
섬모 뿌리는 세포 내부 구조물에 섬모를 연결하는 중요한 역할을 하지만, 그 구조와 기능에 대해서는 잘 알려져 있지 않다. 이 연구에서는 cryo-ET를 이용하여 섬모 뿌리의 3차원 구조를 분석하고, 그 구조적 특징과 기능적 의미를 제시한다.


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Ciliary rootlets are striated bundles of filaments that connect the base of cilia to internal cellular structures. Rootlets are critical for the sensory and motile functions of cilia. However, the mechanisms underlying these functions remain unknown, in part due to a lack of structural information of rootlet organization. In this study, we obtain 3D reconstructions of membrane-associated and purified rootlets using cryo-electron tomography (cryo-ET). We show that flexible protrusions on the rootlet surface, which emanate from the cross-striations, connect to intracellular membranes. In purified rootlets, the striations were classified into amorphous (A)-bands, associated with accumulations on the rootlet surface, and discrete (D)-bands corresponding to punctate lines of density that run through the rootlet. These striations connect a flexible network of longitudinal filaments. Subtomogram averaging suggests the filaments consist of two intertwined coiled coils. The rootlet’s filamentous architecture, with frequent membrane-connecting cross-striations, lends itself well for anchoring large membranes in the cell.

### Competing Interest Statement

The authors have declared no competing interest.

A cryo-ET study of ciliary rootlet organization

마우스-광수용체-세포의-섬모-뿌리-조직에-대한-cryo-et-연구


마우스 광수용체 세포의 섬모 뿌리 조직에 대한 cryo-ET 연구


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수정 과정에서 편모의 구조적 변화가 일어나며, 이는 편모의 운동 정지에 필수적이다.


Mammalian sperm delve into the female reproductive tract to fertilize the female gamete. The available information about how sperm regulate their motility during the final journey to the fertilization site is extremely limited. In this work, we investigated the structural and functional changes in the sperm flagellum after AE and during the interaction with the eggs. The evidence demonstrates that the double helix actin network surrounding the mitochondrial sheath of the midpiece undergoes structural changes prior to the motility cessation. This structural modification is accompanied by a decrease in diameter of the midpiece and is driven by intracellular calcium changes that occur concomitant with a reorganization of the actin helicoidal cortex. Midpiece contraction occurs in a subset of cells that undergo AE, live-cell imaging during in vitro fertilization showed that the midpiece contraction is required for motility cessation after fusion is initiated. These findings provide the first evidence of the F-actin network’s role in regulating sperm motility, adapting its function to meet specific cellular requirements during fertilization, and highlighting the broader significance of understanding sperm motility.

Significant statement In this work, we demonstrate that the helical structure of polymerized actin in the flagellum undergoes a rearrangement at the time of sperm-egg fusion. This process is driven by intracellular calcium and promotes a decrease in the sperm midpiece diameter as well as the arrest in motility, which is observed after the fusion process is initiated.

### Competing Interest Statement

The authors have declared no competing interest.

Reorganization of the Flagellum Scaffolding Induces a Sperm Standstill During Fertilization

난자-수정을-위해-필요한-편모-지지체의-재구성


난자 수정을 위해 필요한 편모 지지체의 재구성



리소좀은 미토콘드리아 내막의 손상된 부분을 선별적으로 제거하는 새로운 경로를 제공한다.


We show that cytosolic inner mitochondrial membrane vesicles, devoid of outer mitochondrial membrane or mitochondrial matrix, are formed during resting state and directly herniate into lysosomes through pores formed by voltage-dependent anion channel 1 in the outer mitochondrial membrane, thereby allowing their selective removal.

Lysosomes drive the piecemeal removal of mitochondrial inner membrane - Nature

미토콘드리아-내막의-점진적-제거를-주도하는-리소좀


미토콘드리아 내막의 점진적 제거를 주도하는 리소좀



다중섬모세포에서 섬모 재생은 전이대 없이 시작되며, 전이대 단백질은 새로 합성되어야 한다. 또한 세포는 제한된 단백질 공급 상황에서 더 긴 섬모를 가진 적은 수의 섬모를 선택적으로 재생한다.


Cilia regeneration is a physiological event, and while studied extensively in unicellular organisms, it remains poorly understood in vertebrates. In this study, using Xenopus multiciliated cells (MCCs) as a model, we demonstrate that, unlike unicellular organisms, deciliation removes the transition zone (TZ) and the ciliary axoneme. While MCCs immediately begin the regeneration of the ciliary axoneme, surprisingly, the assembly of TZ is delayed. However, ciliary tip proteins, Sentan and Clamp, localize to regenerating cilia without delay. Using cycloheximide (CHX) to block new protein synthesis, we show that the TZ protein B9d1 is not a component of the cilia precursor pool and requires new transcription/translation, providing insights into the delayed repair of TZ. Moreover, MCCs in CHX treatment assemble fewer (∼ 10 vs. ∼150 in controls) but near wild-type length (ranging between 60 to 90%) cilia by gradually concentrating ciliogenesis proteins like IFTs at a select few basal bodies. Using mathematical modeling, we show that cilia length compared to cilia number influences the force generated by MCCs more. In summary, our results question the requirement of TZ in motile cilia assembly and provide insights into how cells determine organelle size and number.

### Competing Interest Statement

The authors have declared no competing interest.

Mechanisms of cilia regeneration in Xenopus multiciliated epithelium in vivo

다양한-척추동물에서-다중섬모세포의-섬모-재생-메커니즘


다양한 척추동물에서 다중섬모세포의 섬모 재생 메커니즘



CD9과 CD81 테트라스파닌은 터널링 나노튜브 형성과 기능에 있어 서로 다른 비중복적인 역할을 한다. CD9은 나노튜브의 안정화에 기여하고, CD81은 나노튜브를 통한 소포 전달에 필요하다.


Tunneling nanotubes (TNTs) are open actin- and membrane-based channels, connecting remote cells and allowing direct transfer of cellular material (e.g. vesicles, mRNAs, protein aggregates) from cytoplasm to cytoplasm. Although they are important especially in pathological conditions (e.g., cancers, neurodegenerative diseases), their precise composition and their regulation were still poorly described. Here, using a biochemical approach allowing to separate TNTs from cell bodies and from extracellular vesicles and particles (EVPs), we obtained the full composition of TNTs compared to EVPs. We then focused to two major components of our proteomic data, the CD9 and CD81 tetraspanins, and further investigated their specific roles in TNT formation and function. We show that these two tetraspanins have distinct non-redundant functions: CD9 participates in stabilizing TNTs, whereas CD81 expression is required to allow the functional transfer of vesicle in the newly formed TNTs, possibly by regulating docking to or fusion with the opposing cell.

### Competing Interest Statement

The authors have declared no competing interest.

Proteomic landscape of tunneling nanotubes reveals CD9 and CD81 tetraspanins as key regulators

터널링-나노튜브의-프로테오믹-분석을-통해-cd9과-cd81-테트라스파닌이-핵심-조절자임이-밝혀짐


터널링 나노튜브의 프로테오믹 분석을 통해 CD9과 CD81 테트라스파닌이 핵심 조절자임이 밝혀짐



단백질 응집체 성숙화 과정에서 스핑고지질 대사 경로가 핵심적인 역할을 하며, 이를 통해 세포는 병원성 단백질 응집체의 독성을 해독할 수 있다.


Due to proteostasis stress induced by aging or disease, misfolded proteins can form toxic intermediate species of aggregates and eventually mature into less toxic inclusion bodies (IBs). Here, using a yeast imaging-based screen, we identified 84 potential synphilin-1 (SY1) IB regulators and isolated the conserved sphingolipid metabolic components in the most enriched groups. Furthermore, we show that, in both yeast cells and mammalian cells, SY1 IBs are associated with mitochondria. Pharmacological inhibition of the sphingolipid metabolism pathway or knockout of its key genes results in a delayed IB maturation and increased SY1 cytotoxicity. We postulate that SY1 IB matures by association with the mitochondrion membrane, and that sphingolipids stimulate the maturation via their membrane-modulating function and thereby protecting cells from SY1 cytotoxicity. Our findings identify a conserved cellular component essential for IB maturation and suggest a mechanism by which cells may detoxify the pathogenic protein aggregates through forming mitochondrion-associated IBs.

### Competing Interest Statement

The authors have declared no competing interest.

Maturation and detoxification of synphilin-1 inclusion bodies regulated by sphingolipids

단백질-응집체-성숙화와-해독화에-있어-스핑고지질의-역할


단백질 응집체 성숙화와 해독화에 있어 스핑고지질의 역할



filopodia 내에는 수백 개의 myosin 10 분자가 밀집되어 있으며, 이는 actin 위에 교통 체증을 일으킬 수 있다.


Myosin 10 (Myo10) is a motor protein well known for its role in filopodia formation. Although Myo10-driven filopodial dynamics have been characterized, there is no information about the absolute number of Myo10 molecules during the filopodial lifecycle. To better understand molecular stoichiometries and packing restraints in filopodia, we measured Myo10 abundance in these structures. Here we combined SDS-PAGE densitometry with epifluorescence microscopy to quantitate HaloTag-labeled Myo10 in U2OS cells. About 6% of total intracellular Myo10 localizes to filopodia, where it is enriched at opposite ends of the cell. Hundreds of Myo10 are found in a typical filopodium, and their distribution across filopodia is log-normal. Some filopodial tips even contain more Myo10 than accessible binding sites on the actin filament bundle. Live-cell movies reveal a dense cluster of over a hundred Myo10 molecules that initiates filopodial elongation. Hundreds of Myo10 molecules continue to accumulate during filopodial growth, but that accumulation ceases when filopodia begin to retract. Rates of filopodial elongation, second-phase elongation, and retraction are inversely related to Myo10 quantities. Our estimates of Myo10 molecules in filopodia provide insight into the physics of packing Myo10, its cargo, and other filopodia-associated proteins in narrow membrane compartments. Our protocol provides a framework for future work analyzing Myo10 abundance and distribution upon perturbation.

### Competing Interest Statement

RSR is a consultant for Cyntegron Therapeutics.

Pushed to the edge: hundreds of myosin 10s pack into filopodia and could cause traffic jams on actin

세포-내-수백-개의-myosin-10이-filopodia에-밀집되어-있어-actin-위에-교통-체증을-일으킬-수-있다


세포 내 수백 개의 myosin 10이 filopodia에 밀집되어 있어 actin 위에 교통 체증을 일으킬 수 있다



델타-튜불린, 엡실론-튜불린, TEDC1, TEDC2 단백질이 삼중 소관 미세관 구조와 중심체 내부 단백질 복합체 형성에 필수적인 역할을 한다.


Centrioles have a unique, conserved architecture formed by three linked “triplet” microtubules arranged in nine-fold symmetry. The mechanisms by which these triplet microtubules are formed are not understood, but likely involve the noncanonical tubulins delta-tubulin and epsilon-tubulin. Previously, we found that human cells deficient in delta-tubulin or epsilon-tubulin form abnormal centrioles, characterized by an absence of triplet microtubules, lack of central core protein POC5, and a futile cycle of centriole formation and disintegration ([Wang et al., 2017][1]). Here, we show that human cells lacking either of the associated proteins TEDC1 and TEDC2 have these same phenotypes. Using ultrastructure expansion microscopy, we identified the roles of these proteins and triplet microtubules in centriole architecture by mapping the locations of centriolar proteins throughout the cell cycle. We find that mutant centrioles have normal architecture during S-phase. By G2-phase, mutant centrioles grow to the same length as control centrioles, but fail to recruit inner scaffold proteins of the central core. Instead, the inner lumen of centrioles is filled with an expanded proximal region, indicating that these proteins, or the triplet microtubules themselves, may be required for recruiting central core proteins and restricting the length of the proximal end. During mitosis, the mutant centrioles elongate further before fragmenting and disintegrating. All four proteins physically interact and TEDC1 and TEDC2 are capable of interacting in the absence of the tubulins. These results support an AlphaFold Multimer structural prediction model for the tetrameric complex, in which delta-tubulin and epsilon-tubulin are predicted to form a heterodimer. TEDC1 and TEDC2 localize to centrosomes and are mutually dependent on each other and on delta-tubulin and epsilon-tubulin for localization. These results indicate that delta-tubulin, epsilon-tubulin, TEDC1, and TEDC2 function together in promoting robust centriole architecture. This work also lays the groundwork for future dissection of this complex, which will provide a basis for determining the mechanisms that underlie the assembly and interplay between compound microtubules and inner centriole structure.

### Competing Interest Statement

The authors have declared no competing interest.

 [1]: #ref-54

A delta-tubulin/epsilon-tubulin/Ted protein complex is required for centriole architecture

삼중-소관-미세관-구조를-위한-델타-튜불린-엡실론-튜불린-ted-단백질-복합체의-필요성


삼중 소관 미세관 구조를 위한 델타-튜불린/엡실론-튜불린/Ted 단백질 복합체의 필요성



암유발 유전자 유도 노화 과정에서 TPR은 세포질 크로마틴 단편 형성과 이를 통한 NF-κB 신호 활성화에 필수적이다.


During oncogene-induced senescence there are striking changes in the structure of the nucleus and the organisation of heterochromatin. This is accompanied by activation of a pro-inflammatory gene expression programme – the senescence associated secretory phenotype (SASP) – driven by transcription factors such as NF-κB. Here we show that TPR, a protein of the nuclear pore complex basket, is required for the very early activation of NF-κB signalling during the stress-response phase of oncogene-induced senescence. This is prior to activation of the SASP and occurs without affecting NF-κB nuclear import. We show that TPR is required for the activation of TBK1 signalling at these early stages of senescence and we link this to the formation of heterochromatin-enriched cytoplasmic chromatin fragments thought to bleb off from the nuclear periphery. These cytoplasmic chromatin fragments appear to lack nuclear pore components. Our data suggest that TPR at the nuclear pore is involved in the loss of structural integrity of the nuclear periphery during senescence. We propose that this acts as a trigger for activation of cytoplasmic nucleic acid sensing, NF-κB signalling, and activation of the SASP, during senescence.

### Competing Interest Statement

The authors have declared no competing interest.

TPR is required for cytoplasmic chromatin fragment formation during senescence

암유발-유전자-유도-노화-과정에서-세포질-크로마틴-단편-형성에-tpr이-필요하다


암유발 유전자 유도 노화 과정에서 세포질 크로마틴 단편 형성에 TPR이 필요하다



세포 이동은 세포 내 신호 전달의 전방-후방 극성화에 의해 구동되며, 소포체-세포막 접촉 부위의 구배가 이러한 극성화를 매개하여 세포 이동을 지시한다.


The density and size of contact gradients between the endoplasmic reticulum and the plasma membrane control the speed and direction of cell migration by restricting receptor signalling to the front.

Endoplasmic reticulum–plasma membrane contact gradients direct cell migration - Nature

세포-이동을-이끄는-소포체-세포막-접촉-구배


세포 이동을 이끄는 소포체-세포막 접촉 구배
