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1.
Phys Rev Lett ; 126(8): 080601, 2021 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-33709722

RESUMO

The energy dissipation rate in a nonequilibrium reaction system can be determined by the reaction rates in the underlying reaction network. By developing a coarse-graining process in state space and a corresponding renormalization procedure for reaction rates, we find that energy dissipation rate has an inverse power-law dependence on the number of microscopic states in a coarse-grained state. The dissipation scaling law requires self-similarity of the underlying network, and the scaling exponent depends on the network structure and the probability flux correlation. Existence of the inverse dissipation scaling law is shown in realistic biochemical systems such as biochemical oscillators and microtubule-kinesin active flow systems.


Assuntos
Modelos Teóricos , Metabolismo Energético , Entropia , Cinesina/química , Cinesina/metabolismo , Cinética , Microtúbulos/química , Microtúbulos/metabolismo
2.
Nat Commun ; 12(1): 1547, 2021 03 11.
Artigo em Inglês | MEDLINE | ID: mdl-33707436

RESUMO

Hypertension, exercise, and pregnancy are common triggers of cardiac remodeling, which occurs primarily through the hypertrophy of individual cardiomyocytes. During hypertrophy, stress-induced signal transduction increases cardiomyocyte transcription and translation, which promotes the addition of new contractile units through poorly understood mechanisms. The cardiomyocyte microtubule network is also implicated in hypertrophy, but via an unknown role. Here, we show that microtubules are indispensable for cardiac growth via spatiotemporal control of the translational machinery. We find that the microtubule motor Kinesin-1 distributes mRNAs and ribosomes along microtubule tracks to discrete domains within the cardiomyocyte. Upon hypertrophic stimulation, microtubules redistribute mRNAs and new protein synthesis to sites of growth at the cell periphery. If the microtubule network is disrupted, mRNAs and ribosomes collapse around the nucleus, which results in mislocalized protein synthesis, the rapid degradation of new proteins, and a failure of growth, despite normally increased translation rates. Together, these data indicate that mRNAs and ribosomes are actively transported to specific sites to facilitate local translation and assembly of contractile units, and suggest that properly localized translation - and not simply translation rate - is a critical determinant of cardiac hypertrophy. In this work, we find that microtubule based-transport is essential to couple augmented transcription and translation to productive cardiomyocyte growth during cardiac stress.


Assuntos
Cardiomegalia/patologia , Microtúbulos/metabolismo , Miócitos Cardíacos/patologia , Biossíntese de Proteínas/fisiologia , RNA Mensageiro/metabolismo , Ribossomos/metabolismo , Animais , Remodelamento Atrial/fisiologia , Transporte Biológico/fisiologia , Células Cultivadas , Humanos , Cinesina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ratos , Transdução de Sinais/fisiologia , Remodelação Ventricular/fisiologia
3.
Nat Commun ; 12(1): 1401, 2021 03 03.
Artigo em Inglês | MEDLINE | ID: mdl-33658516

RESUMO

Effective treatments for patients suffering from heat hypersensitivity are lacking, mostly due to our limited understanding of the pathogenic mechanisms underlying this disorder. In the nervous system, activating transcription factor 4 (ATF4) is involved in the regulation of synaptic plasticity and memory formation. Here, we show that ATF4 plays an important role in heat nociception. Indeed, loss of ATF4 in mouse dorsal root ganglion (DRG) neurons selectively impairs heat sensitivity. Mechanistically, we show that ATF4 interacts with transient receptor potential cation channel subfamily M member-3 (TRPM3) and mediates the membrane trafficking of TRPM3 in DRG neurons in response to heat. Loss of ATF4 also significantly decreases the current and KIF17-mediated trafficking of TRPM3, suggesting that the KIF17/ATF4/TRPM3 complex is required for the neuronal response to heat stimuli. Our findings unveil the non-transcriptional role of ATF4 in the response to heat stimuli in DRG neurons.


Assuntos
Fator 4 Ativador da Transcrição/metabolismo , Nociceptividade/fisiologia , Células Receptoras Sensoriais/fisiologia , Canais de Cátion TRPM/metabolismo , Fator 4 Ativador da Transcrição/genética , Animais , Membrana Celular/metabolismo , Quimiocina CXCL12/metabolismo , Gânglios Espinais/citologia , Gânglios Espinais/fisiologia , Células HEK293 , Temperatura Alta , Humanos , Injeções Espinhais , Cinesina/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Técnicas de Patch-Clamp , Transporte Proteico , Receptores CXCR4/metabolismo , Canais de Cátion TRPM/genética
4.
Nat Commun ; 12(1): 1463, 2021 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-33674590

RESUMO

Kinesin-1 is a processive motor protein that uses ATP-derived energy to transport a variety of intracellular cargoes toward the cell periphery. The ability to visualize and monitor kinesin transport in live cells is critical to study the myriad of functions associated with cargo trafficking. Herein we report the discovery of a fluorogenic small molecule substrate (QPD-OTf) for kinesin-1 that yields a precipitating dye along its walking path on microtubules (MTs). QPD-OTf enables to monitor native kinesin-1 transport activity in cellulo without external modifications. In vitro assays show that kinesin-1 and MTs are sufficient to yield fluorescent crystals; in cells, kinesin-1 specific transport of cargo from the Golgi appears as trails of fluorescence over time. These findings are further supported by docking studies, which suggest the binding of the activity-based substrate in the nucleotide binding site of kinesin-1.


Assuntos
Cinesina/química , Cinesina/metabolismo , Microtúbulos/metabolismo , Trifosfato de Adenosina , Animais , Sítios de Ligação , Complexo de Golgi/metabolismo , Células HEK293 , Células HeLa , Humanos , Cinesina/genética , Camundongos , Paclitaxel , Transporte Proteico , Células RAW 264.7
5.
Int J Mol Sci ; 22(4)2021 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-33557020

RESUMO

KIF1A is a kinesin family protein that moves over a long distance along the microtubule (MT) to transport synaptic vesicle precursors in neurons. A single KIF1A molecule can move toward the plus-end of MT in the monomeric form, exhibiting the characteristics of biased Brownian motion. However, how the bias is generated in the Brownian motion of KIF1A has not yet been firmly established. To elucidate this, we conducted a set of molecular dynamics simulations and observed the binding of KIF1A to MT. We found that KIF1A exhibits biased Brownian motion along MT as it binds to MT. Furthermore, we show that the bias toward the plus-end is generated by the ratchet-like energy landscape for the KIF1A-MT interaction, in which the electrostatic interaction and the negatively-charged C-terminal tail (CTT) of tubulin play an essential role. The relevance to the post-translational modifications of CTT is also discussed.


Assuntos
Fenômenos Biofísicos , Cinesina/química , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Domínios e Motivos de Interação entre Proteínas , Tubulina (Proteína)/química , Algoritmos , Transporte Axonal , Cinesina/metabolismo , Microtúbulos/química , Microtúbulos/metabolismo , Modelos Teóricos , Neurônios/metabolismo , Ligação Proteica , Tubulina (Proteína)/metabolismo
6.
Virulence ; 12(1): 615-629, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-33538234

RESUMO

It is now clear that the intercellular transport on microtubules by dynein and kinesin-1 motors has an important role in the replication and spread of many viruses. Porcine epidemic diarrhea virus (PEDV) is an enveloped, single-stranded RNA virus of the Coronavirus family, which can infect swine of all ages and cause severe economic losses in the swine industry. Elucidating the molecular mechanisms of the intercellular transport of PEDV through microtubule, dynein and kinesin-1 will be crucial for understanding its pathogenesis. Here, we demonstrate that microtubule, dynein, and kinesin-1 are involved in PEDV infection and can influence PEDV fusion and accumulation in the perinuclear region but cannot affect PEDV attachment or internalization. Furthermore, we adopted a single-virus tracking technique to dynamically observe PEDV intracellular transport with five different types: unidirectional movement toward microtubule plus ends; unidirectional movement toward microtubule minus ends; bidirectional movement along the same microtubule; bidirectional movement along different microtubules and motionless state. Among these types, the functions of dynein and kinesin-1 in PEDV intercellular transport were further analyzed by single-virus tracking and found that dynein and kinesin-1 mainly transport PEDV to the minus and plus ends of the microtubules, respectively; meanwhile, they also can transport PEDV to the opposite ends of the microtubules different from their conventional transport directions and also coordinate the bidirectional movement of PEDV along the same or different microtubules through their cooperation. These results provided deep insights and references to understand the pathogenesis of PEDV as well as to develop vaccines and treatments.


Assuntos
Dineínas/metabolismo , Cinesina/metabolismo , Microtúbulos/metabolismo , Vírus da Diarreia Epidêmica Suína/fisiologia , Animais , Transporte Biológico , Chlorocebus aethiops , Citoplasma/metabolismo , Dineínas/antagonistas & inibidores , Cinesina/genética , Fusão de Membrana , Microscopia de Fluorescência , RNA Interferente Pequeno , Células Vero
7.
Nat Commun ; 12(1): 1213, 2021 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-33619254

RESUMO

Chromosomal instability (CIN) is a hallmark of tumor cells caused by changes in the dynamics and control of microtubules that compromise the mitotic spindle. Thus, CIN cells may respond differently than diploid cells to treatments that target mitotic spindle regulation. Here, we test this idea by inhibiting a subset of kinesin motor proteins involved in mitotic spindle control. KIF18A is required for proliferation of CIN cells derived from triple negative breast cancer or colorectal cancer tumors but is not required in near-diploid cells. Following KIF18A inhibition, CIN tumor cells exhibit mitotic delays, multipolar spindles, and increased cell death. Sensitivity to KIF18A knockdown is strongly correlated with centrosome fragmentation, which requires dynamic microtubules but does not depend on bipolar spindle formation or mitotic arrest. Our results indicate the altered spindle microtubule dynamics characteristic of CIN tumor cells can be exploited to reduce the proliferative capacity of CIN cells.


Assuntos
Instabilidade Cromossômica , Cinesina/metabolismo , Neoplasias/genética , Neoplasias/patologia , Pontos de Checagem do Ciclo Celular , Morte Celular , Linhagem Celular Tumoral , Proliferação de Células , Centrossomo/metabolismo , Humanos , Microtúbulos/metabolismo , Mitose , Modelos Biológicos , Nocodazol/farmacologia , Paclitaxel/farmacologia , Fuso Acromático/metabolismo
8.
Science ; 371(6530)2021 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-33574186

RESUMO

Kinesin motors are essential for the transport of cellular cargo along microtubules. How the motors step, detach, and cooperate with each other is still unclear. To dissect the molecular motion of kinesin-1, we developed germanium nanospheres as ultraresolution optical trapping probes. We found that single motors took 4-nanometer center-of-mass steps. Furthermore, kinesin-1 never detached from microtubules under hindering load conditions. Instead, it slipped on microtubules in microsecond-long, 8-nanometer steps and remained in this slip state before detaching or reengaging in directed motion. Unexpectedly, reengagement and thus rescue of directed motion was more frequent. Our observations broaden our knowledge on the mechanochemical cycle and slip state of kinesin. This state and rescue need to be accounted for to understand long-range transport by teams of motors.


Assuntos
Germânio , Cinesina/química , Cinesina/metabolismo , Nanosferas , Pinças Ópticas , Trifosfato de Adenosina/metabolismo , Transporte Biológico , Cinética , Bicamadas Lipídicas , Microtúbulos/metabolismo , Modelos Biológicos , Imagem Individual de Molécula
9.
J Mol Biol ; 433(5): 166765, 2021 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-33484719

RESUMO

Human PEX14 plays a dual role as docking protein in peroxisomal protein import and as peroxisomal anchor for microtubules (MT), which relates to peroxisome motility. For docking, the conserved N-terminal domain of PEX14 (PEX14-NTD) binds amphipathic alpha-helical ligands, typically comprising one or two aromatic residues, of which human PEX5 possesses eight. Here, we show that the PEX14-NTD also binds to microtubular filaments in vitro with a dissociation constant in nanomolar range. PEX14 interacts with two motifs in the C-terminal region of human ß-tubulin. At least one of the binding motifs is in spatial proximity to the binding site of microtubules (MT) for kinesin. Both PEX14 and kinesin can bind to MT simultaneously. Notably, binding of PEX14 to tubulin can be prevented by its association with PEX5. The data suggest that PEX5 competes peroxisome anchoring to MT by occupying the ß-tubulin-binding site of PEX14. The competitive correlation of matrix protein import and motility may facilitate the homogeneous dispersion of peroxisomes in mammalian cells.


Assuntos
Proteínas de Membrana/química , Microtúbulos/metabolismo , Receptor 1 de Sinal de Orientação para Peroxissomos/química , Peroxissomos/metabolismo , Proteínas Repressoras/química , Tubulina (Proteína)/química , Sequência de Aminoácidos , Sítios de Ligação , Ligação Competitiva , Transporte Biológico , Linhagem Celular , Escherichia coli/genética , Escherichia coli/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Expressão Gênica , Humanos , Cinesina/genética , Cinesina/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Modelos Moleculares , Receptor 1 de Sinal de Orientação para Peroxissomos/genética , Receptor 1 de Sinal de Orientação para Peroxissomos/metabolismo , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Transdução de Sinais , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismo
10.
Proc Natl Acad Sci U S A ; 118(1)2021 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-33443153

RESUMO

The differentiation of cells depends on a precise control of their internal organization, which is the result of a complex dynamic interplay between the cytoskeleton, molecular motors, signaling molecules, and membranes. For example, in the developing neuron, the protein ADAP1 (ADP-ribosylation factor GTPase-activating protein [ArfGAP] with dual pleckstrin homology [PH] domains 1) has been suggested to control dendrite branching by regulating the small GTPase ARF6. Together with the motor protein KIF13B, ADAP1 is also thought to mediate delivery of the second messenger phosphatidylinositol (3,4,5)-trisphosphate (PIP3) to the axon tip, thus contributing to PIP3 polarity. However, what defines the function of ADAP1 and how its different roles are coordinated are still not clear. Here, we studied ADAP1's functions using in vitro reconstitutions. We found that KIF13B transports ADAP1 along microtubules, but that PIP3 as well as PI(3,4)P2 act as stop signals for this transport instead of being transported. We also demonstrate that these phosphoinositides activate ADAP1's enzymatic activity to catalyze GTP hydrolysis by ARF6. Together, our results support a model for the cellular function of ADAP1, where KIF13B transports ADAP1 until it encounters high PIP3/PI(3,4)P2 concentrations in the plasma membrane. Here, ADAP1 disassociates from the motor to inactivate ARF6, promoting dendrite branching.


Assuntos
Fatores de Ribosilação do ADP/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Fosfatidilinositóis/metabolismo , Fatores de Ribosilação do ADP/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Animais , Axônios/metabolismo , Transporte Biológico/fisiologia , Membrana Celular/metabolismo , Citoesqueleto/metabolismo , Proteínas Ativadoras de GTPase/metabolismo , Humanos , Fosfatos de Inositol/metabolismo , Cinesina/metabolismo , Microtúbulos/metabolismo , Proteínas do Tecido Nervoso/fisiologia , Fosfatos de Fosfatidilinositol/metabolismo , Transdução de Sinais
11.
Nature ; 590(7846): 486-491, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33505028

RESUMO

Selective targeting of aneuploid cells is an attractive strategy for cancer treatment1. However, it is unclear whether aneuploidy generates any clinically relevant vulnerabilities in cancer cells. Here we mapped the aneuploidy landscapes of about 1,000 human cancer cell lines, and analysed genetic and chemical perturbation screens2-9 to identify cellular vulnerabilities associated with aneuploidy. We found that aneuploid cancer cells show increased sensitivity to genetic perturbation of core components of the spindle assembly checkpoint (SAC), which ensures the proper segregation of chromosomes during mitosis10. Unexpectedly, we also found that aneuploid cancer cells were less sensitive than diploid cells to short-term exposure to multiple SAC inhibitors. Indeed, aneuploid cancer cells became increasingly sensitive to inhibition of SAC over time. Aneuploid cells exhibited aberrant spindle geometry and dynamics, and kept dividing when the SAC was inhibited, resulting in the accumulation of mitotic defects, and in unstable and less-fit karyotypes. Therefore, although aneuploid cancer cells could overcome inhibition of SAC more readily than diploid cells, their long-term proliferation was jeopardized. We identified a specific mitotic kinesin, KIF18A, whose activity was perturbed in aneuploid cancer cells. Aneuploid cancer cells were particularly vulnerable to depletion of KIF18A, and KIF18A overexpression restored their response to SAC inhibition. Our results identify a therapeutically relevant, synthetic lethal interaction between aneuploidy and the SAC.


Assuntos
Aneuploidia , Pontos de Checagem da Fase M do Ciclo Celular/efeitos dos fármacos , Neoplasias/patologia , Cariótipo Anormal/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Segregação de Cromossomos/efeitos dos fármacos , Diploide , Genes Letais , Humanos , Cinesina/deficiência , Cinesina/genética , Cinesina/metabolismo , Neoplasias/genética , Fuso Acromático/efeitos dos fármacos , Mutações Sintéticas Letais/efeitos dos fármacos , Mutações Sintéticas Letais/genética , Fatores de Tempo
12.
Nature ; 590(7846): 492-497, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33505027

RESUMO

Whole-genome doubling (WGD) is common in human cancers, occurring early in tumorigenesis and generating genetically unstable tetraploid cells that fuel tumour development1,2. Cells that undergo WGD (WGD+ cells) must adapt to accommodate their abnormal tetraploid state; however, the nature of these adaptations, and whether they confer vulnerabilities that can be exploited therapeutically, is unclear. Here, using sequencing data from roughly 10,000 primary human cancer samples and essentiality data from approximately 600 cancer cell lines, we show that WGD gives rise to common genetic traits that are accompanied by unique vulnerabilities. We reveal that WGD+ cells are more dependent than WGD- cells on signalling from the spindle-assembly checkpoint, DNA-replication factors and proteasome function. We also identify KIF18A, which encodes a mitotic kinesin protein, as being specifically required for the viability of WGD+ cells. Although KIF18A is largely dispensable for accurate chromosome segregation during mitosis in WGD- cells, its loss induces notable mitotic errors in WGD+ cells, ultimately impairing cell viability. Collectively, our results suggest new strategies for specifically targeting WGD+ cancer cells while sparing the normal, non-transformed WGD- cells that comprise human tissue.


Assuntos
Genoma Humano/genética , Mitose/efeitos dos fármacos , Neoplasias/genética , Neoplasias/patologia , Tetraploidia , Cariótipo Anormal/efeitos dos fármacos , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Feminino , Genes Letais/genética , Humanos , Cinesina/deficiência , Cinesina/genética , Cinesina/metabolismo , Pontos de Checagem da Fase M do Ciclo Celular/efeitos dos fármacos , Masculino , Mitose/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Reprodutibilidade dos Testes , Fuso Acromático/efeitos dos fármacos
13.
Nat Commun ; 12(1): 20, 2021 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-33397932

RESUMO

Drug resistance and tumor recurrence are major challenges in cancer treatment. Cancer cells often display centrosome amplification. To maintain survival, cancer cells achieve bipolar division by clustering supernumerary centrosomes. Targeting centrosome clustering is therefore considered a promising therapeutic strategy. However, the regulatory mechanisms of centrosome clustering remain unclear. Here we report that KIFC1, a centrosome clustering regulator, is positively associated with tumor recurrence. Under DNA damaging treatments, the ATM and ATR kinases phosphorylate KIFC1 at Ser26 to selectively maintain the survival of cancer cells with amplified centrosomes via centrosome clustering, leading to drug resistance and tumor recurrence. Inhibition of KIFC1 phosphorylation represses centrosome clustering and tumor recurrence. This study identified KIFC1 as a prognostic tumor recurrence marker, and revealed that tumors can acquire therapeutic resistance and recurrence via triggering centrosome clustering under DNA damage stresses, suggesting that blocking KIFC1 phosphorylation may open a new vista for cancer therapy.


Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Centrossomo/metabolismo , Cinesina/metabolismo , Recidiva Local de Neoplasia/metabolismo , Sequência de Aminoácidos , Animais , Linhagem Celular Tumoral , Instabilidade Cromossômica , Dano ao DNA , Resistencia a Medicamentos Antineoplásicos , Humanos , Cinesina/química , Camundongos , Recidiva Local de Neoplasia/patologia , Fosforilação , Fosfosserina/metabolismo
14.
Biochem Pharmacol ; 184: 114364, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33310050

RESUMO

Eg5, the product of Kif11 gene, also known as kinesin spindle protein, is a motor protein involved in the proper establishment of a bipolar mitotic spindle. Eg5 is one of the 45 different kinesins coded in the human genome of the kinesin motor protein superfamily. Over the last three decades Eg5 has attracted great interest as a promising new mitotic target. The identification of monastrol as specific inhibitor of the ATPase activity of the motor domain of Eg5 inhibiting the Eg5 microtubule motility in vitro and in cellulo sparked an intense interest in academia and industry to pursue the identification of novel small molecules that target Eg5 in order to be used in cancer chemotherapy based on the anti-mitotic strategy. Several Eg5 inhibitors entered clinical trials. Currently the field is faced with the problem that most of the inhibitors tested exhibited only limited efficacy. However, one Eg5 inhibitor, Arry-520 (clinical name filanesib), has demonstrated clinical efficacy in patients with multiple myeloma and is scheduled to enter phase III clinical trials. At the same time, new trends in Eg5 inhibitor research are emerging, including an increased interest in novel inhibitor binding sites and a focus on drug synergy with established antitumor agents to improve chemotherapeutic efficacy. This review presents an updated view of the structure and function of Eg5-inhibitor complexes, traces the possible development of resistance to Eg5 inhibitors and their potential therapeutic applications, and surveys the current challenges and future directions of this active field in drug discovery.


Assuntos
Antimitóticos/farmacologia , Antineoplásicos/farmacologia , Cinesina/antagonistas & inibidores , Cinesina/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Antimitóticos/química , Antimitóticos/farmacocinética , Antineoplásicos/farmacocinética , Sítios de Ligação , Produtos Biológicos/química , Produtos Biológicos/farmacologia , Ensaios Clínicos como Assunto , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/fisiologia , Humanos , Cinesina/química , Terapia de Alvo Molecular/métodos
15.
Elife ; 92020 12 21.
Artigo em Inglês | MEDLINE | ID: mdl-33346730

RESUMO

When a T cell and an antigen-presenting cell form an immunological synapse, rapid dynein-driven translocation of the centrosome toward the contact site leads to reorganization of microtubules and associated organelles. Currently, little is known about how the regulation of microtubule dynamics contributes to this process. Here, we show that the knockout of KIF21B, a kinesin-4 linked to autoimmune disorders, causes microtubule overgrowth and perturbs centrosome translocation. KIF21B restricts microtubule length by inducing microtubule pausing typically followed by catastrophe. Catastrophe induction with vinblastine prevented microtubule overgrowth and was sufficient to rescue centrosome polarization in KIF21B-knockout cells. Biophysical simulations showed that a relatively small number of KIF21B molecules can restrict mirotubule length and promote an imbalance of dynein-mediated pulling forces that allows the centrosome to translocate past the nucleus. We conclude that proper control of microtubule length is important for allowing rapid remodeling of the cytoskeleton and efficient T cell polarization.


Assuntos
Centrossomo/metabolismo , Cinesina/metabolismo , Microtúbulos/metabolismo , Linfócitos T/imunologia , Actinas/metabolismo , Células Apresentadoras de Antígenos/imunologia , Citoesqueleto/metabolismo , Humanos , Sinapses Imunológicas/metabolismo , Ativação Linfocitária
16.
Cell Death Dis ; 11(12): 1056, 2020 12 12.
Artigo em Inglês | MEDLINE | ID: mdl-33311452

RESUMO

RNA-binding proteins (RBPs) predominantly contribute to abnormal posttranscriptional gene modulation and disease progression in cancer. Sorbin and SH3 domain-containing 2 (SORBS2), an RBP, has been reported to be a potent tumor suppressor in several cancer types. Through integrative analysis of clinical specimens, we disclosed that the expression level of SORBS2 was saliently decreased in metastatic tissues and positively correlated with overall survival. We observed that overexpression of SORBS2 brought about decreased metastatic capacity in ccRCC cell lines. Transcriptome-wide analysis revealed that SORBS2 notably increased microtubule-associated tumor-suppressor 1 gene (MTUS1) expression. In-depth mechanistic exploring discovered that the Cys2-His2 zinc finger (C2H2-ZnF) domain of SORBS2 directly bound to the 3' untranslated region (3'UTR) of MTUS1 mRNA, which increased MTUS1 mRNA stability. In addition, we identified that MTUS1 regulated microtubule dynamics via promoting KIF2CS192 phosphorylation by Aurora B. Together, our research identified SORBS2 as a suppressor of ccRCC metastasis by enhancing MTUS1 mRNA stability, providing a novel understanding of RBPs during ccRCC progression.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Carcinoma de Células Renais/genética , Estabilidade de RNA/genética , Proteínas de Ligação a RNA/metabolismo , Proteínas Supressoras de Tumor/genética , Regiões 3' não Traduzidas/genética , Aurora Quinase B/metabolismo , Linhagem Celular Tumoral , Regulação para Baixo/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Cinesina/metabolismo , Masculino , Microtúbulos/metabolismo , Pessoa de Meia-Idade , Fosforilação , Prognóstico , Ligação Proteica/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Análise de Sobrevida , Proteínas Supressoras de Tumor/metabolismo , Regulação para Cima/genética
17.
Nat Commun ; 11(1): 5510, 2020 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-33139737

RESUMO

In living cells, dynamics of the endoplasmic reticulum (ER) are driven by the cytoskeleton motor machinery as well as the action of ER-shaping proteins such as atlastin GTPases including RHD3 in Arabidopsis. It is not known if the two systems interplay, and, if so, how they do. Here we report the identification of ARK1 (Armadillo-Repeat Kinesin1) via a genetic screen for enhancers of the rhd3 mutant phenotype. In addition to defects in microtubule dynamics, ER organization is also defective in mutants lacking a functional ARK1. In growing root hair cells, ARK1 comets predominantly localize on the growing-end of microtubules and partially overlap with RHD3 in the cortex of the subapical region. ARK1 co-moves with RHD3 during tip growth of root hair cells. We show that there is a functional interdependence between ARK1 and RHD3. ARK1 physically interacts with RHD3 via its armadillo domain (ARM). In leaf epidermal cells where a polygonal ER network can be resolved, ARK1, but not ARK1ΔARM, moves together with RHD3 to pull an ER tubule toward another and stays with the newly formed 3-way junction of the ER for a while. We conclude that ARK1 acts together with RHD3 to move the ER on microtubules to generate a fine ER network.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Cinesina/metabolismo , Microtúbulos/metabolismo , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Proteínas do Domínio Armadillo , Proteínas de Ligação ao GTP/genética , Mutação , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas
18.
Nat Commun ; 11(1): 4471, 2020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32901010

RESUMO

A human cell contains hundreds to thousands of mitochondrial DNA (mtDNA) packaged into nucleoids. Currently, the segregation and allocation of nucleoids are thought to be passively determined by mitochondrial fusion and division. Here we provide evidence, using live-cell super-resolution imaging, that nucleoids can be actively transported via KIF5B-driven mitochondrial dynamic tubulation (MDT) activities that predominantly occur at the ER-mitochondria contact sites (EMCS). We further demonstrate that a mitochondrial inner membrane protein complex MICOS links nucleoids to Miro1, a KIF5B receptor on mitochondria, at the EMCS. We show that such active transportation is a mechanism essential for the proper distribution of nucleoids in the peripheral zone of the cell. Together, our work identifies an active transportation mechanism of nucleoids, with EMCS serving as a key platform for the interplay of nucleoids, MICOS, Miro1, and KIF5B to coordinate nucleoids segregation and transportation.


Assuntos
DNA Mitocondrial/metabolismo , Retículo Endoplasmático/metabolismo , Mitocôndrias/metabolismo , Dinâmica Mitocondrial/fisiologia , Animais , Transporte Biológico Ativo , Células COS , Células Cultivadas , Chlorocebus aethiops , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Células HEK293 , Humanos , Cinesina/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Camundongos , Membranas Mitocondriais/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Modelos Biológicos , Ratos , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transfecção , Proteínas rho de Ligação ao GTP/metabolismo
19.
PLoS Biol ; 18(8): e3000820, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32866173

RESUMO

Mutations in the gene encoding the microtubule-severing protein spastin (spastic paraplegia 4 [SPG4]) cause hereditary spastic paraplegia (HSP), associated with neurodegeneration, spasticity, and motor impairment. Complicated forms (complicated HSP [cHSP]) further include cognitive deficits and dementia; however, the etiology and dysfunctional mechanisms of cHSP have remained unknown. Here, we report specific working and associative memory deficits upon spastin depletion in mice. Loss of spastin-mediated severing leads to reduced synapse numbers, accompanied by lower miniature excitatory postsynaptic current (mEPSC) frequencies. At the subcellular level, mutant neurons are characterized by longer microtubules with increased tubulin polyglutamylation levels. Notably, these conditions reduce kinesin-microtubule binding, impair the processivity of kinesin family protein (KIF) 5, and reduce the delivery of presynaptic vesicles and postsynaptic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. Rescue experiments confirm the specificity of these results by showing that wild-type spastin, but not the severing-deficient and disease-associated K388R mutant, normalizes the effects at the synaptic, microtubule, and transport levels. In addition, short hairpin RNA (shRNA)-mediated reduction of tubulin polyglutamylation on spastin knockout background normalizes KIF5 transport deficits and attenuates the loss of excitatory synapses. Our data provide a mechanism that connects spastin dysfunction with the regulation of kinesin-mediated cargo transport, synapse integrity, and cognition.


Assuntos
Ácido Glutâmico/metabolismo , Cinesina/metabolismo , Transtornos da Memória/metabolismo , Transtornos da Memória/fisiopatologia , Memória de Curto Prazo , Neurônios/metabolismo , Espastina/deficiência , Tubulina (Proteína)/metabolismo , Potenciais de Ação , Animais , Membrana Celular/metabolismo , Espinhas Dendríticas/metabolismo , Espinhas Dendríticas/ultraestrutura , Potenciais Pós-Sinápticos Excitadores , Hipocampo/patologia , Hipocampo/fisiopatologia , Camundongos Knockout , Microtúbulos/metabolismo , Microtúbulos/ultraestrutura , Atividade Motora , Neurônios/patologia , Neurônios/ultraestrutura , Transporte Proteico , Espastina/metabolismo , Sinapses/metabolismo , Sinapses/ultraestrutura , Vesículas Sinápticas/metabolismo
20.
Nat Commun ; 11(1): 4092, 2020 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-32796837

RESUMO

Single nucleotide polymorphisms (SNPs) in the gene encoding kinesin family member 3A, KIF3A, have been associated with atopic dermatitis (AD), a chronic inflammatory skin disorder. We find that KIF3A SNP rs11740584 and rs2299007 risk alleles create cytosine-phosphate-guanine sites, which are highly methylated and result in lower KIF3A expression, and this methylation is associated with increased transepidermal water loss (TEWL) in risk allele carriers. Kif3aK14∆/∆ mice have increased TEWL, disrupted junctional proteins, and increased susceptibility to develop AD. Thus, KIF3A is required for skin barrier homeostasis whereby decreased KIF3A skin expression causes disrupted skin barrier function and promotes development of AD.


Assuntos
Dermatite Atópica/metabolismo , Cinesina/metabolismo , Pele/metabolismo , Adolescente , Adulto , Alelos , Animais , Criança , Dermatite Atópica/genética , Dermatite Atópica/patologia , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Cinesina/genética , Masculino , Metilação , Camundongos , Reação em Cadeia da Polimerase em Tempo Real , Pele/patologia , Adulto Jovem
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