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Connexin43 promotes exocytosis of damaged lysosomes through actin remodelling. Connexin43通过肌动蛋白重塑促进受损溶酶体的外排。
IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-01 Epub Date: 2024-07-23 DOI: 10.1038/s44318-024-00177-3
Neuza Domingues, Steve Catarino, Beatriz Cristóvão, Lisa Rodrigues, Filomena A Carvalho, Maria João Sarmento, Mónica Zuzarte, Jani Almeida, Teresa Ribeiro-Rodrigues, Ânia Correia-Rodrigues, Fábio Fernandes, Paulo Rodrigues-Santos, Trond Aasen, Nuno C Santos, Viktor I Korolchuk, Teresa Gonçalves, Ira Milosevic, Nuno Raimundo, Henrique Girão

A robust and efficient cellular response to lysosomal membrane damage prevents leakage from the lysosome lumen into the cytoplasm. This response is understood to happen through either lysosomal membrane repair or lysophagy. Here we report exocytosis as a third response mechanism to lysosomal damage, which is further potentiated when membrane repair or lysosomal degradation mechanisms are impaired. We show that Connexin43 (Cx43), a protein canonically associated with gap junctions, is recruited from the plasma membrane to damaged lysosomes, promoting their secretion and accelerating cell recovery. The effects of Cx43 on lysosome exocytosis are mediated by a reorganization of the actin cytoskeleton that increases plasma membrane fluidity and decreases cell stiffness. Furthermore, we demonstrate that Cx43 interacts with the actin nucleator Arp2, the activity of which was shown to be necessary for Cx43-mediated actin rearrangement and lysosomal exocytosis following damage. These results define a novel mechanism of lysosomal quality control whereby Cx43-mediated actin remodelling potentiates the secretion of damaged lysosomes.

细胞对溶酶体膜损伤的强大而有效的反应可以防止溶酶体从腔内渗漏到细胞质中。据了解,这种反应是通过溶酶体膜修复或溶酶体吞噬发生的。在这里,我们报告了溶酶体损伤的第三种反应机制--外渗,当溶酶体膜修复或溶酶体降解机制受损时,外渗会进一步增强。我们发现,Connexin43(Cx43)是一种与缝隙连接相关的蛋白质,它能从质膜被招募到受损的溶酶体,促进溶酶体分泌并加速细胞恢复。Cx43 对溶酶体外泌的影响是由肌动蛋白细胞骨架的重组介导的,这种重组增加了质膜的流动性并降低了细胞的硬度。此外,我们还证明了 Cx43 与肌动蛋白成核因子 Arp2 的相互作用,而 Arp2 的活性被证明是 Cx43 介导的肌动蛋白重排和损伤后溶酶体外排所必需的。这些结果确定了一种新的溶酶体质量控制机制,即 Cx43 介导的肌动蛋白重排可促进受损溶酶体的分泌。
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引用次数: 0
Real-time assessment of mitochondrial DNA heteroplasmy dynamics at the single-cell level. 在单细胞水平上实时评估线粒体 DNA 异构动态。
IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-08-05 DOI: 10.1038/s44318-024-00183-5
Rodaria Roussou, Dirk Metzler, Francesco Padovani, Felix Thoma, Rebecca Schwarz, Boris Shraiman, Kurt M Schmoller, Christof Osman

Mitochondrial DNA (mtDNA) is present in multiple copies within cells and is required for mitochondrial ATP generation. Even within individual cells, mtDNA copies can differ in their sequence, a state known as heteroplasmy. The principles underlying dynamic changes in the degree of heteroplasmy remain incompletely understood, due to the inability to monitor this phenomenon in real time. Here, we employ mtDNA-based fluorescent markers, microfluidics, and automated cell tracking, to follow mtDNA variants in live heteroplasmic yeast populations at the single-cell level. This approach, in combination with direct mtDNA tracking and data-driven mathematical modeling reveals asymmetric partitioning of mtDNA copies during cell division, as well as limited mitochondrial fusion and fission frequencies, as critical driving forces for mtDNA variant segregation. Given that our approach also facilitates assessment of segregation between intact and mutant mtDNA, we anticipate that it will be instrumental in elucidating the mechanisms underlying the purifying selection of mtDNA.

线粒体 DNA(mtDNA)在细胞内有多个拷贝,是线粒体 ATP 生成所必需的。即使在单个细胞内,mtDNA拷贝的序列也可能不同,这种状态被称为异质性。由于无法对这一现象进行实时监测,人们对异质性程度动态变化的基本原理仍然不甚了解。在这里,我们采用基于 mtDNA 的荧光标记、微流控技术和自动细胞追踪技术,在单细胞水平上追踪活的异质酵母群体中的 mtDNA 变异。这种方法与直接 mtDNA 跟踪和数据驱动的数学建模相结合,揭示了细胞分裂过程中 mtDNA 拷贝的非对称分割,以及线粒体融合和分裂频率的限制,是 mtDNA 变异分离的关键驱动力。鉴于我们的方法还有助于评估完整 mtDNA 和突变 mtDNA 之间的分离,我们预计它将有助于阐明 mtDNA 纯化选择的内在机制。
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引用次数: 0
The kinesin-3 KIF1C undergoes liquid-liquid phase separation for accumulation of specific transcripts at the cell periphery. 驱动蛋白-3 KIF1C 经过液-液相分离,在细胞外围积累特定的转录本。
IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-08-01 Epub Date: 2024-06-19 DOI: 10.1038/s44318-024-00147-9
Qi Geng, Jakia Jannat Keya, Takashi Hotta, Kristen J Verhey

In cells, mRNAs are transported to and positioned at subcellular areas to locally regulate protein production. Recent studies have identified the kinesin-3 family member motor protein KIF1C as an RNA transporter. However, it is not clear how KIF1C interacts with RNA molecules. Here, we show that the KIF1C C-terminal tail domain contains an intrinsically disordered region (IDR) that drives liquid-liquid phase separation (LLPS). KIF1C forms dynamic puncta in cells that display physical properties of liquid condensates and incorporate RNA molecules in a sequence-selective manner. Endogenous KIF1C forms condensates in cellular protrusions, where mRNAs are enriched in an IDR-dependent manner. Purified KIF1C tail constructs undergo LLPS in vitro at near-endogenous nM concentrations and in the absence of crowding agents and can directly recruit RNA molecules. Overall, our work uncovers an intrinsic correlation between the LLPS activity of KIF1C and its role in mRNA positioning. In addition, the LLPS activity of KIF1C's tail represents a new mode of motor-cargo interaction that extends our current understanding of cytoskeletal motor proteins.

在细胞中,mRNA 被运输到亚细胞区域并被定位,以局部调节蛋白质的产生。最近的研究发现,驱动蛋白-3 家族的运动蛋白 KIF1C 是一种 RNA 转运体。然而,KIF1C 如何与 RNA 分子相互作用尚不清楚。在这里,我们发现 KIF1C C 端尾部结构域包含一个本征无序区(IDR),它能驱动液-液相分离(LLPS)。KIF1C 在细胞中形成动态点,显示出液体凝结物的物理特性,并以序列选择性的方式结合 RNA 分子。内源性 KIF1C 在细胞突起中形成凝聚体,其中的 mRNA 以 IDR 依赖性方式富集。纯化的 KIF1C 尾部构建体在体外以接近内源性 nM 的浓度并在没有拥挤剂的情况下进行 LLPS,并能直接招募 RNA 分子。总之,我们的工作发现了 KIF1C 的 LLPS 活性与其在 mRNA 定位中的作用之间的内在联系。此外,KIF1C 尾部的 LLPS 活性代表了电机与货物相互作用的一种新模式,拓展了我们目前对细胞骨架电机蛋白的认识。
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引用次数: 0
Targeting neuronal epigenomes for brain rejuvenation. 以神经元表观基因组为目标,实现大脑年轻化。
IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-08-01 Epub Date: 2024-07-15 DOI: 10.1038/s44318-024-00148-8
Sara Zocher

Aging is associated with a progressive decline of brain function, and the underlying causes and possible interventions to prevent this cognitive decline have been the focus of intense investigation. The maintenance of neuronal function over the lifespan requires proper epigenetic regulation, and accumulating evidence suggests that the deterioration of the neuronal epigenetic landscape contributes to brain dysfunction during aging. Epigenetic aging of neurons may, however, be malleable. Recent reports have shown age-related epigenetic changes in neurons to be reversible and targetable by rejuvenation strategies that can restore brain function during aging. This review discusses the current evidence that identifies neuronal epigenetic aging as a driver of cognitive decline and a promising target of brain rejuvenation strategies, and it highlights potential approaches for the specific manipulation of the aging neuronal epigenome to restore a youthful epigenetic state in the brain.

衰老与大脑功能的逐渐衰退有关,而防止认知功能衰退的根本原因和可能的干预措施一直是人们研究的重点。神经元功能在整个生命周期中的维持需要适当的表观遗传调控,越来越多的证据表明,神经元表观遗传景观的恶化导致了衰老过程中的大脑功能障碍。然而,神经元的表观遗传老化可能是可塑的。最近的报告显示,神经元中与年龄相关的表观遗传变化是可逆的,并且可以通过恢复活力策略进行靶向治疗,从而在衰老过程中恢复大脑功能。本综述讨论了目前有证据表明神经元表观遗传老化是认知能力下降的一个驱动因素,也是大脑年轻化策略的一个有希望的目标,并重点介绍了具体操纵老化神经元表观遗传组以恢复大脑年轻表观遗传状态的潜在方法。
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引用次数: 0
Multi-signal regulation of the GSK-3β homolog Rim11 controls meiosis entry in budding yeast. GSK-3β 同源物 Rim11 的多信号调控控制着芽殖酵母的减数分裂过程。
IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-08-01 Epub Date: 2024-06-17 DOI: 10.1038/s44318-024-00149-7
Johanna Kociemba, Andreas Christ Sølvsten Jørgensen, Nika Tadić, Anthony Harris, Theodora Sideri, Wei Yee Chan, Fairouz Ibrahim, Elçin Ünal, Mark Skehel, Vahid Shahrezaei, Orlando Argüello-Miranda, Folkert Jacobus van Werven

Starvation in diploid budding yeast cells triggers a cell-fate program culminating in meiosis and spore formation. Transcriptional activation of early meiotic genes (EMGs) hinges on the master regulator Ime1, its DNA-binding partner Ume6, and GSK-3β kinase Rim11. Phosphorylation of Ume6 by Rim11 is required for EMG activation. We report here that Rim11 functions as the central signal integrator for controlling Ume6 phosphorylation and EMG transcription. In nutrient-rich conditions, PKA suppresses Rim11 levels, while TORC1 retains Rim11 in the cytoplasm. Inhibition of PKA and TORC1 induces Rim11 expression and nuclear localization. Remarkably, nuclear Rim11 is required, but not sufficient, for Rim11-dependent Ume6 phosphorylation. In addition, Ime1 is an anchor protein enabling Ume6 phosphorylation by Rim11. Subsequently, Ume6-Ime1 coactivator complexes form and induce EMG transcription. Our results demonstrate how various signaling inputs (PKA/TORC1/Ime1) converge through Rim11 to regulate EMG expression and meiosis initiation. We posit that the signaling-regulatory network elucidated here generates robustness in cell-fate control.

二倍体芽殖酵母细胞的饥饿会触发细胞命运程序,最终导致减数分裂和孢子形成。早期减数分裂基因(EMG)的转录激活取决于主调节因子Ime1、其DNA结合伙伴Ume6和GSK-3β激酶Rim11。EMG 激活需要 Rim11 对 Ume6 进行磷酸化。我们在此报告了 Rim11 作为控制 Ume6 磷酸化和 EMG 转录的中心信号整合器的功能。在营养丰富的条件下,PKA 会抑制 Rim11 的水平,而 TORC1 则会将 Rim11 保留在细胞质中。抑制 PKA 和 TORC1 会诱导 Rim11 的表达和核定位。值得注意的是,核 Rim11 是 Rim11 依赖性 Ume6 磷酸化所必需的,但还不够。此外,Ime1 是一种锚蛋白,能使 Ume6 通过 Rim11 磷酸化。随后,Ume6-Ime1 辅激活剂复合物形成并诱导 EMG 转录。我们的研究结果表明了各种信号输入(PKA/TORC1/Ime1)是如何通过 Rim11 汇聚到一起,调节 EMG 的表达和减数分裂的启动。我们认为,本文所阐明的信号调控网络可在细胞命运控制中产生稳健性。
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引用次数: 0
Human mitochondrial carriers of the SLC25 family function as monomers exchanging substrates with a ping-pong kinetic mechanism. SLC25 家族的人类线粒体载体以单体形式发挥作用,通过乒乓动力学机制交换底物。
IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-08-01 Epub Date: 2024-06-27 DOI: 10.1038/s44318-024-00150-0
Camila Cimadamore-Werthein, Martin S King, Denis Lacabanne, Eva Pyrihová, Stephany Jaiquel Baron, Edmund Rs Kunji

Members of the SLC25 mitochondrial carrier family link cytosolic and mitochondrial metabolism and support cellular maintenance and growth by transporting compounds across the mitochondrial inner membrane. Their monomeric or dimeric state and kinetic mechanism have been a matter of long-standing debate. It is believed by some that they exist as homodimers and transport substrates with a sequential kinetic mechanism, forming a ternary complex where both exchanged substrates are bound simultaneously. Some studies, in contrast, have provided evidence indicating that the mitochondrial ADP/ATP carrier (SLC25A4) functions as a monomer, has a single substrate binding site, and operates with a ping-pong kinetic mechanism, whereby ADP is imported before ATP is exported. Here we reanalyze the oligomeric state and kinetic properties of the human mitochondrial citrate carrier (SLC25A1), dicarboxylate carrier (SLC25A10), oxoglutarate carrier (SLC25A11), and aspartate/glutamate carrier (SLC25A13), all previously reported to be dimers with a sequential kinetic mechanism. We demonstrate that they are monomers, except for dimeric SLC25A13, and operate with a ping-pong kinetic mechanism in which the substrate import and export steps occur consecutively. These observations are consistent with a common transport mechanism, based on a functional monomer, in which a single central substrate-binding site is alternately accessible.

SLC25 线粒体载体家族的成员将细胞代谢和线粒体代谢联系在一起,并通过跨线粒体内膜转运化合物来支持细胞的维持和生长。它们的单体或二聚体状态以及动力学机制一直是争论不休的问题。一些人认为,它们以同二聚体形式存在,以顺序动力学机制运输底物,形成一个三元复合物,其中同时结合两种交换底物。相反,一些研究提供的证据表明,线粒体 ADP/ATP 载体(SLC25A4)以单体形式存在,只有一个底物结合位点,并以乒乓动力学机制运行,即在输出 ATP 之前先输入 ADP。在这里,我们重新分析了人类线粒体柠檬酸盐载体(SLC25A1)、二羧酸盐载体(SLC25A10)、氧谷氨酸盐载体(SLC25A11)和天冬氨酸盐/谷氨酸盐载体(SLC25A13)的寡聚状态和动力学特性。我们证明,除了二聚体 SLC25A13 外,它们都是单体,并以乒乓动力学机制运行,其中底物的导入和导出步骤连续发生。这些观察结果与一种基于功能单体的常见转运机制相一致,在这种机制中,单个中心底物结合位点可交替访问。
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引用次数: 0
Optimized PAR-2 RING dimerization mediates cooperative and selective membrane binding for robust cell polarity. 优化的 PAR-2 RING 二聚化介导了合作性和选择性膜结合,从而实现稳健的细胞极性。
IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-08-01 Epub Date: 2024-06-21 DOI: 10.1038/s44318-024-00123-3
Tom Bland, Nisha Hirani, David C Briggs, Riccardo Rossetto, KangBo Ng, Ian A Taylor, Neil Q McDonald, David Zwicker, Nathan W Goehring

Cell polarity networks are defined by quantitative features of their constituent feedback circuits, which must be tuned to enable robust and stable polarization, while also ensuring that networks remain responsive to dynamically changing cellular states and/or spatial cues during development. Using the PAR polarity network as a model, we demonstrate that these features are enabled by the dimerization of the polarity protein PAR-2 via its N-terminal RING domain. Combining theory and experiment, we show that dimer affinity is optimized to achieve dynamic, selective, and cooperative binding of PAR-2 to the plasma membrane during polarization. Reducing dimerization compromises positive feedback and robustness of polarization. Conversely, enhanced dimerization renders the network less responsive due to kinetic trapping of PAR-2 on internal membranes and reduced sensitivity of PAR-2 to the anterior polarity kinase, aPKC/PKC-3. Thus, our data reveal a key role for a dynamically oligomeric RING domain in optimizing interaction affinities to support a robust and responsive cell polarity network, and highlight how optimization of oligomerization kinetics can serve as a strategy for dynamic and cooperative intracellular targeting.

细胞极性网络是由其组成的反馈回路的定量特征定义的,这些反馈回路必须经过调整才能实现稳健而稳定的极性化,同时还要确保网络在发育过程中对动态变化的细胞状态和/或空间线索保持响应。我们以 PAR 极性网络为模型,证明了极性蛋白 PAR-2 通过其 N 端 RING 结构域的二聚化可以实现这些特征。结合理论和实验,我们发现二聚体的亲和力得到了优化,从而在极化过程中实现了 PAR-2 与质膜的动态、选择性和合作性结合。减少二聚化会损害极化的正反馈和稳健性。相反,由于 PAR-2 在内部膜上的动力学捕获以及 PAR-2 对前极性激酶 aPKC/PKC-3 的敏感性降低,二聚化增强会降低网络的反应能力。因此,我们的数据揭示了动态寡聚的 RING 结构域在优化相互作用亲和力以支持稳健且反应灵敏的细胞极性网络中的关键作用,并强调了寡聚动力学的优化如何作为一种动态、合作的细胞内靶向策略。
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引用次数: 0
Antonello Covacci (1957-2023), a visionary and beautiful mind. 安东内罗-科瓦奇(Antonello Covacci,1957-2023 年),一个富有远见的美丽心灵。
IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-08-01 Epub Date: 2024-07-15 DOI: 10.1038/s44318-024-00167-5
Rino Rappuoli, Duccio Medini
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引用次数: 0
YTHDC1 m6A-dependent and m6A-independent functions converge to preserve the DNA damage response. YTHDC1 的 m6A 依赖性和 m6A 非依赖性功能共同维护 DNA 损伤反应。
IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-08-01 Epub Date: 2024-07-01 DOI: 10.1038/s44318-024-00153-x
Daniel Elvira-Blázquez, José Miguel Fernández-Justel, Aida Arcas, Luisa Statello, Enrique Goñi, Jovanna González, Benedetta Ricci, Sara Zaccara, Ivan Raimondi, Maite Huarte

Cells have evolved a robust and highly regulated DNA damage response to preserve their genomic integrity. Although increasing evidence highlights the relevance of RNA regulation, our understanding of its impact on a fully efficient DNA damage response remains limited. Here, through a targeted CRISPR-knockout screen, we identify RNA-binding proteins and modifiers that participate in the p53 response. Among the top hits, we find the m6A reader YTHDC1 as a master regulator of p53 expression. YTHDC1 binds to the transcription start sites of TP53 and other genes involved in the DNA damage response, promoting their transcriptional elongation. YTHDC1 deficiency also causes the retention of introns and therefore aberrant protein production of key DNA damage factors. While YTHDC1-mediated intron retention requires m6A, TP53 transcriptional pause-release is promoted by YTHDC1 independently of m6A. Depletion of YTHDC1 causes genomic instability and aberrant cancer cell proliferation mediated by genes regulated by YTHDC1. Our results uncover YTHDC1 as an orchestrator of the DNA damage response through distinct mechanisms of co-transcriptional mRNA regulation.

细胞进化出了一种强大而高度调控的 DNA 损伤反应,以保持其基因组的完整性。尽管越来越多的证据凸显了 RNA 调节的相关性,但我们对其对 DNA 损伤响应的影响的了解仍然有限。在这里,我们通过靶向 CRISPR 基因敲除筛选,确定了参与 p53 响应的 RNA 结合蛋白和修饰因子。在最热门的研究中,我们发现 m6A 阅读器 YTHDC1 是 p53 表达的主调控因子。YTHDC1 与 TP53 和其他参与 DNA 损伤反应的基因的转录起始位点结合,促进它们的转录伸长。YTHDC1 缺乏也会导致内含子的保留,从而导致关键 DNA 损伤因子的蛋白质生成异常。YTHDC1 介导的内含子保留需要 m6A,而 TP53 转录暂停释放则独立于 m6A 由 YTHDC1 促进。耗尽 YTHDC1 会导致基因组不稳定和由 YTHDC1 调控的基因介导的癌细胞异常增殖。我们的研究结果揭示了 YTHDC1 通过不同的共转录 mRNA 调控机制成为 DNA 损伤反应的协调者。
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引用次数: 0
Whole-body replacement of larval myofibers generates permanent adult myofibers in zebrafish. 斑马鱼幼体肌纤维的全身替代可产生永久性成体肌纤维。
IF 9.4 1区 生物学 Q1 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-08-01 Epub Date: 2024-06-05 DOI: 10.1038/s44318-024-00136-y
Uday Kumar, Chun-Yi Fang, Hsiao-Yuh Roan, Shao-Chun Hsu, Chung-Han Wang, Chen-Hui Chen

Drastic increases in myofiber number and size are essential to support vertebrate post-embryonic growth. However, the collective cellular behaviors that enable these increases have remained elusive. Here, we created the palmuscle myofiber tagging and tracking system for in toto monitoring of the growth and fates of ~5000 fast myofibers in developing zebrafish larvae. Through live tracking of individual myofibers within the same individuals over extended periods, we found that many larval myofibers readily dissolved during development, enabling the on-site addition of new and more myofibers. Remarkably, whole-body surveillance of multicolor-barcoded myofibers further unveiled a gradual yet extensive elimination of larval myofiber populations, resulting in near-total replacement by late juvenile stages. The subsequently emerging adult myofibers are not only long-lasting, but also morphologically and functionally distinct from the larval populations. Furthermore, we determined that the elimination-replacement process is dependent on and driven by the autophagy pathway. Altogether, we propose that the whole-body replacement of larval myofibers is an inherent yet previously unnoticed process driving organismic muscle growth during vertebrate post-embryonic development.

肌纤维数量和大小的急剧增加对支持脊椎动物胚胎后生长至关重要。然而,实现这些增长的细胞集体行为仍然难以捉摸。在这里,我们创建了掌肌肌纤维标记和跟踪系统,用于全面监测发育中斑马鱼幼体中约 5000 条快速肌纤维的生长和命运。通过对同一个体内的单个肌纤维进行长时间的实时跟踪,我们发现许多幼体肌纤维在发育过程中很容易溶解,从而可以现场添加新的和更多的肌纤维。值得注意的是,对全身多色条纹肌纤维的监测进一步揭示了幼体肌纤维群逐渐而广泛地消亡的过程,结果是幼体后期几乎完全被替代。随后出现的成体肌纤维不仅持续时间长,而且在形态和功能上都与幼体不同。此外,我们还确定,消除-替代过程依赖于自噬途径,并由自噬途径驱动。总之,我们提出,幼体肌纤维的全身替换是脊椎动物胚后发育过程中驱动有机体肌肉生长的一个固有但以前未被注意的过程。
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引用次数: 0
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