Biochimica et biophysica acta. Molecular cell research最新文献_第2页

Editorial for the BBA-MCR Special Issue on "Biogenesis and Function Iron-sulfur Proteins". BBA-MCR 特刊 "铁硫蛋白的生物生成和功能 "编辑。

IF 4.6 2区生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Molecular cell research

Pub Date : 2024-10-24 DOI: 10.1016/j.bbamcr.2024.119867

Roland Lill, Frederic Barras

引用次数: 0

Involvement of aquaporins in Shiga toxin-induced swelling and water transport dysfunction in human renal microvascular endothelial cells 水蒸发蛋白参与滋贺毒素诱导的人肾微血管内皮细胞肿胀和水转运功能障碍。

IF 4.6 2区生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Molecular cell research

Pub Date : 2024-10-21 DOI: 10.1016/j.bbamcr.2024.119866

Fernando D. Gomez , Julieta Reppetti , Romina S. Alvarez , Daniel C. Girón Reyes , Flavia Sacerdoti , Alejandro Balestracci , Alicia E. Damiano , Nora A. Martínez , Gisela Di Giusto , María M. Amaral

One of the hallmarks of Shiga toxin-producing Escherichia coli-associated hemolytic uremic syndrome (STEC-HUS) is kidney damage. Our previous research demonstrated that Shiga toxin type 2 (Stx2a) decreases cell viability and induces swelling of human glomerular endothelial cells (HGEC). However, Stx2a can disrupt net water transport across HGEC monolayers without affecting cell viability. This work aimed to elucidate the possible mechanisms involved in the water transport disruption caused by Stx2a across HGEC monolayers. We investigated paracellular and transcellular water transfer across HGEC by analyzing the passage of FITC-Dextran and the hydrostatic pressure (Phydr) and measuring the osmotic pressure (Posm), respectively. Stx2a selectively affected the transcellular pathway without impacting the paracellular route. Furthermore, Stx2a cell swelling was prevented by pretreatment with aquaporin inhibitors tetraethylammonium chloride (TEA), Mercury (II) chloride (HgCl₂) or TGN-020, suggesting aquaporin involvement in this process. Confocal microscopy revealed that Stx2a increased HGEC total volume, which TEA and TGN-020 counteracted. Additionally, we identified in HGEC not only the expression of aquaporin-1 (AQP1) but also the expression of aquaporin-4 (AQP4). Surprisingly, we observed a decrease in the expression of both AQPs after Stx2a exposure. Our findings suggest that Stx2a may induce water movement into HGEC via AQP1 and AQP4, increasing total cell volume. Subsequently, decreased AQP1 and AQP4 expression could inhibit transcellular water transfer, potentially as a protective mechanism against excessive water entry and cell lysis.

产志贺毒素大肠埃希菌相关溶血性尿毒症（STEC-HUS）的特征之一是肾脏损伤。我们之前的研究表明，2 型志贺毒素（Stx2a）会降低细胞活力，并诱导人肾小球内皮细胞（HGEC）肿胀。然而，Stx2a 能破坏 HGEC 单层细胞的净水运输，而不影响细胞活力。这项研究旨在阐明 Stx2a 在 HGEC 单层细胞上造成水转运中断的可能机制。我们通过分析 FITC-二聚体的通过量、静水压（Pydr）和渗透压（Posm），分别研究了 HGEC 的胞旁和跨细胞水转运。Stx2a 选择性地影响跨细胞途径，而不影响旁细胞途径。此外，在使用水蒸发蛋白抑制剂四乙基氯化铵（TEA）、氯化汞（HgCl2）或 TGN-020 进行预处理后，Stx2a 细胞肿胀也会被阻止，这表明水蒸发蛋白参与了这一过程。共聚焦显微镜显示，Stx2a 增加了 HGEC 的总体积，而 TEA 和 TGN-020 则抵消了这一作用。此外，我们在 HGEC 中不仅发现了水通道蛋白-1（AQP1）的表达，还发现了水通道蛋白-4（AQP4）的表达。令人惊讶的是，在接触 Stx2a 后，我们观察到这两种 AQPs 的表达都有所下降。我们的研究结果表明，Stx2a 可能会诱导水分通过 AQP1 和 AQP4 进入 HGEC，从而增加细胞的总体积。随后，AQP1 和 AQP4 表达的减少可能会抑制跨细胞的水分转移，这可能是防止水分过度进入和细胞溶解的一种保护机制。

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引用次数: 0

Synergy of native mass spectrometry and other biophysical techniques in studies of iron‑sulfur cluster proteins and their assembly 本机质谱和其他生物物理技术在铁硫簇蛋白质及其组装研究中的协同作用。

IF 4.6 2区生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Molecular cell research

Pub Date : 2024-10-21 DOI: 10.1016/j.bbamcr.2024.119865

Jason C. Crack, Nick E. Le Brun

The application of mass spectrometric methodologies has revolutionised biological chemistry, from identification through to structural and conformational studies of proteins and other macromolecules. Native mass spectrometry (MS), in which proteins retain their native structure, is a rapidly growing field. This is particularly the case for studies of metalloproteins, where non-covalently bound cofactors remain bound following ionisation. Such metalloproteins include those that contain an iron‑sulfur (FeS) cluster and, despite their fragility and O₂ sensitivity, they have been a particular focus for applications of native MS because of its capacity to accurately monitor mass changes that reveal chemical changes at the cluster. Here we review recent advances in these applications of native MS, which, together with data from more traditionally applied biophysical methods, have yielded a remarkable breadth of information about the FeS species present, and provided key mechanistic insight not only for FeS cluster proteins themselves, but also their assembly.

从蛋白质和其他大分子的鉴定到结构和构象研究，质谱方法的应用彻底改变了生物化学。保持蛋白质原生结构的原生质谱（MS）是一个发展迅速的领域。尤其是在研究金属蛋白时，非共价结合的辅助因子在电离后仍保持结合。此类金属蛋白包括那些含有铁硫（FeS）簇的金属蛋白，尽管它们很脆弱且对氧气很敏感，但它们一直是原生质谱应用的重点，因为质谱能够准确监测质量变化，从而揭示簇的化学变化。在此，我们回顾了本机质谱应用的最新进展，这些进展与更多传统生物物理方法的应用数据一起，产生了有关 FeS 物种的大量信息，不仅为 FeS 簇蛋白质本身，也为它们的组装提供了关键的机理认识。

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引用次数: 0

IL-17A's role in exacerbating radiation-induced lung injury: Autophagy impairment via the PP2A-mTOR pathway IL-17A 在加剧辐射诱导的肺损伤中的作用：通过 PP2A-mTOR 通路的自噬损伤

IF 4.6 2区生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Molecular cell research

Pub Date : 2024-10-20 DOI: 10.1016/j.bbamcr.2024.119864

Liangzhong Liu , GuangMing Yi , Xiaohong Li , Cai Chen , Kehong Chen , Hengqiu He , Jinjin Li , Fanghao Cai , Yuan Peng , Zhenzhou Yang , Xiaoyue Zhang

Objective

Radiation-induced lung injury (RILI) is a serious complication of radiotherapy, and the role of IL-17A in this process is not well understood. While IL-17A has been shown to modulate autophagy, conflicting reports exist regarding its activation or inhibition of autophagy. This study investigates the role of IL-17A in RILI and its effects on autophagy via the PP2A-mTOR pathway, with a focus on the PP2A B56α subunit.

Methods

C57BL/6J mice and human lung epithelial cells (BEAS-2B) were exposed to radiation with or without recombinant IL-17A. Autophagy markers were analyzed using Western blotting, immunofluorescence, and autophagy flux assays. PP2A activity, specifically the B56α subunit, was measured. A PP2A agonist (DT-061) was used to verify its role in reversing IL-17A-mediated autophagy inhibition.

Results

IL-17A inhibited autophagy in lung epithelial cells exposed to radiation by suppressing PP2A activity, particularly through downregulation of the B56α subunit, leading to mTOR activation and reduced autophagosome formation. Treatment with DT-061 restored autophagic activity and improved cell viability. These findings align with reports suggesting that IL-17A inhibits autophagy in certain contexts, while other studies have shown opposing effects.

Conclusion

IL-17A inhibits autophagy in RILI through the PP2A B56α-mTOR pathway, exacerbating lung damage. Further research is needed to clarify the role of IL-17A in different cell types and conditions. Targeting the IL-17A-PP2A B56α-mTOR axis may offer new therapeutic strategies for RILI management.

目的：放疗引起的肺损伤（RILI）是放疗的一种严重并发症，而 IL-17A 在这一过程中的作用尚不十分清楚。虽然已证明 IL-17A 可调节自噬，但关于其激活或抑制自噬的报道却相互矛盾。本研究探讨了 IL-17A 在 RILI 中的作用及其通过 PP2A-mTOR 途径对自噬的影响，重点是 PP2A B56α 亚基：方法：将 C57BL/6J 小鼠和人肺上皮细胞（BEAS-2B）暴露于含有或不含重组 IL-17A 的辐射中。使用 Western 印迹、免疫荧光和自噬通量测定分析自噬标记物。测定了 PP2A 的活性，特别是 B56α 亚基。使用 PP2A 激动剂（DT-061）验证其在逆转 IL-17A 介导的自噬抑制中的作用：结果：IL-17A通过抑制PP2A的活性，特别是通过下调B56α亚基，导致mTOR激活和自噬体形成减少，从而抑制了暴露于辐射的肺上皮细胞的自噬。使用 DT-061 治疗可恢复自噬活性并提高细胞活力。这些发现与IL-17A在某些情况下抑制自噬的报道一致，而其他研究则显示了相反的作用：结论：IL-17A通过PP2A B56α-mTOR通路抑制RILI中的自噬，加剧肺损伤。要明确IL-17A在不同细胞类型和条件下的作用，还需要进一步的研究。以IL-17A-PP2A B56α-mTOR轴为靶点可能会为RILI的治疗提供新的治疗策略。

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引用次数: 0

Mitoception, or transfer of normal cell mitochondria to cancer cells, reverses remodeling of store-operated Ca2+ entry in tumor cells 线粒体接收，或将正常细胞线粒体转移到癌细胞中，可逆转肿瘤细胞中储存操作的 Ca2+ 进入的重塑。

IF 4.6 2区生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Molecular cell research

Pub Date : 2024-10-20 DOI: 10.1016/j.bbamcr.2024.119862

Verónica Feijóo , Sendoa Tajada , Alejandra Méndez-Mena , Lucía Núñez , Carlos Villalobos

Most cancer cells show the Warburg effect, the rewiring of aerobic metabolism to glycolysis due to defective mitochondrial ATP synthesis. As a consequence, tumor cells display enhanced mitochondrial potential (∆Ψ), the driving force for mitochondrial Ca²⁺ uptake. Mitochondria control the Ca²⁺-dependent inactivation of store-operated channels (SOCs), leading to enhanced and sustained store-operated Ca²⁺ entry (SOCE) involved in cancer hallmarks. We asked here whether the transfer of mitochondria (mitoception) from normal cells to tumor cells may reverse SOCE remodeling in cancer cells. For this end, we labeled mitochondria in normal NCM460 human colonic cells, isolated them and transferred them to tumor HT29 cells. We tested the viability and efficiency of mitoception using flow cytometry and confocal microscopy, as well as calcium imaging to investigate the effects of mitoception on SOCE. Our results show that mitoception of tumor HT29 cells with normal mitochondria restores a low ∆Ψ and SOCE. Conversely, self-mitoception of tumor HT29 cells with tumor cell mitochondria increases further ∆Ψ and SOCE, thus excluding the possibility that effects of mitoception are due to increased mitochondrial mass. Strikingly, mitoception of normal NCM460 cells with tumor cell mitochondria has no effects on either ∆Ψ or SOCE. These results are consistent with the previous proposal that transformed mitochondria may modulate SOC channels involved in SOCE. Further research is warranted to test whether mitoception of cancer cells with normal mitochondria may reverse Ca²⁺ remodeling associated to cancer.

大多数癌细胞都表现出沃伯格效应，即由于线粒体 ATP 合成缺陷，有氧代谢被重新安排为糖酵解。因此，肿瘤细胞的线粒体电位（ΔΨ）增强，这是线粒体摄取 Ca2+ 的驱动力。线粒体控制着贮存操作通道（SOC）的 Ca2+ 失活，从而导致贮存操作 Ca2+ 进入（SOCE）的增强和持续，这与癌症的特征有关。我们在此提出的问题是，将线粒体从正常细胞转移到肿瘤细胞（线粒体接收）是否会逆转癌细胞中的 SOCE 重塑。为此，我们标记了正常 NCM460 人结肠细胞中的线粒体，将其分离并转移到肿瘤 HT29 细胞中。我们利用流式细胞术和共聚焦显微镜检测了线粒体的活力和接收效率，并通过钙成像研究了线粒体接收对 SOCE 的影响。我们的结果表明，线粒体正常的肿瘤 HT29 细胞的线粒体接收可恢复较低的ΔΨ和 SOCE。相反，具有肿瘤细胞线粒体的肿瘤 HT29 细胞的自我线粒体感知进一步增加了ΔΨ 和 SOCE，从而排除了线粒体感知的影响是由于线粒体质量增加的可能性。令人吃惊的是，用肿瘤细胞线粒体诱导正常 NCM460 细胞对 ∆Ψ 和 SOCE 都没有影响。这些结果与之前的提议一致，即转化线粒体可能会调节参与 SOCE 的 SOC 通道。还需要进一步研究，以检验用正常线粒体诱导癌细胞是否可以逆转与癌症相关的 Ca2+ 重塑。

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引用次数: 0

Interleukin-2 receptor α (IL-2Rα/CD25) shedding is differentially regulated by N- and O-glycosylation 白细胞介素-2受体α（IL-2Rα/CD25）脱落受N-和O-糖基化的不同调节。

IF 4.6 2区生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Molecular cell research

Pub Date : 2024-10-19 DOI: 10.1016/j.bbamcr.2024.119863

Amelie Franke , Sophia Dahl , Monika Funck , Hans Bakker , Christoph Garbers , Juliane Lokau

The cytokine interleukin-2 (IL-2) is a critical regulator of immune responses, with an especially well-characterized role in regulating T-cell homeostasis. IL-2 signaling involves three distinct receptor subunits: the IL-2Rα (CD25), IL-2Rβ, and IL-2Rγ. The intracellular transduction of IL-2-induced signals is strictly dependent on IL-2Rβ and IL-2Rγ, while the IL-2Rα is not directly involved in signaling. Instead, it has the highest affinity towards IL-2 and is thus responsible for regulating the affinity of a cell for IL-2. In addition to the membrane-bound IL-2Rα, a soluble form of the receptor (sIL-2Rα) has been described, which is present in the blood of healthy individuals, increased under various pathological conditions, and able to bind IL-2 and thus modulate its function. The sIL-2Rα is generated by proteolytic cleavage of the membrane-bound receptor. Here, we analyze whether glycosylation of the IL-2Rα regulates its proteolysis. We find that constitutive IL-2Rα shedding is affected by glycosylation and discover distinct roles for N- and O-glycosylation. Furthermore, we show that induced shedding by the metalloproteases ADAM10 and ADAM17 is also differentially regulated by distinct types of glycans. Finally, we identify a specific role for an N-glycan at an exosite in ADAM17-mediated proteolysis that does not affect ADAM10, indicating distinct substrate recognition mechanisms. These results further the understanding of the mechanisms leading to sIL-2Rα generation, and thus offer the opportunity to specifically modulate the generation of the soluble receptor.

细胞因子白细胞介素-2（IL-2）是免疫反应的关键调节因子，在调节 T 细胞稳态方面的作用尤其明显。IL-2 信号转导涉及三种不同的受体亚基：IL-2Rα（CD25）、IL-2Rβ 和 IL-2Rγ。IL-2 诱导的细胞内信号转导严格依赖于 IL-2Rβ 和 IL-2Rγ，而 IL-2Rα 并不直接参与信号转导。相反，它对 IL-2 的亲和力最高，因此负责调节细胞对 IL-2 的亲和力。除了膜结合型 IL-2Rα，还有一种可溶性形式的受体（sIL-2Rα）被描述出来，这种受体存在于健康人的血液中，在各种病理情况下会增加，能够结合 IL-2，从而调节其功能。sIL-2Rα是由膜结合受体蛋白水解产生的。在这里，我们分析了 IL-2Rα 的糖基化是否会调节其蛋白水解。我们发现组成型 IL-2Rα 脱落受糖基化的影响，并发现 N 型和 O 型糖基化的不同作用。此外，我们还发现金属蛋白酶 ADAM10 和 ADAM17 诱导的脱落也受到不同类型糖基的不同调控。最后，我们发现在 ADAM17 介导的蛋白水解过程中，外切物上的 N-聚糖起着特殊作用，而 ADAM10 却不受影响，这表明它们有不同的底物识别机制。这些结果进一步加深了人们对 sIL-2Rα 生成机制的理解，从而为特异性调节可溶性受体的生成提供了机会。

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引用次数: 0

The special issue of the ECS2022 meeting in Cork, Ireland. 在爱尔兰科克举行的 ECS2022 会议特刊。

IF 4.6 2区生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Molecular cell research

Pub Date : 2024-10-10 DOI: 10.1016/j.bbamcr.2024.119861

Geert Bultynck, Björn-Philipp Diercks, Enikö Kallay, John Mackrill

引用次数: 0

USP7 facilitates deubiquitination of LRRC42 in colorectal cancer to accelerate tumorigenesis and augment Wnt/β-catenin signaling USP7 在结直肠癌中促进 LRRC42 的去泛素化，从而加速肿瘤发生并增强 Wnt/β-catenin 信号传导。

IF 4.6 2区生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Molecular cell research

Pub Date : 2024-10-09 DOI: 10.1016/j.bbamcr.2024.119859

Yunze Li , Xin Sun , Zhe Huang

Colorectal cancer is a prevalent malignancy with an increasing incidence worldwide. Leucine-rich repeat-containing protein 42 (LRRC42) is known to be dysregulated in tumor tissues, yet its role in colorectal cancer remains largely unexplored. Herein, the function of LRRC42 in colorectal cancer was investigated using clinical samples, cellular experiments, animal models, and multiple omics techniques. The results demonstrated that LRRC42 was highly expressed in colorectal cancer tissues and was associated with poor clinical outcomes. Silencing LRRC42 suppressed cell proliferation, induced G0/G1 phase arrest, and promoted apoptosis by reducing Bcl2 expression while elevating the expression of Bax, cleaved PARP and cleaved caspase 3. Conversely, LRRC42 overexpression exhibited the opposite effects. Consistent findings were observed in vivo. Additionally, ubiquitin specific peptidase 7 was identified as a potential LRRC42-interacting protein through immunoprecipitation-mass spectrometry, with ubiquitin specific peptidase 7 stabilizing LRRC42 expression by promoting its deubiquitination. Notably, LRRC42 overexpression partially reversed the effects of ubiquitin specific peptidase 7 silencing on tumor cell proliferation and apoptosis. mRNA sequencing analysis revealed that differentially expressed genes in LRRC42 overexpressing cells were linked to Wnt signaling pathway, suggesting that LRRC42 overexpression may activate this pathway. Furthermore, LRRC42 was proved to elevate the levels of ki67, cyclin D1 and WNT3, while reducing the level of p-β-catenin. These findings suggest that LRRC42 perhaps serve as a potential oncogenic factor in colorectal cancer, regulated by ubiquitin specific peptidase 7 and capable of activating Wnt/β-catenin signaling pathway.

结直肠癌是一种常见的恶性肿瘤，在全球的发病率不断上升。众所周知，富亮氨酸重复序列蛋白 42（LRRC42）在肿瘤组织中调控失调，但其在结直肠癌中的作用仍未得到充分探究。在此，研究人员利用临床样本、细胞实验、动物模型和多种全息技术研究了LRRC42在结直肠癌中的功能。结果表明，LRRC42在结直肠癌组织中高表达，并与不良临床预后相关。沉默 LRRC42 可抑制细胞增殖，诱导 G0/G1 期停滞，并通过降低 Bcl2 表达，同时提高 Bax、裂解 PARP 和裂解 Caspase 3 的表达来促进细胞凋亡。相反，LRRC42 过表达则表现出相反的效果。在体内也观察到了一致的结果。此外，通过免疫沉淀-质谱分析，泛素特异性肽酶 7 被确定为潜在的 LRRC42 相互作用蛋白，泛素特异性肽酶 7 通过促进 LRRC42 的去泛素化来稳定其表达。mRNA测序分析发现，LRRC42过表达细胞中的差异表达基因与Wnt信号通路有关，表明LRRC42的过表达可能激活了这一通路。此外，LRRC42 还能提高 ki67、细胞周期蛋白 D1 和 WNT3 的水平，同时降低 p-β-catenin 的水平。这些研究结果表明，LRRC42 可能是结直肠癌的潜在致癌因子，它受泛素特异性肽酶 7 的调控，能够激活 Wnt/β-catenin 信号通路。

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引用次数: 0

Atrial natriuretic peptide (ANP) modulates stress-induced autophagy in endothelial cells 心房利钠肽（ANP）可调节内皮细胞中由压力诱导的自噬。

IF 4.6 2区生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Molecular cell research

Pub Date : 2024-10-09 DOI: 10.1016/j.bbamcr.2024.119860

Maurizio Forte , Simona Marchitti , Flavio di Nonno , Donatella Pietrangelo , Rosita Stanzione , Maria Cotugno , Luca D'Ambrosio , Alessandra D'Amico , Vittoria Cammisotto , Gianmarco Sarto , Erica Rocco , Beatrice Simeone , Sonia Schiavon , Daniele Vecchio , Roberto Carnevale , Salvatore Raffa , Giacomo Frati , Massimo Volpe , Sebastiano Sciarretta , Speranza Rubattu

Atrial natriuretic peptide (ANP), a cardiac hormone involved in the regulation of water/sodium balance and blood pressure, is also secreted by endothelial cells, where it exerts protective effects in response to stress. Autophagy is an intracellular self-renewal process involved in the degradation of dysfunctional cytoplasmic elements. ANP was recently reported to act as an extracellular regulator of cardiac autophagy. However, its role in the regulation of endothelial autophagy has never been investigated. Here, we tested the effects of ANP in the regulation of autophagy in human umbilical vein endothelial cells (HUVECs). We found that ANP rapidly increases autophagy and autophagic flux at physiological concentrations through its predominant pathway, mediated by natriuretic peptide receptor type A (NPR-A) and protein kinase G (PKG). We further observed that ANP is rapidly secreted by HUVEC under stress conditions, where it mediates stress-induced autophagy through autocrine and paracrine mechanisms. Finally, we found that the protective effects of ANP in response to high-salt loading or tumor necrosis factor (TNF)-α are blunted by concomitant inhibition of autophagy. Overall, our results suggest that ANP acts as an endogenous autophagy activator in endothelial cells. The autophagy mechanism mediates the protective endothelial effects exerted by ANP.

心房利钠肽（ANP）是一种参与调节水/钠平衡和血压的心脏激素，它也由内皮细胞分泌，在应对压力时发挥保护作用。自噬是一种细胞内自我更新过程，参与降解功能失调的细胞质元素。最近有报道称，ANP 是心脏自噬的细胞外调节因子。然而，它在调节内皮自噬中的作用却从未被研究过。在这里，我们测试了 ANP 在调节人脐静脉内皮细胞（HUVECs）自噬中的作用。我们发现，在生理浓度下，ANP 通过其主要途径迅速增加自噬和自噬通量，该途径由钠肽受体 A 型（NPR-A）和蛋白激酶 G（PKG）介导。我们进一步观察到，HUVEC 在应激条件下会快速分泌 ANP，ANP 通过自分泌和旁分泌机制介导应激诱导的自噬。最后，我们发现 ANP 对高盐负荷或肿瘤坏死因子（TNF）-α 的保护作用会因同时抑制自噬而减弱。总之，我们的研究结果表明，ANP 是内皮细胞中的内源性自噬激活剂。自噬机制介导了 ANP 对内皮细胞的保护作用。

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引用次数: 0

Structural analysis of microtubule binding by minus-end targeting protein Spiral2 负端靶向蛋白 Spiral2 与微管结合的结构分析

IF 4.6 2区生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Molecular cell research

Pub Date : 2024-10-04 DOI: 10.1016/j.bbamcr.2024.119858

Marina Ohno , Yuuki Higuchi , Kazune Yamai , Sotaro Fuchigami , Takema Sasaki , Yoshihisa Oda , Ikuko Hayashi

Microtubules (MTs) are dynamic cytoskeletal polymers that play a critical role in determining cell polarity and shape. In plant cells, acentrosomal MTs are localized on the cell surface and are referred to as cortical MTs. Cortical MTs nucleate in the cell cortex and detach from nucleation sites. The released MT filaments perform treadmilling, with the plus-ends of MTs polymerizing and the minus-ends depolymerizing. Minus-end targeting proteins, -TIPs, include Spiral2, which regulates the minus-end dynamics of acentrosomal MTs. Spiral2 accumulates autonomously at MT minus-ends and inhibits filament shrinkage, but the mechanism by which Spiral2 specifically recognizes minus-ends of MTs remains unknown. Here we describe the crystal structure of Spiral2's N-terminal MT-binding domain. The structural properties of this domain resemble those of the HEAT repeat structure of the tumor overexpressed gene (TOG) domain, but the number of HEAT repeats is different and the conformation is highly arched. Gel filtration and co-sedimentation analyses demonstrate that the domain binds preferentially to MT filaments rather than the tubulin dimer, and that the tubulin-binding mode of Spiral2 via the basic surface is similar to that of the TOG domain. We constructed an in silico model of the Spiral2-tubulin complex to identify residues that potentially recognize tubulin. Mutational analysis revealed that the key residues inferred in the model are involved in microtubule recognition, and provide insight into the mechanism by which end-targeting proteins stabilize MT ends.

微管（MT）是一种动态细胞骨架聚合物，在决定细胞极性和形状方面起着至关重要的作用。在植物细胞中，顶体 MT 定位于细胞表面，被称为皮层 MT。皮层 MT 在细胞皮层成核，并从成核点分离。释放出来的MT丝进行踩踏运动，MT的正端聚合，负端解聚。负端靶向蛋白（-TIPs）包括螺旋2（Spiral2），它能调节顶体MT的负端动态。Spiral2 在 MT 负端自主聚集并抑制纤维收缩，但 Spiral2 特异性识别 MT 负端的机制仍不清楚。在这里，我们描述了 Spiral2 N 端 MT 结合结构域的晶体结构。该结构域的结构特性与肿瘤过表达基因（TOG）结构域的 HEAT 重复结构相似，但 HEAT 重复的数量不同，而且构象呈高度弧形。凝胶过滤和共沉淀分析表明，该结构域优先与 MT 细丝结合，而不是与微管蛋白二聚体结合，而且 Spiral2 通过基本表面与微管蛋白结合的模式与 TOG 结构域类似。我们构建了一个Spiral2-微管蛋白复合物的硅学模型，以确定可能识别微管蛋白的残基。突变分析表明，模型中推断出的关键残基参与了微管识别，并为末端靶向蛋白稳定MT末端的机制提供了启示。

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引用次数: 0