Pub Date : 2025-02-01DOI: 10.1016/j.bbagen.2024.130742
Joanne Jennifer E. Tan , Marvin M. Bilog , Adam A. Profit , Francisco M. Heralde III , Ruel Z.B. Desamero
<div><div>Atherosclerosis, the major underlying cause of cardiovascular disease, is believed to arise from the accumulation of low-density lipoprotein (LDL) in the arterial subendothelial space, ultimately leading to plaque formation. It is proposed that the accumulation of LDL is linked to its intrinsic aggregation propensity. Although the native LDL is not prone to aggregation, LDL(−), an electronegative LDL characterized in the plasma, has been shown to prime LDL aggregation in a domino-like behavior similar to amyloidogenic proteins. LDL(−) has also been observed to have a misfolded apolipoprotein B-100 (apo B-100), a huge protein consisting of 4563 amino acid residues. As misfolding of proteins is commonly associated with amyloid formation, apo B-100 is therefore being considered as the possible triggering factor in LDL aggregation. Previous computational studies have implicated the α2 domain to be the aggregation-prone region of apo B-100. In this study, the amyloidogenic properties of the α2 domain of apo B-100 were interrogated using both <em>in silico</em> and <em>in vitro</em> techniques.</div><div>Since the crystal structure of the 570-amino acid α2 domain of apo B-100 is yet to be solved, we used several secondary structure prediction tools to model putative helical regions that make up the α2 domain. The stability of each of the 17 helices thus identified was further probed using molecular dynamics (MD), with the least stable of the helices considered as potentially amyloidogenic. In a 100 ns simulation window, helices <strong><em>k</em></strong> (YFEKLVGFIDDAVK), <strong><em>m</em></strong> (YHQFVDETNDKIREVTQRLNGEIQA), and <strong><em>p</em></strong> (QQELQRYLSLVGQVYS) were the least stable and appeared to transition to β-structures, the hallmark of amyloidogenesis. When the simulation was extended to longer times, only helices <strong><em>k</em></strong> and <strong><em>p</em></strong> formed stable β-sheets that persisted. Analysis of the data indicates that the final β-sheet conformation was stabilized by the π-π stacking interactions between the aromatic rings of Tyr-1 and Phe-8 for helix <strong><em>k</em></strong> and likely π-π stacking contacts between Arg-6 guanidino group and Tyr-15 ring for helix <strong><em>p</em></strong>.</div><div>Based on the <em>in silico</em> work, we proceeded to synthesize and spectroscopically characterize helices <strong><em>k</em></strong>, <strong><em>m</em></strong><sub><strong><em>17</em></strong></sub><sub><strong><em>–</em></strong></sub><sub><strong><em>25</em></strong></sub> (QRLNGEIQA), and <strong><em>p</em></strong>. As expected, <strong><em>k</em></strong> and <strong><em>p</em></strong> formed detectable amyloids, with the latter appearing to be substantially more amyloidogenic based on kinetic aggregation assays. Amyloid fibrils formed by <strong><em>p</em></strong> were confirmed using circular dichroism spectroscopy and transmission electron microscopy. Data obtained could be ex
动脉粥样硬化是心血管疾病的主要潜在原因，被认为是由动脉内皮下空间低密度脂蛋白（LDL）的积累引起的，最终导致斑块的形成。有人提出LDL的积累与其内在的聚集倾向有关。虽然天然LDL不容易聚集，但LDL(-)，一种在血浆中具有电负性的LDL，已被证明以类似于淀粉样蛋白的多米诺骨牌行为引发LDL聚集。LDL（-）也被观察到有一个错误折叠的载脂蛋白B-100（载脂蛋白B-100），一个由4563个氨基酸残基组成的巨大蛋白质。由于蛋白质的错误折叠通常与淀粉样蛋白的形成有关，因此载脂蛋白B-100被认为是LDL聚集的可能触发因素。先前的计算研究表明α2结构域是载脂蛋白B-100的聚集易发区域。在这项研究中，我们用硅和体外技术研究了载脂蛋白B-100 α2结构域的淀粉样变性。由于载脂蛋白B-100的570个氨基酸α2结构域的晶体结构尚未解决，我们使用了几个二级结构预测工具来模拟推定的构成α2结构域的螺旋区域。用分子动力学（MD）进一步研究了17个螺旋的稳定性，其中最不稳定的螺旋被认为是潜在的淀粉样蛋白。在100 ns的模拟窗口中，螺旋k (YFEKLVGFIDDAVK), m （YHQFVDETNDKIREVTQRLNGEIQA）和p （QQELQRYLSLVGQVYS）是最不稳定的，并且似乎转变为β-结构，这是淀粉样变性的标志。当模拟延长到更长的时间时，只有螺旋k和p形成了持续存在的稳定β片。数据分析表明，最终的β-薄片构象是由螺旋k的tyr1和phe8芳香环之间的π-π堆叠相互作用以及螺旋p的Arg-6胍基与tyr15环之间可能的π-π堆叠相互作用稳定的。在硅工作的基础上，我们继续合成和光谱表征了螺旋k， m17-25 （QRLNGEIQA）和p。根据动力学聚集试验，后者似乎更具有淀粉样变性。通过圆二色光谱和透射电镜证实了p形成的淀粉样原纤维。获得的数据可用于进一步研究载脂蛋白B-100 α2结构域螺旋衍生的肽在触发LDL聚集中的作用。根据初步数据，基于这项工作设计的肽之一减少了LDL的聚集。
{"title":"Computational analysis of the alpha−2 domain of apolipoprotein B − 100, a potential triggering factor in LDL aggregation","authors":"Joanne Jennifer E. Tan , Marvin M. Bilog , Adam A. Profit , Francisco M. Heralde III , Ruel Z.B. Desamero","doi":"10.1016/j.bbagen.2024.130742","DOIUrl":"10.1016/j.bbagen.2024.130742","url":null,"abstract":"<div><div>Atherosclerosis, the major underlying cause of cardiovascular disease, is believed to arise from the accumulation of low-density lipoprotein (LDL) in the arterial subendothelial space, ultimately leading to plaque formation. It is proposed that the accumulation of LDL is linked to its intrinsic aggregation propensity. Although the native LDL is not prone to aggregation, LDL(−), an electronegative LDL characterized in the plasma, has been shown to prime LDL aggregation in a domino-like behavior similar to amyloidogenic proteins. LDL(−) has also been observed to have a misfolded apolipoprotein B-100 (apo B-100), a huge protein consisting of 4563 amino acid residues. As misfolding of proteins is commonly associated with amyloid formation, apo B-100 is therefore being considered as the possible triggering factor in LDL aggregation. Previous computational studies have implicated the α2 domain to be the aggregation-prone region of apo B-100. In this study, the amyloidogenic properties of the α2 domain of apo B-100 were interrogated using both <em>in silico</em> and <em>in vitro</em> techniques.</div><div>Since the crystal structure of the 570-amino acid α2 domain of apo B-100 is yet to be solved, we used several secondary structure prediction tools to model putative helical regions that make up the α2 domain. The stability of each of the 17 helices thus identified was further probed using molecular dynamics (MD), with the least stable of the helices considered as potentially amyloidogenic. In a 100 ns simulation window, helices <strong><em>k</em></strong> (YFEKLVGFIDDAVK), <strong><em>m</em></strong> (YHQFVDETNDKIREVTQRLNGEIQA), and <strong><em>p</em></strong> (QQELQRYLSLVGQVYS) were the least stable and appeared to transition to β-structures, the hallmark of amyloidogenesis. When the simulation was extended to longer times, only helices <strong><em>k</em></strong> and <strong><em>p</em></strong> formed stable β-sheets that persisted. Analysis of the data indicates that the final β-sheet conformation was stabilized by the π-π stacking interactions between the aromatic rings of Tyr-1 and Phe-8 for helix <strong><em>k</em></strong> and likely π-π stacking contacts between Arg-6 guanidino group and Tyr-15 ring for helix <strong><em>p</em></strong>.</div><div>Based on the <em>in silico</em> work, we proceeded to synthesize and spectroscopically characterize helices <strong><em>k</em></strong>, <strong><em>m</em></strong><sub><strong><em>17</em></strong></sub><sub><strong><em>–</em></strong></sub><sub><strong><em>25</em></strong></sub> (QRLNGEIQA), and <strong><em>p</em></strong>. As expected, <strong><em>k</em></strong> and <strong><em>p</em></strong> formed detectable amyloids, with the latter appearing to be substantially more amyloidogenic based on kinetic aggregation assays. Amyloid fibrils formed by <strong><em>p</em></strong> were confirmed using circular dichroism spectroscopy and transmission electron microscopy. Data obtained could be ex","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1869 2","pages":"Article 130742"},"PeriodicalIF":2.8,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142833772","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-01DOI: 10.1016/j.bbagen.2024.130743
Bartosz Klebowski , Karolina Kosinska , Agnieszka Bukowska , Piotr M. Zieliński , Magdalena Parlinska-Wojtan , Joanna Depciuch
Titanium oxide nanoparticles (TiO2 NPs) are currently used as ingredients in medicines and sunscreens. Unfortunately, recent information about TiO2 NPs indicates their undesirable biological effect on colon cells. Therefore, the aim of this work was to synthesize and evaluate the physicochemical characterization of spherical (TiO2 NSs) and rods-like (TiO2 NRs) NPs, followed by assessment their cytotoxicity. For this purpose, both normal colon epithelial cells (CRL-1790) and cancerous colon cells (SW480) were used. Scanning transmission electron microscopy (STEM) showed that TiO2 NSs with a diameter of ≈10 nm and TiO2 NRs with the size of the longer axis ≈25 nm and shorter axis ≈3 nm were obtained. Based on the selected area electron diffraction (SAED) patterns, it was found that crystalline phases were obtained for both TiO2 NPs. The UV–Vis spectra showed no contamination of TiO2 NPs. Zeta potential values were 9.7 mV and 3.1 mV for NSs and NRs, respectively. Cytotoxicity of TiO2 NPs was assessed using the MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxy-methoxy-phenyl)-2-(4-sulfophenyl)-2H-tetrazolium) test for various concentration of NPs. The cytotoxic effect for both TiO2 NPs was visible for concentration of 75 μg/ml (for CRL-1790) and 50 μg/ml (for SW480) and higher, and it did not depend on the shape. Moreover, both types of TiO2 NPs (in higher concentration) induce the generation of reactive oxygen species (ROS) in cells cultured with these NPs. Holotomographic microscopy studies showed increased cellular uptake of TiO2 NPs by SW480. The obtained results for the synthesized TiO2 NPs are a promising prospect for their use in biomedical application.
{"title":"Synthesis of spherical and rods-like titanium oxide nanoparticles (TiO2 NPs) and evaluation of their cytotoxicity towards colon cells in vitro","authors":"Bartosz Klebowski , Karolina Kosinska , Agnieszka Bukowska , Piotr M. Zieliński , Magdalena Parlinska-Wojtan , Joanna Depciuch","doi":"10.1016/j.bbagen.2024.130743","DOIUrl":"10.1016/j.bbagen.2024.130743","url":null,"abstract":"<div><div>Titanium oxide nanoparticles (TiO<sub>2</sub> NPs) are currently used as ingredients in medicines and sunscreens. Unfortunately, recent information about TiO<sub>2</sub> NPs indicates their undesirable biological effect on colon cells. Therefore, the aim of this work was to synthesize and evaluate the physicochemical characterization of spherical (TiO<sub>2</sub> NSs) and rods-like (TiO<sub>2</sub> NRs) NPs, followed by assessment their cytotoxicity. For this purpose, both normal colon epithelial cells (CRL-1790) and cancerous colon cells (SW480) were used. Scanning transmission electron microscopy (STEM) showed that TiO<sub>2</sub> NSs with a diameter of ≈10 nm and TiO<sub>2</sub> NRs with the size of the longer axis ≈25 nm and shorter axis ≈3 nm were obtained. Based on the selected area electron diffraction (SAED) patterns, it was found that crystalline phases were obtained for both TiO<sub>2</sub> NPs. The UV–Vis spectra showed no contamination of TiO<sub>2</sub> NPs. Zeta potential values were 9.7 mV and 3.1 mV for NSs and NRs, respectively. Cytotoxicity of TiO<sub>2</sub> NPs was assessed using the MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxy-methoxy-phenyl)-2-(4-sulfophenyl)-2H-tetrazolium) test for various concentration of NPs. The cytotoxic effect for both TiO<sub>2</sub> NPs was visible for concentration of 75 μg/ml (for CRL-1790) and 50 μg/ml (for SW480) and higher, and it did not depend on the shape. Moreover, both types of TiO<sub>2</sub> NPs (in higher concentration) induce the generation of reactive oxygen species (ROS) in cells cultured with these NPs. Holotomographic microscopy studies showed increased cellular uptake of TiO<sub>2</sub> NPs by SW480. The obtained results for the synthesized TiO<sub>2</sub> NPs are a promising prospect for their use in biomedical application.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1869 2","pages":"Article 130743"},"PeriodicalIF":2.8,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142833775","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Obesity-induced monocyte chemoattractant protein 1 (MCP-1) production leads to the infiltration of monocytes/macrophages into white adipose tissue (WAT), which contributes to systemic insulin resistance. Peroxisome proliferator-activated receptor gamma (PPARγ) agonists are known to reduce MCP-1 production in both humans and mice; however, the underlying mechanism in WAT remains unclear. Here, we propose a novel mechanism for the reduction in MCP-1 production in preadipocytes. The PPARγ agonist rosiglitazone (RSG) reduced MCP-1 production and secretion in response to lipopolysaccharide (LPS) in 3T3-L1 preadipocytes and mouse stromal vascular fraction–derived primary preadipocytes. Both RSG and SP600125 (a c-Jun N-terminal kinase (JNK) inhibitor) inhibited LPS-induced degradation of silent information regulator 2 homolog 1 (SIRT1), a negative regulator of MCP-1 production in 3T3-L1 preadipocytes. Furthermore, RSG inhibited LPS-induced activation of nuclear factor-κB. These effects of RSG were abolished in 3T3-L1 preadipocytes transfected with Pparg siRNA. These findings highlight a novel mechanism by which PPARγ activation inhibits JNK/SIRT1 signaling in preadipocytes and contributes to the reduction in MCP-1 production, suggesting that preadipocytes could be a potential therapeutic target for the treatment of insulin resistance.
肥胖诱导的单核细胞化学引诱蛋白1 （MCP-1）的产生导致单核细胞/巨噬细胞浸润到白色脂肪组织（WAT），这有助于全身性胰岛素抵抗。已知过氧化物酶体增殖物激活受体γ (PPARγ)激动剂可减少人类和小鼠MCP-1的产生；然而，WAT的潜在机制尚不清楚。在这里，我们提出了一种减少前脂肪细胞中MCP-1产生的新机制。PPARγ激动剂罗格列酮（RSG）减少了3 T3-L1前脂肪细胞和小鼠间质血管部分来源的原发性前脂肪细胞对脂多糖（LPS）的反应中MCP-1的产生和分泌。RSG和SP600125（一种c-Jun n -末端激酶（JNK）抑制剂）均抑制lps诱导的沉默信息调节因子2同源物1 （SIRT1）的降解，SIRT1是3 T3-L1前脂肪细胞中MCP-1产生的负调节因子。此外，RSG抑制lps诱导的核因子-κB活化。RSG的这些作用在3 T3-L1前脂肪细胞转染Pparg siRNA后被消除。这些发现强调了PPARγ激活抑制前脂肪细胞中JNK/SIRT1信号传导并有助于减少MCP-1产生的新机制，表明前脂肪细胞可能是治疗胰岛素抵抗的潜在治疗靶点。
{"title":"Reduction in MCP-1 production in preadipocytes is mediated by PPARγ activation and JNK/SIRT1 signaling","authors":"Atsushi Sawamoto, Ibuki Itagaki, Satoshi Okuyama, Mitsunari Nakajima","doi":"10.1016/j.bbagen.2024.130737","DOIUrl":"10.1016/j.bbagen.2024.130737","url":null,"abstract":"<div><div>Obesity-induced monocyte chemoattractant protein 1 (MCP-1) production leads to the infiltration of monocytes/macrophages into white adipose tissue (WAT), which contributes to systemic insulin resistance. Peroxisome proliferator-activated receptor gamma (PPARγ) agonists are known to reduce MCP-1 production in both humans and mice; however, the underlying mechanism in WAT remains unclear. Here, we propose a novel mechanism for the reduction in MCP-1 production in preadipocytes. The PPARγ agonist rosiglitazone (RSG) reduced MCP-1 production and secretion in response to lipopolysaccharide (LPS) in 3T3-L1 preadipocytes and mouse stromal vascular fraction–derived primary preadipocytes. Both RSG and SP600125 (a c-Jun N-terminal kinase (JNK) inhibitor) inhibited LPS-induced degradation of silent information regulator 2 homolog 1 (SIRT1), a negative regulator of MCP-1 production in 3T3-L1 preadipocytes. Furthermore, RSG inhibited LPS-induced activation of nuclear factor-κB. These effects of RSG were abolished in 3T3-L1 preadipocytes transfected with <em>Pparg</em> siRNA. These findings highlight a novel mechanism by which PPARγ activation inhibits JNK/SIRT1 signaling in preadipocytes and contributes to the reduction in MCP-1 production, suggesting that preadipocytes could be a potential therapeutic target for the treatment of insulin resistance.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1869 2","pages":"Article 130737"},"PeriodicalIF":2.8,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142821763","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-01DOI: 10.1016/j.bbagen.2024.130745
Yangyang Xiao , Dan Zou , Jianan Liu , Fanfan Dai , Ansha Zhao , Ping Yang
Endothelial cell-sourced exosomes are potential participants in the process of atherosclerosis, and their function is mainly affected by concentration. By studying the effects of exosome concentrations on vascular cells, atherosclerosis can be better intervened. In this study, exosomes with concentrations of 0, 0.07, 0.35, 1.75 and 8.75 μg/mL were set to interact with endothelial cells, macrophages and smooth muscle cells respectively. The results suggested that EC-Exo altered vascular cells' proliferation, migration and nitric oxide release abilities, increasing with EC-Exo concentrate from 0 to 1.75 μg/mL and varing with cell types at 8.75 μg/mL. The effects of exosome on cells is dose-responsive，and endothelial cells-sourced exosome favors vascular repair within the concentration of 0.35–1.75 μg/mL，showing potential for atherosclerosis regulation.
{"title":"Dose-responsive effects of endothelial cell-sourced exosomes on vascular cell proliferation and phenotype transition","authors":"Yangyang Xiao , Dan Zou , Jianan Liu , Fanfan Dai , Ansha Zhao , Ping Yang","doi":"10.1016/j.bbagen.2024.130745","DOIUrl":"10.1016/j.bbagen.2024.130745","url":null,"abstract":"<div><div>Endothelial cell-sourced exosomes are potential participants in the process of atherosclerosis, and their function is mainly affected by concentration. By studying the effects of exosome concentrations on vascular cells, atherosclerosis can be better intervened. In this study, exosomes with concentrations of 0, 0.07, 0.35, 1.75 and 8.75 μg/mL were set to interact with endothelial cells, macrophages and smooth muscle cells respectively. The results suggested that EC-Exo altered vascular cells' proliferation, migration and nitric oxide release abilities, increasing with EC-Exo concentrate from 0 to 1.75 μg/mL and varing with cell types at 8.75 μg/mL. The effects of exosome on cells is dose-responsive，and endothelial cells-sourced exosome favors vascular repair within the concentration of 0.35–1.75 μg/mL，showing potential for atherosclerosis regulation.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1869 2","pages":"Article 130745"},"PeriodicalIF":2.8,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142845530","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-01DOI: 10.1016/j.bbagen.2024.130741
Krzysztof Żamojć , Dan Milaș , Ola Grabowska , Dariusz Wyrzykowski , Magdalena Mańkowska , Karol Krzymiński
The study delves into the binding properties of acridine-9-amine and its selected, mainly N-substituted derivatives (A9As), with calf thymus deoxyribonucleic acid (CT-DNA). This investigation, conducted using UV–Vis spectrophotometry, steady-state fluorescence spectroscopy and isothermal titration calorimetry, provides insights into the relationship between their structure and activity. The absorption spectra of the A9As exhibited a slight red shift and significant hypochromic effects, while the fluorescence emission intensities decreased in the presence of CT-DNA. These results suggest that all fluorescent substrates intercalate into the double helix of native DNA to varying degrees. The binding constants for the A9As/CT-DNA complexes (log(KA) were determined using various techniques in the range from 2.59 to 5.50). The thermodynamic parameters of A9As binding to DNA were obtained from ITC measurements (ΔG from – 7.51 to – 6.75 kcal·mol−1, ΔH from – 11.58 to – 3.83 kcal·mol−1, and TΔS from – 4.83 to 3.68 kcal·mol−1) and indicated that the formation of all the investigated A9As–DNA complexes is an enthalpy-driven process. The study also discusses the influence of the emitters' structure and electronic properties of substituents on intercalation efficiency. This knowledge serves as a guide for further research and offers directions for functionalising new acridines as potential reagents. It also provides the latest information on the ability of intercalation to DNA, which can be instrumental in studies on the mechanism of binding small aromatic molecules to DNA and can potentially contribute to new anticancer drug designs.
{"title":"Insight into the intercalation of N-substituted acridine-9-amines into DNA based on spectroscopic and calorimetric analysis","authors":"Krzysztof Żamojć , Dan Milaș , Ola Grabowska , Dariusz Wyrzykowski , Magdalena Mańkowska , Karol Krzymiński","doi":"10.1016/j.bbagen.2024.130741","DOIUrl":"10.1016/j.bbagen.2024.130741","url":null,"abstract":"<div><div>The study delves into the binding properties of acridine-9-amine and its selected, mainly <em>N</em>-substituted derivatives (A9As), with calf thymus deoxyribonucleic acid (CT-DNA). This investigation, conducted using UV–Vis spectrophotometry, steady-state fluorescence spectroscopy and isothermal titration calorimetry, provides insights into the relationship between their structure and activity. The absorption spectra of the A9As exhibited a slight red shift and significant hypochromic effects, while the fluorescence emission intensities decreased in the presence of CT-DNA. These results suggest that all fluorescent substrates intercalate into the double helix of native DNA to varying degrees. The binding constants for the A9As/CT-DNA complexes (log(<em>K</em><sub>A</sub>) were determined using various techniques in the range from 2.59 to 5.50). The thermodynamic parameters of A9As binding to DNA were obtained from ITC measurements (Δ<em>G</em> from – 7.51 to – 6.75 kcal·mol<sup>−1</sup>, Δ<em>H</em> from – 11.58 to – 3.83 kcal·mol<sup>−1</sup>, and <em>T</em>Δ<em>S</em> from – 4.83 to 3.68 kcal·mol<sup>−1</sup>) and indicated that the formation of all the investigated A9As–DNA complexes is an enthalpy-driven process. The study also discusses the influence of the emitters' structure and electronic properties of substituents on intercalation efficiency. This knowledge serves as a guide for further research and offers directions for functionalising new acridines as potential reagents. It also provides the latest information on the ability of intercalation to DNA, which can be instrumental in studies on the mechanism of binding small aromatic molecules to DNA and can potentially contribute to new anticancer drug designs.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1869 2","pages":"Article 130741"},"PeriodicalIF":2.8,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142852299","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-01DOI: 10.1016/j.bbagen.2024.130748
Anne M.L. Nilsen , Galice Hoarau , Irina Smolina , James A. Coyer , Christoffer Boström , Martina E.L. Kopp , Alexander Jueterbock
Factors influencing variance of DNA methylation in vegetatively reproducing plants, both terrestrial plants and aquatic seagrasses, is just beginning to be understood. Improving our knowledge of these mechanisms will increase understanding of transgenerational epigenetics in plant clones, of the relationship between DNA methylation and seagrass development, and of the drivers of epigenetic variation, which may underly acclimation in clonally reproducing plants. Here, we sampled leaves, rhizomes and roots of three physically and spatially separated ramet sections from a clonally propagated field of the seagrass Zostera marina. Using reduced methylome sequencing, we studied variations in the methylome of seagrass Zostera marina between the sampled tissue types and across age groups. Our analysis of ramets of different ages showed variations in methylation between older and younger samples in both specific methylation patterns and global methylation levels. Our analysis of tissue types showed a marked differentiation of the roots from the rhizomes and leaves, which showed more similar methylation patterns. These findings are in agreement with the strong connection of DNA methylation and plant development and tissue differentiation. We also suggest an effect of differential environmental exposures on the methylome of the younger versus the older ramets due to the forming of molecular stress memories.
{"title":"The methylome of clonal seagrass shoots shows age-associated variation and differentiation of roots from other tissues","authors":"Anne M.L. Nilsen , Galice Hoarau , Irina Smolina , James A. Coyer , Christoffer Boström , Martina E.L. Kopp , Alexander Jueterbock","doi":"10.1016/j.bbagen.2024.130748","DOIUrl":"10.1016/j.bbagen.2024.130748","url":null,"abstract":"<div><div>Factors influencing variance of DNA methylation in vegetatively reproducing plants, both terrestrial plants and aquatic seagrasses, is just beginning to be understood. Improving our knowledge of these mechanisms will increase understanding of transgenerational epigenetics in plant clones, of the relationship between DNA methylation and seagrass development, and of the drivers of epigenetic variation, which may underly acclimation in clonally reproducing plants. Here, we sampled leaves, rhizomes and roots of three physically and spatially separated ramet sections from a clonally propagated field of the seagrass <em>Zostera marina</em>. Using reduced methylome sequencing, we studied variations in the methylome of seagrass <em>Zostera marina</em> between the sampled tissue types and across age groups. Our analysis of ramets of different ages showed variations in methylation between older and younger samples in both specific methylation patterns and global methylation levels. Our analysis of tissue types showed a marked differentiation of the roots from the rhizomes and leaves, which showed more similar methylation patterns. These findings are in agreement with the strong connection of DNA methylation and plant development and tissue differentiation. We also suggest an effect of differential environmental exposures on the methylome of the younger versus the older ramets due to the forming of molecular stress memories.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1869 2","pages":"Article 130748"},"PeriodicalIF":2.8,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142884785","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
MicroRNAs (miRNAs), which are small non-coding RNAs, are recognized as important significant endogenous bio-molecules that regulate the post-transcriptional processes of target genes. However, predictive methods for significantly working miRNAs are poorly understood. The present study aimed to establish a novel method, miRNA protein analysis of integrative relationship (miR-PAIR), for the identification of effectively working miRNAs involved in physiological or pathological events. To establish the miR-PAIR, comprehensive expression data of miRNAs and proteins were obtained using small RNA-sequence and quantitative proteomics approach in the alveolar epithelial cell line, A549 treated with bleomycin (BLM) and methotrexate (MTX) as pulmonary toxic drugs. Differentially expressed miRNAs and proteins were integrated using TargetScan, a freely available web tool for predicting the target gene of miRNAs. Next, the enrichment of the integrated miRNA-protein pairs was analyzed, followed by the determination of significantly working miRNAs in BLM- and MTX-induced protein expression changes. The miR-PAIR method identified 22 downregulated and 9 upregulated miRNAs. Among them, miR-493-5p (p = 1.71E-05), an upregulated miRNA, suppressed approximately 70 % of the target proteins, and miR-598-3p (p = 1.1E-03), a downregulated miRNA, canceled 50 % of the target protein expression changes induced by BLM and MTX. Thus, a miR-PAIR could be an effective method to identify significantly working miRNAs associated with biological events such as drug-induced lung injury.
{"title":"miR-PAIR: microRNA-protein analysis of integrative relationship for the identification of significantly working miRNAs","authors":"Mizuki Akai , Yuki Maeda , Masashi Kawami , Ryoko Yumoto , Mikihisa Takano , Yasuo Uchida","doi":"10.1016/j.bbagen.2024.130746","DOIUrl":"10.1016/j.bbagen.2024.130746","url":null,"abstract":"<div><div>MicroRNAs (miRNAs), which are small non-coding RNAs, are recognized as important significant endogenous bio-molecules that regulate the post-transcriptional processes of target genes. However, predictive methods for significantly working miRNAs are poorly understood. The present study aimed to establish a novel method, miRNA protein analysis of integrative relationship (miR-PAIR), for the identification of effectively working miRNAs involved in physiological or pathological events. To establish the miR-PAIR, comprehensive expression data of miRNAs and proteins were obtained using small RNA-sequence and quantitative proteomics approach in the alveolar epithelial cell line, A549 treated with bleomycin (BLM) and methotrexate (MTX) as pulmonary toxic drugs. Differentially expressed miRNAs and proteins were integrated using TargetScan, a freely available web tool for predicting the target gene of miRNAs. Next, the enrichment of the integrated miRNA-protein pairs was analyzed, followed by the determination of significantly working miRNAs in BLM- and MTX-induced protein expression changes. The miR-PAIR method identified 22 downregulated and 9 upregulated miRNAs. Among them, miR-493-5p (<em>p</em> = 1.71E-05), an upregulated miRNA, suppressed approximately 70 % of the target proteins, and miR-598-3p (<em>p</em> = 1.1E-03), a downregulated miRNA, canceled 50 % of the target protein expression changes induced by BLM and MTX. Thus, a miR-PAIR could be an effective method to identify significantly working miRNAs associated with biological events such as drug-induced lung injury.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1869 2","pages":"Article 130746"},"PeriodicalIF":2.8,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142871065","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Apolipoprotein E (apoE) polymorphism is associated with different pathologies such as atherosclerosis and Alzheimer's disease. Knowledge of the three-dimensional structure of apoE and isoform-specific structural differences are prerequisites for the rational design of small molecule structure modulators that correct the detrimental effects of pathological isoforms. In this study, cross-linking mass spectrometry (XL-MS) targeting Asp, Glu and Lys residues was used to explore the intramolecular interactions in the E2, E3 and E4 isoforms of apoE. The resulting quantitative XL-MS data combined with molecular modeling revealed isoform-specific characteristics of the N- and C-terminal domain interfaces as well as the isoform-dependent dynamic equilibrium of these interfaces. Finally, the data identified a network of salt bridges formed by R61-R112-E109 residues in the N-terminal helical bundle as a modulator of the interaction with the C-terminal domain making this network a potential drug target.
{"title":"Characterization of the N- and C-terminal domain interface of the three main apoE isoforms: A combined quantitative cross-linking mass spectrometry and molecular modeling study","authors":"Azadeh Mohammadi , Stéphanie Deroo , Alexander Leitner , Florian Stengel , Eva-Maria Krammer , Ruedi Aebersold , Martine Prévost , Vincent Raussens","doi":"10.1016/j.bbagen.2025.130768","DOIUrl":"10.1016/j.bbagen.2025.130768","url":null,"abstract":"<div><div>Apolipoprotein E (apoE) polymorphism is associated with different pathologies such as atherosclerosis and Alzheimer's disease. Knowledge of the three-dimensional structure of apoE and isoform-specific structural differences are prerequisites for the rational design of small molecule structure modulators that correct the detrimental effects of pathological isoforms. In this study, cross-linking mass spectrometry (XL-MS) targeting Asp, Glu and Lys residues was used to explore the intramolecular interactions in the E2, E3 and E4 isoforms of apoE. The resulting quantitative XL-MS data combined with molecular modeling revealed isoform-specific characteristics of the N- and C-terminal domain interfaces as well as the isoform-dependent dynamic equilibrium of these interfaces. Finally, the data identified a network of salt bridges formed by R61-R112-E109 residues in the N-terminal helical bundle as a modulator of the interaction with the C-terminal domain making this network a potential drug target.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1869 4","pages":"Article 130768"},"PeriodicalIF":2.8,"publicationDate":"2025-01-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143063539","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-26DOI: 10.1016/j.bbagen.2025.130770
Sabrina Yamoune , Henner Koch , Daniel Delev , Yvonne Weber , Julia Carolin Stingl
In vitro and ex vivo studies on drug metabolism and stability are vital for drug development and pre-clinical safety assessment. Traditional in vitro models, such as liver enzyme (S9) fractions and microsomes, often fail to account for individual variability. Personalized models, including 3D cell models and organoids, offer promising alternatives but may not fully replicate physiological processes, especially for Cytochrome P450 (CYP) families involved in extrahepatic metabolism. A major challenge in these studies is the low stability and expression of CYP enzymes.
This study aimed to stabilize native CYP activity in vitro by developing an optimized buffer formulation. Initial experiments using recombinant CYP supersomes and liver microsomes identified 45 μM cysteine, 4 mM dithiothreitol (DTT), and 300 μM phosphocholine (PC) as the most effective stabilizers. The applicability of these stabilizers was subsequently confirmed in primary human brain tissue, where they enabled the successful determination of CYP2D6 activity. This highlights the stabilizing buffer's utility for enhancing CYP functionality in diverse tissue types, including the brain, which plays a critical role in cerebral detoxification and drug metabolism.
These findings suggest that specific enzyme stabilization can enable comprehensive evaluations of CYP function in ex vivo tissue samples, advancing the development of organoid human tissue models and supporting drug metabolism research.
{"title":"Evaluation of stabilizing additives to protect activities of cytochrome P450 enzymes for in vitro drug testing and pharmacogenetic studies: Focus on CYP2D6","authors":"Sabrina Yamoune , Henner Koch , Daniel Delev , Yvonne Weber , Julia Carolin Stingl","doi":"10.1016/j.bbagen.2025.130770","DOIUrl":"10.1016/j.bbagen.2025.130770","url":null,"abstract":"<div><div><em>In vitro</em> and <em>ex vivo</em> studies on drug metabolism and stability are vital for drug development and pre-clinical safety assessment. Traditional <em>in vitro</em> models, such as liver enzyme (S9) fractions and microsomes, often fail to account for individual variability. Personalized models, including 3D cell models and organoids, offer promising alternatives but may not fully replicate physiological processes, especially for Cytochrome P450 (CYP) families involved in extrahepatic metabolism. A major challenge in these studies is the low stability and expression of CYP enzymes.</div><div>This study aimed to stabilize native CYP activity <em>in vitro</em> by developing an optimized buffer formulation. Initial experiments using recombinant CYP supersomes and liver microsomes identified 45 μM cysteine, 4 mM dithiothreitol (DTT), and 300 μM phosphocholine (PC) as the most effective stabilizers. The applicability of these stabilizers was subsequently confirmed in primary human brain tissue, where they enabled the successful determination of CYP2D6 activity. This highlights the stabilizing buffer's utility for enhancing CYP functionality in diverse tissue types, including the brain, which plays a critical role in cerebral detoxification and drug metabolism.</div><div>These findings suggest that specific enzyme stabilization can enable comprehensive evaluations of CYP function in <em>ex vivo</em> tissue samples, advancing the development of organoid human tissue models and supporting drug metabolism research.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1869 4","pages":"Article 130770"},"PeriodicalIF":2.8,"publicationDate":"2025-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143057891","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-25DOI: 10.1016/j.bbagen.2025.130769
Yongjia Zheng , Yuxing Lu , Fang Yuan , Yun Kong , Yang Mao , Shengjun Wang
Aberrant glycosylation has been implicated in promoting the progression and metastasis of pancreatic ductal adenocarcinoma (PDAC). However, the contribution of different glycosylation-related genes in PDAC remains to be clarified. In this study, we performed a differential analysis of RNA-Seq data from TCGA and GTEx and found GALNT5 as the most significant upregulated glycosylation-related gene in PDAC. Using publicly available single-cell sequencing data, we further revealed that GALNT5 is predominantly expressed in malignant ductal epithelial cells of PDAC. Correlation analysis indicated that GALNT5 is the essential member of the GALNT family associated with poor prognosis of PDAC. Overexpression of GALNT5 in PANC-1 or MIAPaCa-2 cells with low endogenous GALNT5 enhances migration and invasion. Conversely, knockdown of GALNT5 in AsPC-1 cells with high endogenous GALNT5 inhibits migration and invasion. Mechanistically, we discovered that GALNT5 activates the Erk signaling pathway in PDAC. Our findings suggest GALNT5 is a potential therapeutic target for PDAC.
{"title":"GALNT5 promotes migration and invasion of pancreatic ductal adenocarcinoma cells by activating Erk signaling pathway","authors":"Yongjia Zheng , Yuxing Lu , Fang Yuan , Yun Kong , Yang Mao , Shengjun Wang","doi":"10.1016/j.bbagen.2025.130769","DOIUrl":"10.1016/j.bbagen.2025.130769","url":null,"abstract":"<div><div>Aberrant glycosylation has been implicated in promoting the progression and metastasis of pancreatic ductal adenocarcinoma (PDAC). However, the contribution of different glycosylation-related genes in PDAC remains to be clarified. In this study, we performed a differential analysis of RNA-Seq data from TCGA and GTEx and found GALNT5 as the most significant upregulated glycosylation-related gene in PDAC. Using publicly available single-cell sequencing data, we further revealed that GALNT5 is predominantly expressed in malignant ductal epithelial cells of PDAC. Correlation analysis indicated that GALNT5 is the essential member of the GALNT family associated with poor prognosis of PDAC. Overexpression of GALNT5 in PANC-1 or MIAPaCa-2 cells with low endogenous GALNT5 enhances migration and invasion. Conversely, knockdown of GALNT5 in AsPC-1 cells with high endogenous GALNT5 inhibits migration and invasion. Mechanistically, we discovered that GALNT5 activates the Erk signaling pathway in PDAC. Our findings suggest GALNT5 is a potential therapeutic target for PDAC.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1869 3","pages":"Article 130769"},"PeriodicalIF":2.8,"publicationDate":"2025-01-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143051450","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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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