken Lin, Anna Maciag, William K Gillette, Dwight V Nissley, Frank McCormick, Dhirendra K Simanshu, Andrew G Stephen

{"title":"Biophysical and structural analysis of KRAS switch-II pocket inhibitors reveals allele-specific binding constraints.","authors":"Patrick Alexander, Albert H Chan, Dana Rabara, Monalisa Swain, Erik K Larsen, Marcin Dyba, Oleg Chertov, Mariam Ashraf, Alison Champagne, Ken Lin, Anna Maciag, William K Gillette, Dwight V Nissley, Frank McCormick, Dhirendra K Simanshu, Andrew G Stephen","doi":"10.1016/j.jbc.2025.110331","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.110331","url":null,"abstract":"RAS mutations are observed in 20% of all cancers, with the KRAS isoform highly mutated in colorectal, lung and pancreatic cancers. The last several years have seen the development of clinical compounds that target KRAS G12C mutations, with other compounds under clinical development. In this study, a series of KRAS small-molecule inhibitors were compared for their binding affinity against a panel of KRAS mutant alleles. These inhibitors either covalently target the G12C mutant or reversibly target other mutants by binding in a transient pocket known as the switch-II pocket. Covalent inhibitors bound KRAS-GDP with KD values ranging from 10-9-10-3 M, whereas reversible inhibitors bound in the low nM range. A loss of affinity was observed for KRAS-GppNHp, due in part to rearrangements in switch-II, where the hydrogen bond between G60 and the γ-phosphate needs to break to form the switch-II pocket. Interestingly, these inhibitors had reduced affinity to KRAS Q61R-GppNHp, but not to WT and other mutants. The crystal structure of KRAS Q61R-GppNHp reported here revealed that access to the switch-II pocket was restricted due to R61 forming an additional hydrogen bond with the backbone carbonyl of T35 in switch-I. The restricted access to the switch-II pocket caused a decrease in the association rate of inhibitor binding and resulted in a loss of affinity. These findings across KRAS mutants provide valuable insights into the conformational adaptability of the switch-II pocket and may prove useful in developing the next generation of allele-specific and pan-KRAS small molecule inhibitors.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"110331"},"PeriodicalIF":4.0,"publicationDate":"2025-06-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144234204","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

Biosynthesis of fatty aldehydes and alcohols in the eye and their role in meibogenesis. 眼内脂肪醛和醇的生物合成及其在细胞新生中的作用。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-06-03 DOI: 10.1016/j.jbc.2025.110330

Seher Yuksel, Igor A Butovich

{"title":"Biosynthesis of fatty aldehydes and alcohols in the eye and their role in meibogenesis.","authors":"Seher Yuksel, Igor A Butovich","doi":"10.1016/j.jbc.2025.110330","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.110330","url":null,"abstract":"Fatty alcohols (FAlc) and aldehydes (FAld) are essential intermediates/precursors in the biosynthesis of lipids. However, elevated FAld levels were shown to be geno- and cytotoxic, and, thus, requiring conversion into less toxic FAlc and fatty acids (FA). An increase in FAlc and FAld in tissues of patients with Sjögren-Larsson syndrome was reported before and repeatedly linked to inactivation of ALDH3A2, which oxidizes FAld in FA. Recently, we hypothesized that another group of enzymes, namely SDR16C5/SDR16C6 (EC 1.1.1.105), could control the balance between FA, FAlc, and FAld via a separate mechanism. In this study, we assessed the in vivo biosynthesis of FAlc and FAld in mammals using Meibomian glands (MG) of wild type (WT) and Sdr16c5/Sdr16c6-null (Hom) mice as models. Lipids were extracted from MG of experimental animals and analyzed using LC/MS. Because of high reactivity and instability of FAld, the compounds were initially converted to stable, sodium borohydride-reduced 3-aminopyridine conjugates, while FAlc were analyzed as N-alkyl pyridinium ions. A wide range of saturated and unsaturated FAld, FAlc, and FA ranging from C3 to C28 and longer were found in MG of mice of both genotypes. Our experiments revealed a multifold upregulation of almost all detected straight chain, but not branched, FAlc in MG lipidomes of Hom mice, which implied a previously unknown ability of SDR16C5/SDR16C6 to oxidize a wide range of FAlc in FAld in vivo. We have concluded that SDR16C5/SDR16C6 play a central, and selective, role in FA/FAlc/FAld metabolism in vivo, and proposed a generalized mechanism of these reactions.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"110330"},"PeriodicalIF":4.0,"publicationDate":"2025-06-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144234205","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

Live-cell imaging of DNA damage and cell cycle progression uncovers distinct responses during neural differentiation of hiPSCs. DNA损伤和细胞周期进展的活细胞成像揭示了hiPSCs在神经分化过程中的不同反应。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-06-03 DOI: 10.1016/j.jbc.2025.110328

Mikio Shimada, Yoshihisa Matsumoto, Kensuke Otsuka

{"title":"Live-cell imaging of DNA damage and cell cycle progression uncovers distinct responses during neural differentiation of hiPSCs.","authors":"Mikio Shimada, Yoshihisa Matsumoto, Kensuke Otsuka","doi":"10.1016/j.jbc.2025.110328","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.110328","url":null,"abstract":"Ionizing radiation induces DNA double-strand breaks, which compromise genomic stability and trigger programmed cell death. The cell's differentiation state modulates DNA damage response (DDR) mechanisms, including DNA repair pathways and cell cycle regulation. The accumulation of p53-binding protein 1 (53BP1) at DSB sites serves as a reliable biomarker for such damage. Previously, we developed a fluorescent live-cell imaging system, termed \"Focicle,\" which monitors 53BP1 foci dynamics and cell cycle phases, utilizing fluorescent ubiquitination-based cell cycle indicators (hCdt1 and hGmnn) in mouse cells. In the current study, to investigate the relationship between differentiation state and DDR activity, we generated Focicle-integrated human induced pluripotent stem cells and further differentiated them into neural progenitors and mature neurons using an optimized Focicle cassette adapted for human cell lines. Using laser microirradiation, we observed differentiation-dependent alterations in 53BP1 foci accumulation dynamics and cell cycle progression. The newly established Focicle system represents a valuable tool for elucidating DDR activity during organ development.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"110328"},"PeriodicalIF":4.0,"publicationDate":"2025-06-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144234209","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

Mitochondrial tRNA processing defects reprogram mitochondrial and cellular homeostasis. 线粒体tRNA加工缺陷重编程线粒体和细胞稳态。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-06-03 DOI: 10.1016/j.jbc.2025.110334

Gao Zhu, Yunfan He, Xincheng Li, Yun Xiao, Huisen Zhan, Maoli Duan, Min-Xin Guan

{"title":"Mitochondrial tRNA processing defects reprogram mitochondrial and cellular homeostasis.","authors":"Gao Zhu, Yunfan He, Xincheng Li, Yun Xiao, Huisen Zhan, Maoli Duan, Min-Xin Guan","doi":"10.1016/j.jbc.2025.110334","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.110334","url":null,"abstract":"Mitochondrial tRNA processing defects have been associated with some clinical presentations including deafness. Especially, a deafness-linked m.7516delA mutation impaired the 5' end processing of RNA precursors and mitochondrial translation. In this study, we investigated the mechanism by m.7516delA mutation induced-deficiencies mitigate organellular and cellular integrity. The m.7516delA mutation downregulated the expression of nucleus encoding subunits and upregulated assemble factors of complex IV and altered the assembly and activities of oxidative phosphorylation (OXPHOS) complexes. The impairment of OXPHOS alleviated mitochondrial quality control processes, including the imbalanced mitochondrial dynamics via increasing fission with abnormal mitochondrial morphology. The m.7516delA mutation upregulated both ubiquitin-dependent and independent mitophagy pathways, evidenced by increasing levels of Parkin, BNIP3, NIX and MFN2-ubiquitination and altering interaction between MFN2 and MUL1 or Parkin, to facilitate the degradation of severely damaged mitochondria. Strikingly, the m.7516delA mutation activated integrated stress response (ISR) pathway, evidenced by upregulation of GCN2, P-GCN2, p-eIF2α, CHOP, ATF4 and elevating the nucleus-location of ATF5 to minimizes the damages in defective mitochondria. Both activation of ISR and PINK1/Parkin mitophagy pathways ameliorate the cell homeostasis via elevating the autophagy process and upregulating apoptotic pathways. Our findings provide new insights into underlying aberrant RNA processing-induced dysfunctions reprogrammed organelles and cellular integrity.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"110334"},"PeriodicalIF":4.0,"publicationDate":"2025-06-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144234210","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

Identification of substrates and sequence requirements for CARNMT1-mediated histidine methylation of C3H zinc fingers. carnmt1介导C3H锌指组氨酸甲基化的底物鉴定和序列要求。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-06-03 DOI: 10.1016/j.jbc.2025.110335

Jędrzej M Małecki, Sara Weirich, Manuel Ramirez-Garrastacho, Lars Hagen, Jakin Al-Egly, Jan H Anonsen, Lisa Schroer, Maria C Herrera, Erna Davydova, Geir Slupphaug, Albert Jeltsch, Pål Ø Falnes

{"title":"Identification of substrates and sequence requirements for CARNMT1-mediated histidine methylation of C3H zinc fingers.","authors":"Jędrzej M Małecki, Sara Weirich, Manuel Ramirez-Garrastacho, Lars Hagen, Jakin Al-Egly, Jan H Anonsen, Lisa Schroer, Maria C Herrera, Erna Davydova, Geir Slupphaug, Albert Jeltsch, Pål Ø Falnes","doi":"10.1016/j.jbc.2025.110335","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.110335","url":null,"abstract":"It has recently become clear that protein histidine methylation is widespread and functionally important, and human CARNMT1 was recently reported as a novel protein histidine methyltransferase (HMT). We describe our independent uncovering of CARNMT1's protein HMT activity, and a comprehensive assessment of its methylation targets and substrate specificity. Using a combination of in vitro methylation of cellular extracts and protein mass spectrometry, we identified several CARNMT1 targets that were fully methylated in cells, all of which were C3H zinc finger (ZnF) proteins. These included the previously identified U2AF1, ZC3H15 and ZC3H18, but also the unreported RBM22, PPP1R10, PRR3 and RNF113A. Using peptide arrays, we investigated CARNMT1-mediated methylation of 145 candidate sequences, encompassing all C3H ZnFs and selected non-ZnFs. We found that only ∼30 % of the tested sequences were methylated, with C3H ZnFs constituting the vast majority of the strongly methylated ones, most of which are also methylated in cells. This establishes peptide methylation as a good predictor of in vivo methylation. To investigate the specificity of CARNMT1, we systematically substituted His-proximal residues in four different substrate peptides. This generated four rather different activity profiles, which were still quite restrictive for each peptide, indicating that molecular recognition by CARNMT1 is context-dependent, and that sequence-based prediction of additional CARNMT1 substrates may be challenging. We also identified several homologous methylation events in Caenorhabditis elegans and showed that they could be introduced by nematode CARNMT in vitro. Thus, CARNMT1 is an evolutionary conserved protein HMT with a complex mode of substrate recognition.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"110335"},"PeriodicalIF":4.0,"publicationDate":"2025-06-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144234208","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

Characterization of C. elegans sphingomyelin synthases through heterologous expression. 秀丽隐杆线虫鞘磷脂合成酶的异源表达研究。

IF 4 2区生物学

Journal of Biological Chemistry Pub Date : 2025-06-03 DOI: 10.1016/j.jbc.2025.110300

Gaelen Guzman, Helene Jahn, Scotland E Farley, Lisa Bramer, Jennifer Kyle, Sandra Hoetzl, Joep van den Dikkenberg, Martin Hermansson, Joost C M Holthuis, Fikadu G Tafesse

{"title":"Characterization of C. elegans sphingomyelin synthases through heterologous expression.","authors":"Gaelen Guzman, Helene Jahn, Scotland E Farley, Lisa Bramer, Jennifer Kyle, Sandra Hoetzl, Joep van den Dikkenberg, Martin Hermansson, Joost C M Holthuis, Fikadu G Tafesse","doi":"10.1016/j.jbc.2025.110300","DOIUrl":"https://doi.org/10.1016/j.jbc.2025.110300","url":null,"abstract":"Sphingomyelin (SM) is a major component of mammalian cell membranes and particularly abundant in the myelin sheath that surrounds nerve fibers. Its production is catalyzed by SM synthases SMS1 and SMS2, which interconvert phosphatidylcholine and ceramide to diacylglycerol and SM in the Golgi and at the plasma membrane, respectively. As the lipids participating in this reaction fulfill both structural and signaling functions, SMS enzymes have considerable potential to influence diverse important cellular processes. The nematode Caenorhabditis elegans is an attractive model for studying both animal development and human disease. The organism contains five SMS homologues but none of these have been characterized in any detail. Here, we carried out the first systematic analysis of SMS family members in C. elegans. Using heterologous expression systems, genetic ablation, metabolic labeling and lipidome analyses, we show that C. elegans harbors at least three distinct SM synthases and one ceramide phosphoethanolamine (CPE) synthase. Moreover, C. elegans SMS family members have partially overlapping but also unique sub-cellular distributions and together occupy all principal compartments of the secretory pathway. Our findings shed light on crucial aspects of sphingolipid metabolism in a valuable animal model and opens avenues for exploring the role of SM and its metabolic intermediates in organismal development.","PeriodicalId":15140,"journal":{"name":"Journal of Biological Chemistry","volume":" ","pages":"110300"},"PeriodicalIF":4.0,"publicationDate":"2025-06-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144234206","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