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1.
Annu Int Conf IEEE Eng Med Biol Soc ; 2020: 1811-1815, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-33018351

RESUMO

Automated mitotic detection in time-lapse phase-contrast microscopy provides us much information for cell behavior analysis, and thus several mitosis detection methods have been proposed. However, these methods still have two problems; 1) they cannot detect multiple mitosis events when there are closely placed. 2) they do not consider the annotation gaps, which may occur since the appearances of mitosis cells are very similar before and after the annotated frame. In this paper, we propose a novel mitosis detection method that can detect multiple mitosis events in a candidate sequence and mitigate the human annotation gap via estimating spatial-temporal likelihood map by 3DCNN. In this training, the loss gradually decreases with the gap size between ground-truth and estimation. This mitigates the annotation gaps. Our method outperformed the compared methods in terms of F1-score using challenging dataset that contains the data under four different conditions. Code is publicly available in https://github.com/naivete5656/MDMLM.


Assuntos
Algoritmos , Mitose , Humanos , Microscopia de Contraste de Fase , Probabilidade
2.
Nat Commun ; 11(1): 4983, 2020 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-33020492

RESUMO

The energetic demands of a cell are believed to increase during mitosis, but the rates of ATP synthesis and consumption during mitosis have not been quantified. Here, we monitor mitochondrial membrane potential of single lymphocytic leukemia cells and demonstrate that mitochondria hyperpolarize from the G2/M transition until the metaphase-anaphase transition. This hyperpolarization was dependent on cyclin-dependent kinase 1 (CDK1) activity. By using an electrical circuit model of mitochondria, we quantify mitochondrial ATP synthesis rates in mitosis from the single-cell time-dynamics of mitochondrial membrane potential. We find that mitochondrial ATP synthesis decreases by approximately 50% during early mitosis and increases back to G2 levels during cytokinesis. Consistently, ATP levels and ATP synthesis are lower in mitosis than in G2 in synchronized cell populations. Overall, our results provide insights into mitotic bioenergetics and suggest that cell division is not a highly energy demanding process.


Assuntos
Trifosfato de Adenosina/biossíntese , Divisão Celular , Metabolismo Energético , Animais , Proteína Quinase CDC2/metabolismo , Linhagem Celular Tumoral , Citocinese , Camundongos , Mitocôndrias/metabolismo , Mitose , Modelos Biológicos
3.
Anticancer Res ; 40(10): 5471-5480, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32988869

RESUMO

BACKGROUND/AIM: Accurate regulation of the spindle assembly checkpoint (SAC) and anaphase promoting complex/cyclosome (APC/C) are essential for the correct execution of mitosis. In this work, we focused on MAD2L2 (REV7), a central translesion (TLS) protein, which also functions as a mitotic regulator by inhibiting APC/C in prometaphase. MATERIALS AND METHODS: Using bioinformatics analysis, live cell imaging and APC/C protein binding and degradation assays, we explored the influence of MAD2L2 over-expression in breast cancer. RESULTS: A significant over-expression of MAD2L2 was found in triple negative breast cancers (TNBC), compared to other breast cancers, correlating to poor patient prognosis. We also identified significant over-expression of MAD2L2 in the MDA-MB-157 triple negative (TN) cell line. A high percentage of MDA-MB-157 cells failed to complete mitosis and died during mitosis or shortly after. In addition, these cells completed mitosis at a significantly slower rate than control cells. MDA-MB-157 cells present high levels of mitotic slippage upon nocodazole treatment and acute dysregulation in APC/C function and substrate degradation. Moreover, silencing of MAD2L2 in the MDA-MB-157 cell line improved mitotic phenotypes. CONCLUSION: MAD2L2 over-expression supports the carcinogenic phenotype of MDA-MB-157 cells by promoting uncontrolled mitosis.


Assuntos
Neoplasias da Mama/genética , Proteínas Mad2/genética , Mitose/genética , Neoplasias da Mama/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Células HeLa , Humanos , Pontos de Checagem da Fase M do Ciclo Celular/genética , Prometáfase/genética
4.
PLoS Comput Biol ; 16(9): e1007470, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32941445

RESUMO

Human T-lymphotropic virus type-1 (HTLV-1) persists within hosts via infectious spread (de novo infection) and mitotic spread (infected cell proliferation), creating a population structure of multiple clones (infected cell populations with identical genomic proviral integration sites). The relative contributions of infectious and mitotic spread to HTLV-1 persistence are unknown, and will determine the efficacy of different approaches to treatment. The prevailing view is that infectious spread is negligible in HTLV-1 persistence beyond early infection. However, in light of recent high-throughput data on the abundance of HTLV-1 clones, and recent estimates of HTLV-1 clonal diversity that are substantially higher than previously thought (typically between 104 and 105 HTLV-1+ T cell clones in the body of an asymptomatic carrier or patient with HTLV-1-associated myelopathy/tropical spastic paraparesis), ongoing infectious spread during chronic infection remains possible. We estimate the ratio of infectious to mitotic spread using a hybrid model of deterministic and stochastic processes, fitted to previously published HTLV-1 clonal diversity estimates. We investigate the robustness of our estimates using three alternative estimators. We find that, contrary to previous belief, infectious spread persists during chronic infection, even after HTLV-1 proviral load has reached its set point, and we estimate that between 100 and 200 new HTLV-1 clones are created and killed every day. We find broad agreement between all estimators. The risk of HTLV-1-associated malignancy and inflammatory disease is strongly correlated with proviral load, which in turn is correlated with the number of HTLV-1-infected clones, which are created by de novo infection. Our results therefore imply that suppression of de novo infection may reduce the risk of malignant transformation.


Assuntos
Infecções por HTLV-I , Interações Hospedeiro-Patógeno , Vírus Linfotrópico T Tipo 1 Humano , Linfócitos T CD4-Positivos/virologia , Infecções por HTLV-I/fisiopatologia , Infecções por HTLV-I/virologia , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/fisiologia , Vírus Linfotrópico T Tipo 1 Humano/classificação , Vírus Linfotrópico T Tipo 1 Humano/genética , Vírus Linfotrópico T Tipo 1 Humano/patogenicidade , Humanos , Mitose/genética , Mitose/fisiologia , Modelos Biológicos , Provírus/genética , Provírus/patogenicidade , Carga Viral/genética , Integração Viral/genética
5.
Zhongguo Yi Xue Ke Xue Yuan Xue Bao ; 42(4): 485-490, 2020 Aug 30.
Artigo em Chinês | MEDLINE | ID: mdl-32895100

RESUMO

Objective To explore the molecular mechanism of human papillomavirus subtype 16(HPV-16)E7 oncogene-induced DNA re-replication in response to DNA damage. Methods Flow cytometry was performed to examine the cell cycle changes in RPE1 E7 cells stably expressing HPV-16 E7 and its control cell RPE1 Vector after DNA damage.Immunoblotting assay was used to evaluate the early mitotic inhibitor 1(Emi1)expression in RPE1 E7 and RPE1 Vector cells with or without DNA damage.The changes of the proportion of polyploidy was detected by flow cytometry in DNA-damaged RPE1 E7 cells interfered by Emi1 small interfering RNA. Results Compared with the control cells,the proportion of polyploids in RPE1 E7 cells was significantly increased in response to DNA damage(t=6.397,P=0.0031).Emi1 protein expression was significantly increased in DNA damaged RPE1 E7 cells(t=8.241,P=0.0012).The polyploid ratio of RPE1 E7 cells was significantly reduced after Emi1 was interfered by two independent small interfering RNAs(t=2.916,P=0.0434;t=3.452,P=0.0260). Conclusion In response to DNA damage,Emi1 promoted DNA re-replication caused by HPV-16 E7.


Assuntos
Replicação do DNA , Dano ao DNA , Papillomavirus Humano 16 , Mitose , Proteínas Oncogênicas Virais
6.
Nat Commun ; 11(1): 4455, 2020 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-32901005

RESUMO

Dysregulated alternative splicing (AS) driving carcinogenetic mitosis remains poorly understood. Here, we demonstrate that cancer metastasis-associated antigen 1 (MTA1), a well-known oncogenic chromatin modifier, broadly interacts and co-expresses with RBPs across cancers, contributing to cancerous mitosis-related AS. Using developed fCLIP-seq technology, we show that MTA1 binds abundant transcripts, preferentially at splicing-responsible motifs, influencing the abundance and AS pattern of target transcripts. MTA1 regulates the mRNA level and guides the AS of a series of mitosis regulators. MTA1 deletion abrogated the dynamic AS switches of variants for ATRX and MYBL2 at mitotic stage, which are relevant to mitosis-related tumorigenesis. MTA1 dysfunction causes defective mitotic arrest, leads to aberrant chromosome segregation, and results in chromosomal instability (CIN), eventually contributing to tumorigenesis. Currently, little is known about the RNA splicing during mitosis; here, we uncover that MTA1 binds transcripts and orchestrates dynamic splicing of mitosis regulators in tumorigenesis.


Assuntos
Carcinogênese/genética , Carcinogênese/metabolismo , Montagem e Desmontagem da Cromatina/fisiologia , Mitose/fisiologia , RNA Mensageiro/metabolismo , Proteínas Repressoras/metabolismo , Transativadores/metabolismo , Processamento Alternativo , Animais , Sítios de Ligação/genética , Montagem e Desmontagem da Cromatina/genética , Instabilidade Cromossômica , Feminino , Células HCT116 , Xenoenxertos , Humanos , Camundongos , Camundongos Nus , Mitose/genética , Neoplasias/genética , Neoplasias/metabolismo , Precursores de RNA/genética , Precursores de RNA/metabolismo , Processamento Pós-Transcricional do RNA , RNA Mensageiro/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/genética , Transativadores/antagonistas & inibidores , Transativadores/genética
7.
Science ; 369(6505): 858-862, 2020 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-32792401

RESUMO

The conversion of neural stem cells into neurons is associated with the remodeling of organelles, but whether and how this is causally linked to fate change is poorly understood. We examined and manipulated mitochondrial dynamics during mouse and human cortical neurogenesis. We reveal that shortly after cortical stem cells have divided, daughter cells destined to self-renew undergo mitochondrial fusion, whereas those that retain high levels of mitochondria fission become neurons. Increased mitochondria fission promotes neuronal fate, whereas induction of mitochondria fusion after mitosis redirects daughter cells toward self-renewal. This occurs during a restricted time window that is doubled in human cells, in line with their increased self-renewal capacity. Our data reveal a postmitotic period of fate plasticity in which mitochondrial dynamics are linked with cell fate.


Assuntos
Córtex Cerebral/crescimento & desenvolvimento , Mitocôndrias/fisiologia , Dinâmica Mitocondrial , Mitose , Células-Tronco Neurais/citologia , Neurogênese/fisiologia , Neurônios/citologia , Animais , Córtex Cerebral/citologia , Feminino , Células HEK293 , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Humanos , Masculino , Camundongos , Plasticidade Neuronal/efeitos dos fármacos , Plasticidade Neuronal/fisiologia , Sirtuínas/metabolismo
8.
PLoS Genet ; 16(8): e1008990, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32810142

RESUMO

The kinetochore, a multi-protein complex assembled on centromeres, is essential to segregate chromosomes during cell division. Deficiencies in kinetochore function can lead to chromosomal instability and aneuploidy-a hallmark of cancer cells. Kinetochore function is controlled by recruitment of regulatory proteins, many of which have been documented, however their function often remains uncharacterized and many are yet to be identified. To identify candidates of kinetochore regulation we used a proteome-wide protein association strategy in budding yeast and detected many proteins that are involved in post-translational modifications such as kinases, phosphatases and histone modifiers. We focused on the Polo-like kinase, Cdc5, and interrogated which cellular components were sensitive to constitutive Cdc5 localization. The kinetochore is particularly sensitive to constitutive Cdc5 kinase activity. Targeting Cdc5 to different kinetochore subcomplexes produced diverse phenotypes, consistent with multiple distinct functions at the kinetochore. We show that targeting Cdc5 to the inner kinetochore, the constitutive centromere-associated network (CCAN), increases the levels of centromeric RNA via an SPT4 dependent mechanism.


Assuntos
Proteínas de Ciclo Celular/genética , Centrômero/genética , Proteínas Nucleares/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Proto-Oncogênicas/genética , Proteínas de Saccharomyces cerevisiae/genética , Fatores de Elongação da Transcrição/genética , Anáfase/genética , Proteínas Cromossômicas não Histona/genética , Segregação de Cromossomos/genética , Histonas/genética , Humanos , Cinetocoros/metabolismo , Mitose/genética , Fenótipo , Fosforilação/genética , RNA/genética , Saccharomyces cerevisiae/genética
9.
Nature ; 585(7823): 119-123, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32848252

RESUMO

At the end of mitosis, eukaryotic cells must segregate the two copies of their replicated genome into two new nuclear compartments1. They do this either by first dismantling and later reassembling the nuclear envelope in an 'open mitosis' or by reshaping an intact nucleus and then dividing it into two in a 'closed mitosis'2,3. Mitosis has been studied in a wide variety of eukaryotes for more than a century4, but how the double membrane of the nuclear envelope is split into two at the end of a closed mitosis without compromising the impermeability of the nuclear compartment remains unknown5. Here, using the fission yeast Schizosaccharomyces pombe (a classical model for closed mitosis5), genetics, live-cell imaging and electron tomography, we show that nuclear fission is achieved via local disassembly of nuclear pores within the narrow bridge that links segregating daughter nuclei. In doing so, we identify the protein Les1, which is localized to the inner nuclear envelope and restricts the process of local nuclear envelope breakdown to the bridge midzone to prevent the leakage of material from daughter nuclei. The mechanism of local nuclear envelope breakdown in a closed mitosis therefore closely mirrors nuclear envelope breakdown in open mitosis3, revealing an unexpectedly high conservation of nuclear remodelling mechanisms across diverse eukaryotes.


Assuntos
Mitose , Membrana Nuclear/metabolismo , Schizosaccharomyces/citologia , Divisão Celular , Modelos Biológicos , Poro Nuclear/metabolismo , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Schizosaccharomyces/ultraestrutura
10.
PLoS Biol ; 18(8): e3000762, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32760088

RESUMO

Centrosomes, the main microtubule organizing centers (MTOCs) of metazoan cells, contain an older "mother" and a younger "daughter" centriole. Stem cells either inherit the mother or daughter-centriole-containing centrosome, providing a possible mechanism for biased delivery of cell fate determinants. However, the mechanisms regulating centrosome asymmetry and biased centrosome segregation are unclear. Using 3D-structured illumination microscopy (3D-SIM) and live-cell imaging, we show in fly neural stem cells (neuroblasts) that the mitotic kinase Polo and its centriolar protein substrate Centrobin (Cnb) accumulate on the daughter centriole during mitosis, thereby generating molecularly distinct mother and daughter centrioles before interphase. Cnb's asymmetric localization, potentially involving a direct relocalization mechanism, is regulated by Polo-mediated phosphorylation, whereas Polo's daughter centriole enrichment requires both Wdr62 and Cnb. Based on optogenetic protein mislocalization experiments, we propose that the establishment of centriole asymmetry in mitosis primes biased interphase MTOC activity, necessary for correct spindle orientation.


Assuntos
Proteínas de Ciclo Celular/genética , Centríolos/metabolismo , Centrossomo/metabolismo , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Mitose , Proteínas Serina-Treonina Quinases/genética , Animais , Animais Geneticamente Modificados , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Ciclo Celular/metabolismo , Centríolos/ultraestrutura , Centrossomo/ultraestrutura , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citologia , Drosophila melanogaster/crescimento & desenvolvimento , Drosophila melanogaster/metabolismo , Embrião não Mamífero , Regulação da Expressão Gênica no Desenvolvimento , Genes Reporter , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Interfase , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Optogenética/métodos , Fosforilação , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais
11.
Mol Cell ; 79(6): 902-916.e6, 2020 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-32768407

RESUMO

A long-standing conundrum is how mitotic chromosomes can compact, as required for clean separation to daughter cells, while maintaining close parallel alignment of sister chromatids. Pursuit of this question, by high resolution 3D fluorescence imaging of living and fixed mammalian cells, has led to three discoveries. First, we show that the structural axes of separated sister chromatids are linked by evenly spaced "mini-axis" bridges. Second, when chromosomes first emerge as discrete units, at prophase, they are organized as co-oriented sister linear loop arrays emanating from a conjoined axis. We show that this same basic organization persists throughout mitosis, without helical coiling. Third, from prophase onward, chromosomes are deformed into sequential arrays of half-helical segments of alternating handedness (perversions), accompanied by correlated kinks. These arrays fluctuate dynamically over <15 s timescales. Together these discoveries redefine the foundation for thinking about the evolution of mitotic chromosomes as they prepare for anaphase segregation.


Assuntos
Proteínas de Ciclo Celular/genética , Cromossomos/genética , Proteínas de Ligação a DNA/genética , Mitose/genética , Adenosina Trifosfatases/genética , Anáfase/genética , Animais , Proteínas de Ciclo Celular/isolamento & purificação , Cromátides/genética , Proteínas Cromossômicas não Histona , DNA Topoisomerases Tipo II/genética , Proteínas de Ligação a DNA/isolamento & purificação , Imageamento Tridimensional , Mamíferos , Metáfase/genética , Prófase/genética
12.
PLoS Genet ; 16(8): e1008962, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32750047

RESUMO

Haspin, a highly conserved kinase in eukaryotes, has been shown to be responsible for phosphorylation of histone H3 at threonine 3 (H3T3ph) during mitosis, in mammals and yeast. Here we report that haspin is the kinase that phosphorylates H3T3 in Drosophila melanogaster and it is involved in sister chromatid cohesion during mitosis. Our data reveal that haspin also phosphorylates H3T3 in interphase. H3T3ph localizes in broad silenced domains at heterochromatin and lamin-enriched euchromatic regions. Loss of haspin compromises insulator activity in enhancer-blocking assays and triggers a decrease in nuclear size that is accompanied by changes in nuclear envelope morphology. We show that haspin is a suppressor of position-effect variegation involved in heterochromatin organization. Our results also demonstrate that haspin is necessary for pairing-sensitive silencing and it is required for robust Polycomb-dependent homeotic gene silencing. Haspin associates with the cohesin complex in interphase, mediates Pds5 binding to chromatin and cooperates with Pds5-cohesin to modify Polycomb-dependent homeotic transformations. Therefore, this study uncovers an unanticipated role for haspin kinase in genome organization of interphase cells and demonstrates that haspin is required for homeotic gene regulation.


Assuntos
Cromatina/genética , Proteínas de Drosophila/genética , Mitose/genética , Proteínas Serina-Treonina Quinases/genética , Animais , Proteínas de Ciclo Celular/genética , Centrômero/genética , Proteínas Cromossômicas não Histona/genética , Segregação de Cromossomos/genética , Drosophila melanogaster/genética , Inativação Gênica , Heterocromatina/genética , Histonas/genética , Interfase/genética , Fosforilação , Proteínas do Grupo Polycomb/genética , Troca de Cromátide Irmã/genética , Treonina/genética
13.
Nat Commun ; 11(1): 3521, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32665556

RESUMO

Microtubules (MTs) mediate mitosis, directional signaling, and are therapeutic targets in cancer. Yet in vivo analysis of cancer cell MT behavior within the tumor microenvironment remains challenging. Here we developed an imaging pipeline using plus-end tip tracking and intravital microscopy to quantify MT dynamics in live xenograft tumor models. Among analyzed features, cancer cells in vivo displayed higher coherent orientation of MT dynamics along their cell major axes compared with 2D in vitro cultures, and distinct from 3D collagen gel cultures. This in vivo MT phenotype was reproduced in vitro when cells were co-cultured with IL4-polarized MΦ. MΦ depletion, MT disruption, targeted kinase inhibition, and altered MΦ polarization via IL10R blockade all reduced MT coherence and/or tumor cell elongation. We show that MT coherence is a defining feature for in vivo tumor cell dynamics and migration, modulated by local signaling from pro-tumor macrophages.


Assuntos
Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/metabolismo , Animais , Ciclo Celular/genética , Ciclo Celular/fisiologia , Movimento Celular/genética , Movimento Celular/fisiologia , Feminino , Humanos , Macrófagos/citologia , Macrófagos/metabolismo , Camundongos , Proteínas Associadas aos Microtúbulos/genética , Mitose/genética , Mitose/fisiologia , Análise de Componente Principal , Células RAW 264.7
14.
Nat Commun ; 11(1): 3317, 2020 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-32620775

RESUMO

Oriented cell division is a fundamental mechanism to control asymmetric stem cell division, neural tube elongation and body axis extension, among other processes. During zebrafish gastrulation, when the body axis extends, dorsal epiblast cells display divisions that are robustly oriented along the animal-vegetal embryonic axis. Here, we use a combination of lipidomics, metabolic tracer analysis and quantitative image analysis to show that sphingolipids mediate spindle positioning during oriented division of epiblast cells. We identify the Wnt signaling as a regulator of sphingolipid synthesis that mediates the activity of serine palmitoyltransferase (SPT), the first and rate-limiting enzyme in sphingolipid production. Sphingolipids determine the palmitoylation state of the Anthrax receptor, which then positions the mitotic spindle of dividing epiblast cells. Our data show how Wnt signaling mediates sphingolipid-dependent oriented division and how sphingolipids determine Anthrax receptor palmitoylation, which ultimately controls the activation of Diaphanous to mediate spindle rotation and oriented mitosis.


Assuntos
Embrião não Mamífero/metabolismo , Mitose , Receptores de Peptídeos/metabolismo , Esfingolipídeos/metabolismo , Via de Sinalização Wnt , Sequência de Aminoácidos , Animais , Divisão Celular Assimétrica/genética , Embrião não Mamífero/citologia , Embrião não Mamífero/embriologia , Gastrulação , Regulação da Expressão Gênica no Desenvolvimento , Camadas Germinativas/citologia , Camadas Germinativas/embriologia , Camadas Germinativas/metabolismo , Lipoilação , Tubo Neural/citologia , Tubo Neural/embriologia , Tubo Neural/metabolismo , Receptores de Peptídeos/genética , Homologia de Sequência de Aminoácidos , Serina C-Palmitoiltransferase/genética , Serina C-Palmitoiltransferase/metabolismo , Fuso Acromático/metabolismo , Peixe-Zebra , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
15.
Proc Natl Acad Sci U S A ; 117(30): 18091-18098, 2020 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-32661148

RESUMO

The frequent mutations recovered recently from the pollen of select maize lines resulted from the meiotic mobilization of specific low-copy number long-terminal repeat (LTR) retrotransposons, which differ among lines. Mutations that arise at male meiosis produce kernels with concordant mutant phenotypes in both endosperm and embryo because the two sperms that participate in double fertilization are genetically identical. Those are in a majority. However, a small minority of kernels with a mutant endosperm carry a nonconcordant normal embryo, pointing to a postmeiotic or microgametophytic origin. In this study, we have identified the basis for those nonconcordant mutations. We find that all are produced by transposition of a defective LTR retrotransposon that we have termed dRemp (defective retroelement mobile in pollen). This element has several unique properties. Unlike the mutagenic LTR retrotransposons identified previously, dRemp is present in hundreds of copies in all sequenced lines. It seems to transpose only at the second pollen mitosis because all dRemp insertion mutants are nonconcordant yet recoverable in either the endosperm or the embryo. Although it does not move in most lines, dRemp is highly mobile in the Corn Belt inbred M14, identified earlier by breeders as being highly unstable. Lastly, it can be recovered in an array of structures, ranging from solo LTRs to tandem dRemp repeats containing several internal LTRs, suggestive of extensive recombination during retrotransposition. These results shed further light on the spontaneous mutation process and on the possible basis for inbred instability in maize.


Assuntos
Mitose , Mutação , Pólen , Retroelementos , Zea mays/fisiologia , Evolução Molecular , Mutação em Linhagem Germinativa , Endogamia , Fenótipo , Melhoramento Vegetal
16.
Gene ; 760: 144989, 2020 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-32717307

RESUMO

Kinesin 14 family member KIFC1 is a mitotic kinesin which contains a C-terminal motor domain and plays a vital role for clustering the amplified centrosomes. Overexpression of KIFC1 in prostate cancer (PCa) cells showed resistance to docetaxel (DTX). The present study revealed that small KIFC1 inhibitor AZ82 suppresed the transcription and translation of KIFC1 significantly in PCa cells. AZ82 inhibited the KIFC1 expression both in the cytoplasm and nucleus of PCa cells. Inhibition of KIFC1 by AZ82 caused multipolar mitosis in PCa cells via de-clustering the amplified centrosomes and decreased the rate of cancer cell growth and proliferation. Moreover, depletion of KIFC1 reduced cells entering the cell cycle and caused PCa cells death through apoptosis by increasing the expression of Bax and Cytochrome C. Thereby, KIFC1 silencing and inhibition decreased the PCa cells survival by inducing multipolar mitosis as well as apoptosis, suggesting inhibition of KIFC1 using AZ82 might be a strategy to treat PCa by controlling the cancer cell proliferation.


Assuntos
Alanina/análogos & derivados , Centrossomo/efeitos dos fármacos , Cinesina/antagonistas & inibidores , Neoplasias da Próstata/tratamento farmacológico , Piridinas/farmacologia , Alanina/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Centrossomo/metabolismo , Dineínas/metabolismo , Humanos , Cinesina/genética , Cinesina/metabolismo , Masculino , Mitose/efeitos dos fármacos , Miosinas/metabolismo , Próstata/patologia , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia
17.
Proc Natl Acad Sci U S A ; 117(28): 16527-16536, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32601218

RESUMO

Folate deprivation drives the instability of a group of rare fragile sites (RFSs) characterized by CGG trinucleotide repeat (TNR) sequences. Pathological expansion of the TNR within the FRAXA locus perturbs DNA replication and is the major causative factor for fragile X syndrome, a sex-linked disorder associated with cognitive impairment. Although folate-sensitive RFSs share many features with common fragile sites (CFSs; which are found in all individuals), they are induced by different stresses and share no sequence similarity. It is known that a pathway (termed MiDAS) is employed to complete the replication of CFSs in early mitosis. This process requires RAD52 and is implicated in generating translocations and copy number changes at CFSs in cancers. However, it is unclear whether RFSs also utilize MiDAS and to what extent the fragility of CFSs and RFSs arises by shared or distinct mechanisms. Here, we demonstrate that MiDAS does occur at FRAXA following folate deprivation but proceeds via a pathway that shows some mechanistic differences from that at CFSs, being dependent on RAD51, SLX1, and POLD3. A failure to complete MiDAS at FRAXA leads to severe locus instability and missegregation in mitosis. We propose that break-induced DNA replication is required for the replication of FRAXA under folate stress and define a cellular function for human SLX1. These findings provide insights into how folate deprivation drives instability in the human genome.


Assuntos
Endodesoxirribonucleases/metabolismo , Ácido Fólico/metabolismo , Síndrome do Cromossomo X Frágil/metabolismo , Mitose , Rad51 Recombinase/metabolismo , DNA/genética , DNA/metabolismo , Reparo do DNA , Endodesoxirribonucleases/genética , Síndrome do Cromossomo X Frágil/genética , Síndrome do Cromossomo X Frágil/fisiopatologia , Humanos , Rad51 Recombinase/genética , Proteína Rad52 de Recombinação e Reparo de DNA/genética , Proteína Rad52 de Recombinação e Reparo de DNA/metabolismo , Recombinases/genética , Recombinases/metabolismo
18.
Proc Natl Acad Sci U S A ; 117(28): 16154-16159, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32601228

RESUMO

The metaphase spindle is a dynamic structure orchestrating chromosome segregation during cell division. Recently, soft matter approaches have shown that the spindle behaves as an active liquid crystal. Still, it remains unclear how active force generation contributes to its characteristic spindle-like shape. Here we combine theory and experiments to show that molecular motor-driven forces shape the structure through a barreling-type instability. We test our physical model by titrating dynein activity in Xenopus egg extract spindles and quantifying the shape and microtubule orientation. We conclude that spindles are shaped by the interplay between surface tension, nematic elasticity, and motor-driven active forces. Our study reveals how motor proteins can mold liquid crystalline droplets and has implications for the design of active soft materials.


Assuntos
Metáfase/fisiologia , Fuso Acromático/fisiologia , Animais , Fenômenos Biomecânicos , Dineínas/antagonistas & inibidores , Dineínas/metabolismo , Elasticidade , Cristais Líquidos , Metáfase/efeitos dos fármacos , Microtúbulos/efeitos dos fármacos , Microtúbulos/fisiologia , Mitose , Fuso Acromático/química , Fuso Acromático/efeitos dos fármacos , Tensão Superficial , Proteínas de Xenopus/antagonistas & inibidores , Proteínas de Xenopus/metabolismo , Xenopus laevis
19.
Science ; 369(6505): 787-793, 2020 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-32675289

RESUMO

Although Huntington's disease is a late-manifesting neurodegenerative disorder, both mouse studies and neuroimaging studies of presymptomatic mutation carriers suggest that Huntington's disease might affect neurodevelopment. To determine whether this is actually the case, we examined tissue from human fetuses (13 weeks gestation) that carried the Huntington's disease mutation. These tissues showed clear abnormalities in the developing cortex, including mislocalization of mutant huntingtin and junctional complex proteins, defects in neuroprogenitor cell polarity and differentiation, abnormal ciliogenesis, and changes in mitosis and cell cycle progression. We observed the same phenomena in Huntington's disease mouse embryos, where we linked these abnormalities to defects in interkinetic nuclear migration of progenitor cells. Huntington's disease thus has a neurodevelopmental component and is not solely a degenerative disease.


Assuntos
Proteína Huntingtina/metabolismo , Doença de Huntington/metabolismo , Sistema Nervoso/embriologia , Animais , Ciclo Celular , Endossomos/metabolismo , Feto , Humanos , Proteína Huntingtina/genética , Doença de Huntington/genética , Camundongos , Camundongos Mutantes , Mitose , Mutação , Células Neuroepiteliais/metabolismo , Junções Íntimas/metabolismo , Proteína da Zônula de Oclusão-1/metabolismo
20.
Cancer Sci ; 111(9): 3155-3163, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32594560

RESUMO

The eukaryotic nucleus is not a homogenous single-spaced but a highly compartmentalized organelle, partitioned by various types of membraneless structures, including nucleoli, PML bodies, paraspeckles, DNA damage foci and RNA clouds. Over the past few decades, these nuclear structures have been implicated in biological reactions such as gene regulation and DNA damage response and repair, and are thought to provide "microenvironments," facilitating these reactions in the nucleus. Notably, an altered morphology of these nuclear structures is found in many cancers, which may relate to so-called "nuclear atypia" in histological examinations. While the diagnostic significance of nuclear atypia has been established, its nature has remained largely enigmatic and awaits characterization. Here, we review the emerging biophysical principles that govern biomolecular condensate assembly in the nucleus, namely, liquid-liquid phase separation (LLPS), to investigate the nature of the nuclear microenvironment. In the nucleus, LLPS is typically driven by multivalent interactions between proteins with intrinsically disordered regions, and is also facilitated by protein interaction with nucleic acids, including nuclear non-coding RNAs. Importantly, an altered LLPS leads to dysregulation of nuclear events and epigenetics, and often to tumorigenesis and tumor progression. We further note the possibility that LLPS could represent a new therapeutic target for cancer intervention.


Assuntos
Núcleo Celular/metabolismo , Suscetibilidade a Doenças , Neoplasias/etiologia , Neoplasias/metabolismo , Biomarcadores , Núcleo Celular/genética , Cromatina/genética , Cromatina/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Extração Líquido-Líquido , Mitose , Neoplasias/patologia , Proteômica/métodos , RNA não Traduzido
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