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Heterogeneity of late endosome/lysosomes shown by multiplexed DNA-PAINT imaging. 多重 DNA-PAINT 成像显示晚期内膜体/溶酶体的异质性。
IF 7.4 1区 生物学 Q1 CELL BIOLOGY Pub Date : 2025-01-06 Epub Date: 2024-11-01 DOI: 10.1083/jcb.202403116
Charles Bond, Siewert Hugelier, Jiazheng Xing, Elena M Sorokina, Melike Lakadamyali

Late endosomes/lysosomes (LELs) are crucial for numerous physiological processes and their dysfunction is linked to many diseases. Proteomic analyses have identified hundreds of LEL proteins; however, whether these proteins are uniformly present on each LEL, or if there are cell-type-dependent LEL subpopulations with unique protein compositions is unclear. We employed quantitative, multiplexed DNA-PAINT super-resolution imaging to examine the distribution of seven key LEL proteins (LAMP1, LAMP2, CD63, Cathepsin D, TMEM192, NPC1, and LAMTOR4). While LAMP1, LAMP2, and Cathepsin D were abundant across LELs, marking a common population, most analyzed proteins were associated with specific LEL subpopulations. Our multiplexed imaging approach identified up to eight different LEL subpopulations based on their unique membrane protein composition. Additionally, our analysis of the spatial relationships between these subpopulations and mitochondria revealed a cell-type-specific tendency for NPC1-positive LELs to be closely positioned to mitochondria. Our approach will be broadly applicable to determining organelle heterogeneity with single organelle resolution in many biological contexts.

晚期内体/溶酶体(LELs)对许多生理过程至关重要,其功能障碍与许多疾病有关。蛋白质组学分析已鉴定出数百种 LEL 蛋白;然而,这些蛋白质是否均匀地存在于每个 LEL 上,或者是否存在具有独特蛋白质组成的细胞类型依赖性 LEL 亚群,目前还不清楚。我们采用定量、多重 DNA-PAINT 超分辨率成像技术研究了七种关键 LEL 蛋白(LAMP1、LAMP2、CD63、Cathepsin D、TMEM192、NPC1 和 LAMTOR4)的分布情况。虽然 LAMP1、LAMP2 和 Cathepsin D 在 LEL 中含量丰富,标志着一个共同的群体,但大多数分析的蛋白质都与特定的 LEL 亚群相关。我们的多重成像方法根据 LEL 独特的膜蛋白组成确定了多达八个不同的 LEL 亚群。此外,我们对这些亚群与线粒体之间空间关系的分析表明,NPC1 阳性的 LEL 与线粒体的位置密切相关,这是一种细胞类型特异性趋势。我们的方法将广泛适用于在许多生物环境中以单个细胞器分辨率确定细胞器的异质性。
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引用次数: 0
Postsynaptic BMP signaling regulates myonuclear properties in Drosophila larval muscles. 突触后BMP信号调节果蝇幼虫肌肉的肌核特性。
IF 7.4 1区 生物学 Q1 CELL BIOLOGY Pub Date : 2025-01-06 Epub Date: 2024-10-30 DOI: 10.1083/jcb.202404052
Victoria E von Saucken, Stefanie E Windner, Giovanna Armetta, Mary K Baylies

The syncytial mammalian muscle fiber contains a heterogeneous population of (myo)nuclei. At the neuromuscular junction (NMJ), myonuclei have specialized positioning and gene expression. However, it remains unclear how myonuclei are recruited and what regulates myonuclear output at the NMJ. Here, we identify specific properties of myonuclei located near the Drosophila larval NMJ. These synaptic myonuclei have increased size in relation to their surrounding cytoplasmic domain (size scaling), increased DNA content (ploidy), and increased levels of transcription factor pMad, a readout for BMP signaling activity. Our genetic manipulations show that local BMP signaling affects muscle size, nuclear size, ploidy, and NMJ size and function. In support, RNA sequencing analysis reveals that pMad regulates genes involved in muscle growth, ploidy (i.e., E2f1), and neurotransmission. Our data suggest that muscle BMP signaling instructs synaptic myonuclear output that positively shapes the NMJ synapse. This study deepens our understanding of how myonuclear heterogeneity supports local signaling demands to fine tune cellular function and NMJ activity.

哺乳动物的肌纤维中含有大量不同的(肌)核。在神经肌肉接头(NMJ)处,肌核有专门的定位和基因表达。然而,目前仍不清楚肌核是如何被招募的,以及是什么调控肌核在 NMJ 的输出。在这里,我们确定了位于果蝇幼虫 NMJ 附近的肌核的特殊属性。这些突触肌核的大小相对于其周围的细胞质域增大(大小缩放),DNA含量增加(倍性),转录因子pMad的水平增加,而pMad是BMP信号活动的读数。我们的遗传操作表明,局部 BMP 信号影响肌肉大小、核大小、倍性以及 NMJ 大小和功能。作为佐证,RNA 测序分析显示 pMad 可调控涉及肌肉生长、倍性(即 E2f1)和神经传递的基因。我们的数据表明,肌肉 BMP 信号指示突触肌核输出,从而积极塑造 NMJ 突触。这项研究加深了我们对肌核异质性如何支持局部信号需求以微调细胞功能和 NMJ 活动的理解。
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引用次数: 0
Definition of phosphatidylinositol 4,5-bisphosphate distribution by freeze-fracture replica labeling. 通过冷冻断裂复制标记确定磷脂酰肌醇 4,5-二磷酸的分布。
IF 7.4 1区 生物学 Q1 CELL BIOLOGY Pub Date : 2025-01-06 Epub Date: 2024-11-04 DOI: 10.1083/jcb.202311067
Takuma Tsuji, Junya Hasegawa, Takehiko Sasaki, Toyoshi Fujimoto

Phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] is a phospholipid essential for plasma membrane functions, but its two-dimensional distribution is not clear. Here, we compared the result of sodium dodecyl sulfate-treated freeze-fracture replica labeling (SDS-FRL) of quick-frozen cells with the actual PtdIns(4,5)P2 content and the results obtained by fluorescence biosensor and by labeling of chemically-fixed membranes. In yeast, enrichment of PtdIns(4,5)P2 in the membrane compartment of Can1 (MCC)/eisosome, especially in the curved MCC/eisosome, was evident by SDS-FRL, but not by fluorescence biosensor, GFP-PLC1δ-PH. PtdIns(4,5)P2 remaining after acute ATP depletion and in the stationary phase, 30.0% and 56.6% of the control level, respectively, was not detectable by fluorescence biosensor, whereas the label intensity by SDS-FRL reflected the PtdIns(4,5)P2 amount. In PC12 cells, PtdIns(4,5)P2 was observed in a punctate pattern in the formaldehyde-fixed plasma membrane, whereas it was distributed randomly by SDS-FRL and showed clustering after formaldehyde fixation. The results indicate that the distribution of PtdIns(4,5)P2 can be defined most reliably by SDS-FRL of quick-frozen cells.

磷脂酰肌醇4,5-二磷酸[PtdIns(4,5)P2]是一种对质膜功能至关重要的磷脂,但其二维分布并不清楚。在这里,我们比较了十二烷基硫酸钠处理的速冻细胞冻裂复制标记(SDS-FRL)结果与 PtdIns(4,5)P2 的实际含量,以及荧光生物传感器和化学固定膜标记的结果。在酵母菌中,SDS-FRL检测到PtdIns(4,5)P2在Can1 (MCC)/eisosome的膜区,尤其是在弯曲的MCC/eisosome中富集,但荧光生物传感器GFP-PLC1δ-PH的检测结果并不明显。荧光生物传感器检测不到ATP急性耗竭后和静止期剩余的PtdIns(4,5)P2,分别为对照水平的30.0%和56.6%,而SDS-FRL的标记强度反映了PtdIns(4,5)P2的量。在 PC12 细胞中,PtdIns(4,5)P2 在甲醛固定的质膜上呈点状分布,而在 SDS-FRL 中则呈随机分布,甲醛固定后则呈聚集状。结果表明,速冻细胞的 SDS-FRL 能够最可靠地确定 PtdIns(4,5)P2 的分布。
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引用次数: 0
PINK1 controls RTN3L-mediated ER autophagy by regulating peripheral tubule junctions. PINK1 通过调节外周小管连接控制 RTN3L 介导的 ER 自噬。
IF 7.4 1区 生物学 Q1 CELL BIOLOGY Pub Date : 2024-12-02 Epub Date: 2024-11-18 DOI: 10.1083/jcb.202407193
Ravi Chidambaram, Kamal Kumar, Smriti Parashar, Gowsalya Ramachandran, Shuliang Chen, Susan Ferro-Novick

Here, we report that the RTN3L-SEC24C endoplasmic reticulum autophagy (ER-phagy) receptor complex, the CUL3KLHL12 E3 ligase that ubiquitinates RTN3L, and the FIP200 autophagy initiating protein, target mutant proinsulin (Akita) condensates for lysosomal delivery at ER tubule junctions. When delivery was blocked, Akita condensates accumulated in the ER. In exploring the role of tubulation in these events, we unexpectedly found that loss of the Parkinson's disease protein, PINK1, reduced peripheral tubule junctions and blocked ER-phagy. Overexpression of the PINK1 kinase substrate, DRP1, increased junctions, reduced Akita condensate accumulation, and restored lysosomal delivery in PINK1-depleted cells. DRP1 is a dual-functioning protein that promotes ER tubulation and severs mitochondria at ER-mitochondria contact sites. DRP1-dependent ER tubulating activity was sufficient for suppression. Supporting these findings, we observed PINK1 associating with ER tubules. Our findings show that PINK1 shapes the ER to target misfolded proinsulin for RTN3L-SEC24C-mediated macro-ER-phagy at defined ER sites called peripheral junctions. These observations may have important implications for understanding Parkinson's disease.

在这里,我们报告了RTN3L-SEC24C内质网自噬(ER-phagy)受体复合物、泛素化RTN3L的CUL3KLHL12 E3连接酶以及FIP200自噬启动蛋白在ER小管连接处靶向突变型胰蛋白酶(秋田)凝集物进行溶酶体递送。当输送受阻时,秋田凝集物就会在 ER 中积累。在探索小管在这些事件中的作用时,我们意外地发现,帕金森病蛋白 PINK1 的缺失会减少外周小管连接并阻止 ER 吞噬。在缺失 PINK1 的细胞中,过量表达 PINK1 激酶底物 DRP1 增加了连接,减少了秋田凝集物的积累,并恢复了溶酶体输送。DRP1 是一种具有双重功能的蛋白质,它能促进ER管化,并在ER-线粒体接触点处切断线粒体。依赖 DRP1 的 ER 管化活性足以抑制线粒体。为支持这些发现,我们观察到 PINK1 与 ER 小管有关联。我们的研究结果表明,PINK1能在被称为外周连接点的确定的ER位点上塑造ER,将错误折叠的前胰岛素作为RTN3L-SEC24C介导的大ER吞噬的目标。这些观察结果可能对理解帕金森病具有重要意义。
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引用次数: 0
Stairway to the Golgi: Two paths VPS13B can go by. 通往高尔基体的阶梯VPS13B 有两条路可走
IF 7.4 1区 生物学 Q1 CELL BIOLOGY Pub Date : 2024-12-02 Epub Date: 2024-11-12 DOI: 10.1083/jcb.202411005
Roger Pons Lanau, Felix Campelo

VPS13 proteins mediate lipid transfer across membranes. Among them, VPS13B is associated with Golgi membranes, and VPS13B mutations cause Cohen syndrome. In this issue, Ugur et al. (https://doi.org/10.1083/jcb.202311189) and Du et al. (https://doi.org/10.1083/jcb.202402083) reveal new VPS13B interactors and their functions in Golgi organization and trafficking.

VPS13 蛋白介导跨膜脂质转移。其中,VPS13B 与高尔基体膜有关,VPS13B 突变会导致科恩综合征。在本期中,Ugur 等人 (https://doi.org/10.1083/jcb.202311189) 和 Du 等人 (https://doi.org/10.1083/jcb.202402083) 揭示了新的 VPS13B 相互作用因子及其在高尔基体组织和贩运中的功能。
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引用次数: 0
Visualizing ER-phagy and ER architecture in vivo. 体内ER吞噬和ER结构的可视化。
IF 7.4 1区 生物学 Q1 CELL BIOLOGY Pub Date : 2024-12-02 Epub Date: 2024-11-18 DOI: 10.1083/jcb.202408061
Yongjuan Sang, Boran Li, Tinglin Su, Hanyu Zhan, Yue Xiong, Zhiming Huang, Changjing Wang, Xiaoxia Cong, Mengjie Du, Yang Wu, Hang Yu, Xi Yang, Kezhi Ding, Xuhua Wang, Xiaolong Miao, Weihua Gong, Liang Wang, Jingwei Zhao, Yiting Zhou, Wei Liu, Xinyang Hu, Qiming Sun

ER-phagy is an evolutionarily conserved mechanism crucial for maintaining cellular homeostasis. However, significant gaps persist in our understanding of how ER-phagy and the ER network vary across cell subtypes, tissues, and organs. Furthermore, the pathophysiological relevance of ER-phagy remains poorly elucidated. Addressing these questions requires developing quantifiable methods to visualize ER-phagy and ER architecture in vivo. We generated two transgenic mouse lines expressing an ER lumen-targeting tandem RFP-GFP (ER-TRG) tag, either constitutively or conditionally. This approach enables precise spatiotemporal measurements of ER-phagy and ER structure at single-cell resolution in vivo. Systemic analysis across diverse organs, tissues, and primary cultures derived from these ER-phagy reporter mice unveiled significant variations in basal ER-phagy, both in vivo and ex vivo. Furthermore, our investigation uncovered substantial remodeling of ER-phagy and the ER network in different tissues under stressed conditions such as starvation, oncogenic transformation, and tissue injury. In summary, both reporter models represent valuable resources with broad applications in fundamental research and translational studies.

ER吞噬是一种进化保守的机制,对维持细胞稳态至关重要。然而,我们对ER噬菌过程和ER网络如何在不同细胞亚型、组织和器官之间发生变化的认识仍然存在很大差距。此外,ER-噬的病理生理相关性仍未得到充分阐明。要解决这些问题,需要开发可量化的方法来观察体内的ER噬和ER结构。我们产生了两种转基因小鼠品系,它们组成型或条件型表达ER腔靶向串联RFP-GFP(ER-TRG)标签。这种方法可在体内以单细胞分辨率对ER吞噬和ER结构进行精确的时空测量。对来自这些ER噬菌报告小鼠的不同器官、组织和原代培养物的系统分析揭示了体内和体外基础ER噬菌的显著变化。此外,我们的研究还发现,在饥饿、致癌转化和组织损伤等应激条件下,不同组织中的ER噬功能和ER网络会发生重大重塑。总之,这两种报告模型都是宝贵的资源,可广泛应用于基础研究和转化研究。
{"title":"Visualizing ER-phagy and ER architecture in vivo.","authors":"Yongjuan Sang, Boran Li, Tinglin Su, Hanyu Zhan, Yue Xiong, Zhiming Huang, Changjing Wang, Xiaoxia Cong, Mengjie Du, Yang Wu, Hang Yu, Xi Yang, Kezhi Ding, Xuhua Wang, Xiaolong Miao, Weihua Gong, Liang Wang, Jingwei Zhao, Yiting Zhou, Wei Liu, Xinyang Hu, Qiming Sun","doi":"10.1083/jcb.202408061","DOIUrl":"https://doi.org/10.1083/jcb.202408061","url":null,"abstract":"<p><p>ER-phagy is an evolutionarily conserved mechanism crucial for maintaining cellular homeostasis. However, significant gaps persist in our understanding of how ER-phagy and the ER network vary across cell subtypes, tissues, and organs. Furthermore, the pathophysiological relevance of ER-phagy remains poorly elucidated. Addressing these questions requires developing quantifiable methods to visualize ER-phagy and ER architecture in vivo. We generated two transgenic mouse lines expressing an ER lumen-targeting tandem RFP-GFP (ER-TRG) tag, either constitutively or conditionally. This approach enables precise spatiotemporal measurements of ER-phagy and ER structure at single-cell resolution in vivo. Systemic analysis across diverse organs, tissues, and primary cultures derived from these ER-phagy reporter mice unveiled significant variations in basal ER-phagy, both in vivo and ex vivo. Furthermore, our investigation uncovered substantial remodeling of ER-phagy and the ER network in different tissues under stressed conditions such as starvation, oncogenic transformation, and tissue injury. In summary, both reporter models represent valuable resources with broad applications in fundamental research and translational studies.</p>","PeriodicalId":15211,"journal":{"name":"Journal of Cell Biology","volume":"223 12","pages":""},"PeriodicalIF":7.4,"publicationDate":"2024-12-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142648286","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Non-cell autonomous regulation of cell-cell signaling and differentiation by mitochondrial ROS. 线粒体 ROS 对细胞-细胞信号传递和分化的非细胞自主调控。
IF 7.4 1区 生物学 Q1 CELL BIOLOGY Pub Date : 2024-12-02 Epub Date: 2024-11-13 DOI: 10.1083/jcb.202401084
Yipeng Du, Lei Wang, Lizbeth Perez-Castro, Maralice Conacci-Sorrell, Matthew Sieber

Mitochondrial reactive oxygen species (ROS) function intrinsically within cells to induce cell damage, regulate transcription, and cause genome instability. However, we know little about how mitochondrial ROS production non-cell autonomously impacts cell-cell signaling. Here, we show that mitochondrial dysfunction inhibits the plasma membrane localization of cell surface receptors that drive cell-cell communication during oogenesis. Within minutes, we found that mitochondrial ROS impairs exocyst membrane binding and leads to defective endosomal recycling. This endosomal defect impairs the trafficking of receptors, such as the Notch ligand Delta, during oogenesis. Remarkably, we found that overexpressing RAB11 restores ligand trafficking and rescues the developmental defects caused by ROS production. ROS production from adjacent cells acutely initiates a transcriptional response associated with growth and migration by suppressing Notch signaling and inducing extra cellualr matrix (ECM) remodeling. Our work reveals a conserved rapid response to ROS production that links mitochondrial dysfunction to the non-cell autonomous regulation of cell-cell signaling.

线粒体活性氧(ROS)在细胞内具有内在功能,可诱导细胞损伤、调节转录并导致基因组不稳定。然而,我们对线粒体 ROS 的产生如何非细胞自主地影响细胞-细胞信号传导知之甚少。在这里,我们发现线粒体功能障碍会抑制细胞表面受体的质膜定位,而细胞表面受体会在卵子发生过程中驱动细胞间通信。在几分钟内,我们发现线粒体 ROS 会损害外囊膜结合,并导致内体循环缺陷。这种内体缺陷会影响受体(如 Notch 配体 Delta)在卵子发生过程中的运输。值得注意的是,我们发现过量表达 RAB11 能恢复配体的贩运,并挽救因 ROS 产生而导致的发育缺陷。邻近细胞产生的 ROS 通过抑制 Notch 信号传导和诱导细胞外基质(ECM)重塑,迅速启动了与生长和迁移相关的转录反应。我们的研究揭示了对 ROS 产生的一种保守的快速反应,它将线粒体功能障碍与细胞-细胞信号传导的非细胞自主调节联系起来。
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引用次数: 0
Local glycolysis supports injury-induced axonal regeneration. 局部糖酵解支持损伤诱导的轴突再生。
IF 7.4 1区 生物学 Q1 CELL BIOLOGY Pub Date : 2024-12-02 Epub Date: 2024-10-01 DOI: 10.1083/jcb.202402133
Luca Masin, Steven Bergmans, Annelies Van Dyck, Karl Farrow, Lies De Groef, Lieve Moons

Successful axonal regeneration following injury requires the effective allocation of energy. How axons withstand the initial disruption in mitochondrial energy production caused by the injury and subsequently initiate regrowth is poorly understood. Transcriptomic data showed increased expression of glycolytic genes after optic nerve crush in retinal ganglion cells with the co-deletion of Pten and Socs3. Using retinal cultures in a multicompartment microfluidic device, we observed increased regrowth and enhanced mitochondrial trafficking in the axons of Pten and Socs3 co-deleted neurons. While wild-type axons relied on mitochondrial metabolism, after injury, in the absence of Pten and Socs3, energy production was supported by local glycolysis. Specific inhibition of lactate production hindered injury survival and the initiation of regrowth while slowing down glycolysis upstream impaired regrowth initiation, axonal elongation, and energy production. Together, these observations reveal that glycolytic ATP, combined with sustained mitochondrial transport, is essential for injury-induced axonal regrowth, providing new insights into the metabolic underpinnings of axonal regeneration.

损伤后轴突的成功再生需要有效的能量分配。轴突如何经受住损伤引起的线粒体能量产生的最初中断并随后开始再生,目前尚不清楚。转录组数据显示,视网膜神经节细胞在视神经挤压后,Pten 和 Socs3 基因共同缺失,糖酵解基因的表达增加。利用多室微流体装置中的视网膜培养物,我们观察到 Pten 和 Socs3 共缺失神经元的轴突再生能力增强,线粒体贩运能力增强。野生型轴突依赖线粒体代谢,而损伤后,在缺乏 Pten 和 Socs3 的情况下,能量生产则由局部糖酵解支持。特异性抑制乳酸盐的产生会阻碍损伤后的存活和再生的启动,而减缓上游糖酵解则会损害再生的启动、轴突的伸长和能量的产生。这些观察结果共同揭示了糖酵解 ATP 与线粒体持续转运相结合对损伤诱导的轴突再生至关重要,为轴突再生的代谢基础提供了新的见解。
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引用次数: 0
Dachsous and Fat coordinately repress the Dachs-Dlish-Approximated complex to control growth. Dachsous和Fat协同抑制Dachs-Dlish-Approximated复合体,从而控制生长。
IF 7.4 1区 生物学 Q1 CELL BIOLOGY Pub Date : 2024-12-02 Epub Date: 2024-10-07 DOI: 10.1083/jcb.202406119
Hitoshi Matakatsu, Richard G Fehon

Two protocadherins, Dachsous and Fat, regulate organ growth in Drosophila via the Hippo pathway. Dachsous and Fat bind heterotypically to regulate the abundance and subcellular localization of a "core complex" consisting of Dachs, Dlish, and Approximated. This complex localizes to the junctional cortex where it represses Warts. Dachsous is believed to promote growth by recruiting and stabilizing this complex, while Fat represses growth by promoting its degradation. Here, we examine the functional relationships between the intracellular domains of Dachsous and Fat and the core complex. While Dachsous promotes the accumulation of core complex proteins in puncta, it is not required for their assembly. Indeed, the core complex accumulates maximally in the absence of both Dachsous and Fat. Furthermore, Dachsous represses growth in the absence of Fat by removing the core complex from the junctional cortex. Fat similarly recruits core complex components but promotes their degradation. Our findings reveal that Dachsous and Fat coordinately constrain tissue growth by repressing the core complex.

两种原粘连蛋白 Dachsous 和 Fat 通过 Hippo 途径调节果蝇器官的生长。Dachsous 和 Fat 通过异型结合来调节由 Dachs、Dlish 和 Approximated 组成的 "核心复合体 "的丰度和亚细胞定位。该复合体定位于交界皮层,在那里抑制疣。据信,Dachsous 通过招募和稳定这一复合体来促进生长,而 Fat 则通过促进其降解来抑制生长。在这里,我们研究了 Dachsous 和 Fat 的胞内结构域与核心复合体之间的功能关系。虽然Dachsous能促进核心复合体蛋白在点状结构中的积累,但它们的组装并不需要它。事实上,在没有 Dachsous 和 Fat 的情况下,核心复合体会最大程度地聚集。此外,在没有脂肪的情况下,Dachsous 通过从连接皮层移除核心复合体来抑制生长。脂肪同样也会招募核心复合体成分,但会促进它们的降解。我们的研究结果表明,Dachsous和Fat通过抑制核心复合体协调地限制了组织的生长。
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引用次数: 0
eIF5A controls mitoprotein import by relieving ribosome stalling at TIM50 translocase mRNA. eIF5A 通过缓解 TIM50 易位酶 mRNA 的核糖体停滞来控制有丝分裂蛋白的导入。
IF 7.4 1区 生物学 Q1 CELL BIOLOGY Pub Date : 2024-12-02 Epub Date: 2024-11-07 DOI: 10.1083/jcb.202404094
Marina Barba-Aliaga, Vanessa Bernal, Cynthia Rong, Madeleine E Volfbeyn, Keguang Zhang, Brian M Zid, Paula Alepuz

Efficient import of nuclear-encoded proteins into mitochondria is crucial for proper mitochondrial function. The conserved translation factor eIF5A binds ribosomes, alleviating stalling at polyproline-encoding sequences. eIF5A impacts mitochondrial function across species, though the precise molecular mechanism is unclear. We found that eIF5A depletion in yeast reduces the translation and levels of the TCA cycle and oxidative phosphorylation proteins. Loss of eIF5A causes mitoprotein precursors to accumulate in the cytosol and triggers a mitochondrial import stress response. We identify an essential polyproline protein as a direct target of eIF5A: the mitochondrial inner membrane protein and translocase component Tim50. Thus, eIF5A controls mitochondrial protein import by alleviating ribosome stalling along Tim50 mRNA at the mitochondrial surface. Removal of polyprolines from Tim50 partially rescues the mitochondrial import stress response and translation of oxidative phosphorylation genes. Overall, our findings elucidate how eIF5A impacts the mitochondrial function by promoting efficient translation and reducing ribosome stalling of co-translationally imported proteins, thereby positively impacting the mitochondrial import process.

将核编码蛋白质高效导入线粒体对线粒体功能的正常发挥至关重要。保守的翻译因子 eIF5A 与核糖体结合,缓解了多肽编码序列的停滞。eIF5A 对不同物种的线粒体功能都有影响,但确切的分子机制尚不清楚。我们发现,酵母中的 eIF5A 缺失会降低 TCA 循环和氧化磷酸化蛋白的翻译和水平。eIF5A 的缺失会导致有丝分裂蛋白前体在细胞质中积累,并引发线粒体输入应激反应。我们发现一种重要的多脯氨酸蛋白是 eIF5A 的直接靶标:线粒体内膜蛋白和转运酶成分 Tim50。因此,eIF5A 通过缓解线粒体表面 Tim50 mRNA 上的核糖体停滞来控制线粒体蛋白质的导入。去除 Tim50 中的多脯氨酸可部分缓解线粒体导入应激反应和氧化磷酸化基因的翻译。总之,我们的研究结果阐明了 eIF5A 如何通过促进高效翻译和减少共翻译导入蛋白的核糖体停滞来影响线粒体功能,从而对线粒体导入过程产生积极影响。
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引用次数: 0
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