Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 35
Filtrar
1.
Cell ; 185(23): 4376-4393.e18, 2022 11 10.
Artigo em Inglês | MEDLINE | ID: mdl-36318920

RESUMO

The function of biomolecular condensates is often restricted by condensate dissolution. Whether condensates can be suppressed without condensate dissolution is unclear. Here, we show that upstream regulators of the Hippo signaling pathway form functionally antagonizing condensates, and their coalescence into a common phase provides a mode of counteracting the function of biomolecular condensates without condensate dissolution. Specifically, the negative regulator SLMAP forms Hippo-inactivating condensates to facilitate pathway inhibition by the STRIPAK complex. In response to cell-cell contact or osmotic stress, the positive regulators AMOT and KIBRA form Hippo-activating condensates to facilitate pathway activation. The functionally antagonizing SLMAP and AMOT/KIBRA condensates further coalesce into a common phase to inhibit STRIPAK function. These findings provide a paradigm for restricting the activity of biomolecular condensates without condensate dissolution, shed light on the molecular principles of multiphase organization, and offer a conceptual framework for understanding upstream regulation of the Hippo signaling pathway.


Assuntos
Via de Sinalização Hippo , Proteínas Serina-Treonina Quinases , Transdução de Sinais
2.
Nat Rev Mol Cell Biol ; 24(1): 5, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36008600
3.
Annu Rev Genet ; 53: 67-91, 2019 12 03.
Artigo em Inglês | MEDLINE | ID: mdl-31283358

RESUMO

Cell-cell fusion is indispensable for creating life and building syncytial tissues and organs. Ever since the discovery of cell-cell fusion, how cells join together to form zygotes and multinucleated syncytia has remained a fundamental question in cell and developmental biology. In the past two decades, Drosophila myoblast fusion has been used as a powerful genetic model to unravel mechanisms underlying cell-cell fusion in vivo. Many evolutionarily conserved fusion-promoting factors have been identified and so has a surprising and conserved cellular mechanism. In this review, we revisit key findings in Drosophila myoblast fusion and highlight the critical roles of cellular invasion and resistance in driving cell membrane fusion.


Assuntos
Proteínas de Drosophila/metabolismo , Drosophila/citologia , Mioblastos/citologia , Actinas/metabolismo , Actomiosina/metabolismo , Animais , Moléculas de Adesão Celular/metabolismo , Fusão Celular , Drosophila/embriologia , Drosophila/fisiologia , Proteínas de Drosophila/genética , Embrião não Mamífero/citologia , Bicamadas Lipídicas/metabolismo , Músculos/citologia , Músculos/embriologia , Mioblastos/fisiologia , Pupa/citologia
4.
Nature ; 592(7853): 272-276, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33508854

RESUMO

Cell competition involves a conserved fitness-sensing process during which fitter cells eliminate neighbouring less-fit but viable cells1. Cell competition has been proposed as a surveillance mechanism to ensure normal development and tissue homeostasis, and has also been suggested to act as a barrier to interspecies chimerism2. However, cell competition has not been studied in an interspecies context during early development owing to the lack of an in vitro model. Here we developed an interspecies pluripotent stem cell (PSC) co-culture strategy and uncovered a previously unknown mode of cell competition between species. Interspecies competition between PSCs occurred in primed but not naive pluripotent cells, and between evolutionarily distant species. By comparative transcriptome analysis, we found that genes related to the NF-κB signalling pathway, among others, were upregulated in less-fit 'loser' human cells. Genetic inactivation of a core component (P65, also known as RELA) and an upstream regulator (MYD88) of the NF-κB complex in human cells could overcome the competition between human and mouse PSCs, thereby improving the survival and chimerism of human cells in early mouse embryos. These insights into cell competition pave the way for the study of evolutionarily conserved mechanisms that underlie competitive cell interactions during early mammalian development. Suppression of interspecies PSC competition may facilitate the generation of human tissues in animals.


Assuntos
Competição entre as Células/fisiologia , Quimerismo , Técnicas de Cocultura/métodos , Embrião de Mamíferos/citologia , Células-Tronco Pluripotentes/citologia , Animais , Contagem de Células , Sobrevivência Celular , Feminino , Humanos , Masculino , Camundongos , Fator 88 de Diferenciação Mieloide/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais , Especificidade da Espécie , Fator de Transcrição RelA/metabolismo
5.
J Cell Sci ; 132(18)2019 09 16.
Artigo em Inglês | MEDLINE | ID: mdl-31527149

RESUMO

Cell-cell fusion is a fundamental process underlying fertilization, development, regeneration and physiology of metazoans. It is a multi-step process involving cell recognition and adhesion, actin cytoskeletal rearrangements, fusogen engagement, lipid mixing and fusion pore formation, ultimately resulting in the integration of two fusion partners. Here, we focus on the asymmetric actin cytoskeletal rearrangements at the site of fusion, known as the fusogenic synapse, which was first discovered during myoblast fusion in Drosophila embryos and later also found in mammalian muscle and non-muscle cells. At the asymmetric fusogenic synapse, actin-propelled invasive membrane protrusions from an attacking fusion partner trigger actomyosin-based mechanosensory responses in the receiving cell. The interplay between the invasive and resisting forces generated by the two fusion partners puts the fusogenic synapse under high mechanical tension and brings the two cell membranes into close proximity, promoting the engagement of fusogens to initiate fusion pore formation. In this Cell Science at a Glance article and the accompanying poster, we highlight the molecular, cellular and biophysical events at the asymmetric fusogenic synapse using Drosophila myoblast fusion as a model.


Assuntos
Citoesqueleto de Actina/metabolismo , Proteínas de Drosophila/metabolismo , Mioblastos/citologia , Mioblastos/metabolismo , Citoesqueleto de Actina/genética , Animais , Fusão Celular , Drosophila , Proteínas de Drosophila/genética , Embrião não Mamífero/citologia , Embrião não Mamífero/metabolismo , Mecanotransdução Celular/genética , Mecanotransdução Celular/fisiologia
6.
Proc Natl Acad Sci U S A ; 114(45): 11950-11955, 2017 11 07.
Artigo em Inglês | MEDLINE | ID: mdl-29078404

RESUMO

Skeletal muscle formation requires fusion of mononucleated myoblasts to form multinucleated myofibers. The muscle-specific membrane proteins myomaker and myomixer cooperate to drive mammalian myoblast fusion. Whereas myomaker is highly conserved across diverse vertebrate species, myomixer is a micropeptide that shows relatively weak cross-species conservation. To explore the functional conservation of myomixer, we investigated the expression and function of the zebrafish myomixer ortholog. Here we show that myomixer expression during zebrafish embryogenesis coincides with myoblast fusion, and genetic deletion of myomixer using CRISPR/Cas9 mutagenesis abolishes myoblast fusion in vivo. We also identify myomixer orthologs in other species of fish and reptiles, which can cooperate with myomaker and substitute for the fusogenic activity of mammalian myomixer. Sequence comparison of these diverse myomixer orthologs reveals key amino acid residues and a minimal fusogenic peptide motif that is necessary for promoting cell-cell fusion with myomaker. Our findings highlight the evolutionary conservation of the myomaker-myomixer partnership and provide insights into the molecular basis of myoblast fusion.


Assuntos
Proteínas de Membrana/genética , Desenvolvimento Muscular/genética , Fibras Musculares Esqueléticas/citologia , Proteínas Musculares/genética , Mioblastos/metabolismo , Proteínas de Peixe-Zebra/genética , Sequência de Aminoácidos , Animais , Sistemas CRISPR-Cas/genética , Fusão Celular , Linhagem Celular , Elefantes/genética , Desenvolvimento Muscular/fisiologia , Tubarões/genética , Tartarugas/genética , Peixe-Zebra/embriologia , Peixe-Zebra/genética
7.
Curr Biol ; 34(7): 1438-1452.e6, 2024 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-38513654

RESUMO

Steroid hormones regulate tissue development and physiology by modulating the transcription of a broad spectrum of genes. In insects, the principal steroid hormones, ecdysteroids, trigger the expression of thousands of genes through a cascade of transcription factors (TFs) to coordinate developmental transitions such as larval molting and metamorphosis. However, whether ecdysteroid signaling can bypass transcriptional hierarchies to exert its function in individual developmental processes is unclear. Here, we report that a single non-TF effector gene mediates the transcriptional output of ecdysteroid signaling in Drosophila myoblast fusion, a critical step in muscle development and differentiation. Specifically, we show that the 20-hydroxyecdysone (commonly referred to as "ecdysone") secreted from an extraembryonic tissue, amnioserosa, acts on embryonic muscle cells to directly activate the expression of antisocial (ants), which encodes an essential scaffold protein enriched at the fusogenic synapse. Not only is ants transcription directly regulated by the heterodimeric ecdysone receptor complex composed of ecdysone receptor (EcR) and ultraspiracle (USP) via ecdysone-response elements but also more strikingly, expression of ants alone is sufficient to rescue the myoblast fusion defect in ecdysone signaling-deficient mutants. We further show that EcR/USP and a muscle-specific TF Twist synergistically activate ants expression in vitro and in vivo. Taken together, our study provides the first example of a steroid hormone directly activating the expression of a single key non-TF effector gene to regulate a developmental process via inter-organ signaling and provides a new paradigm for understanding steroid hormone signaling in other developmental and physiological processes.


Assuntos
Proteínas de Drosophila , Receptores de Esteroides , Animais , Proteínas de Ligação a DNA/metabolismo , Ecdisona , Ecdisteroides , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Muda/fisiologia , Drosophila/fisiologia , Regulação da Expressão Gênica no Desenvolvimento
8.
bioRxiv ; 2024 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-39416162

RESUMO

Skeletal muscle regeneration is a multistep process involving the activation, proliferation, differentiation, and fusion of muscle stem cells, known as satellite cells. The fusion of satellite cell-derived mononucleated muscle cells (SCMs) is indispensable for the generation of multinucleated, contractile myofibers during muscle repair. However, the molecular and cellular mechanisms underlying SCM fusion during muscle regeneration remain poorly understood. In this study, we uncovered an essential role for branched actin polymerization in SCM fusion. Using conditional knockouts of the Arp2/3 complex and its actin nucleation-promoting factors, N-WASP and WAVE, we demonstrated that branched actin polymerization is required for the SCM fusion, but not for satellite cell proliferation, differentiation, and migration. We showed that the N-WASP and WAVE complexes have partially redundant functions in regulating SCM fusion. Furthermore, we showed that branched actin polymerization is essential for generating invasive protrusions at the fusogenic synapses in SCMs. Taken together, our study has identified new components of the myoblast fusion machinery in skeletal muscle regeneration and demonstrated a critical role for branched actin-propelled invasive protrusions in this process.

9.
Front Cell Dev Biol ; 12: 1358583, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38827528

RESUMO

Breast cancer metastases exhibit many different genetic alterations, including copy number amplifications (CNA). CNA are genetic alterations that are increasingly becoming relevant to breast oncology clinical practice. Here we identify CNA in metastatic breast tumor samples using publicly available datasets and characterize their expression and function using a metastatic mouse model of breast cancer. Our findings demonstrate that our organoid generation can be implemented to study clinically relevant features that reflect the genetic heterogeneity of individual tumors.

10.
Nat Commun ; 15(1): 2755, 2024 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-38553438

RESUMO

Projection imaging accelerates volumetric interrogation in fluorescence microscopy, but for multi-cellular samples, the resulting images may lack contrast, as many structures and haze are summed up. Here, we demonstrate rapid projective light-sheet imaging with parameter selection (props) of imaging depth, position and viewing angle. This allows us to selectively image different sub-volumes of a sample, rapidly switch between them and exclude background fluorescence. Here we demonstrate the power of props by functional imaging within distinct regions of the zebrafish brain, monitoring calcium firing inside muscle cells of moving Drosophila larvae, super-resolution imaging of selected cell layers, and by optically unwrapping the curved surface of a Drosophila embryo. We anticipate that props will accelerate volumetric interrogation, ranging from subcellular to mesoscopic scales.


Assuntos
Drosophila , Peixe-Zebra , Animais , Microscopia de Fluorescência/métodos , Encéfalo/ultraestrutura , Larva
11.
bioRxiv ; 2023 Jan 31.
Artigo em Inglês | MEDLINE | ID: mdl-36778256

RESUMO

Breast cancer metastases exhibit many different genetic alterations, including copy number amplifications. Using publicly available datasets, we identify copy number amplifications in metastatic breast tumor samples and using our organoid-based metastasis assays, and we validate FGFR1 is amplified in collectively migrating organoids. Because the heterogeneity of breast tumors is increasingly becoming relevant to clinical practice, we demonstrate our organoid method captures genetic heterogeneity of individual tumors.

12.
bioRxiv ; 2023 Mar 09.
Artigo em Inglês | MEDLINE | ID: mdl-36945615

RESUMO

Interspecies chimera formation with human pluripotent stem cells (PSCs) holds great promise to generate humanized animal models and provide donor organs for transplant. However, the approach is currently limited by low levels of human cells ultimately represented in chimeric embryos. Different strategies have been developed to improve chimerism by genetically editing donor human PSCs. To date, however, it remains unexplored if human chimerism can be enhanced in animals through modifying the host embryos. Leveraging the interspecies PSC competition model, here we discovered retinoic acid-inducible gene I (RIG-I)-like receptor (RLR) signaling, an RNA sensor, in "winner" cells plays an important role in the competitive interactions between co-cultured mouse and human PSCs. We found that genetic inactivation of Ddx58/Ifih1-Mavs-Irf7 axis compromised the "winner" status of mouse PSCs and their ability to outcompete PSCs from evolutionarily distant species during co-culture. Furthermore, by using Mavs-deficient mouse embryos we substantially improved unmodified donor human cell survival. Comparative transcriptome analyses based on species-specific sequences suggest contact-dependent human-to-mouse transfer of RNAs likely plays a part in mediating the cross-species interactions. Taken together, these findings establish a previously unrecognized role of RNA sensing and innate immunity in "winner" cells during cell competition and provides a proof-of-concept for modifying host embryos, rather than donor PSCs, to enhance interspecies chimerism.

13.
Dev Cell ; 12(4): 571-86, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17419995

RESUMO

Myoblast fusion is an essential step during muscle differentiation. Previous studies in Drosophila have revealed a signaling pathway that relays the fusion signal from the plasma membrane to the actin cytoskeleton. However, the function for the actin cytoskeleton in myoblast fusion remains unclear. Here we describe the characterization of solitary (sltr), a component of the myoblast fusion signaling cascade. sltr encodes the Drosophila ortholog of the mammalian WASP-interacting protein. Sltr is recruited to sites of fusion by the fusion-competent cell-specific receptor Sns and acts as a positive regulator for actin polymerization at these sites. Electron microscopy analysis suggests that formation of F-actin-enriched foci at sites of fusion is involved in the proper targeting and coating of prefusion vesicles. These studies reveal a surprising cell-type specificity of Sltr-mediated actin polymerization in myoblast fusion, and demonstrate that targeted exocytosis of prefusion vesicles is a critical step prior to plasma membrane fusion.


Assuntos
Actinas/metabolismo , Drosophila melanogaster/metabolismo , Exocitose , Desenvolvimento Muscular , Mioblastos/fisiologia , Actinas/fisiologia , Sequência de Aminoácidos , Animais , Animais Geneticamente Modificados , Fusão Celular , Linhagem Celular , Membrana Celular/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/embriologia , Drosophila melanogaster/genética , Drosophila melanogaster/fisiologia , Embrião não Mamífero , Camundongos , Modelos Biológicos , Dados de Sequência Molecular , Mioblastos/metabolismo , Proteínas Proto-Oncogênicas c-crk/metabolismo , Homologia de Sequência de Aminoácidos , Transfecção , Vesículas Transportadoras/metabolismo , Proteína da Síndrome de Wiskott-Aldrich/genética , Proteína da Síndrome de Wiskott-Aldrich/metabolismo
14.
Nat Genet ; 54(8): 1202-1213, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35835915

RESUMO

Epigenetic remodeling is essential for oncogene-induced cellular transformation and malignancy. In contrast to histone post-translational modifications, how DNA methylation is remodeled by oncogenic signaling remains poorly understood. The oncoprotein YAP, a coactivator of the TEAD transcription factors mediating Hippo signaling, is widely activated in human cancers. Here, we identify the 5-methylcytosine dioxygenase TET1 as a direct YAP target and a master regulator that coordinates the genome-wide epigenetic and transcriptional reprogramming of YAP target genes in the liver. YAP activation induces the expression of TET1, which physically interacts with TEAD to cause regional DNA demethylation, histone H3K27 acetylation and chromatin opening in YAP target genes to facilitate transcriptional activation. Loss of TET1 not only reverses YAP-induced epigenetic and transcriptional changes but also suppresses YAP-induced hepatomegaly and tumorigenesis. These findings exemplify how oncogenic signaling regulates the site specificity of DNA demethylation to promote tumorigenesis and implicate TET1 as a potential target for modulating YAP signaling in physiology and disease.


Assuntos
Proteínas de Ligação a DNA , Fatores de Transcrição , Carcinogênese/genética , Carcinogênese/metabolismo , Proteínas de Ciclo Celular , Proteínas de Ligação a DNA/metabolismo , Epigênese Genética , Histonas/genética , Humanos , Fígado/metabolismo , Oxigenases de Função Mista/genética , Oxigenases de Função Mista/metabolismo , Oncogenes , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
15.
Dev Cell ; 57(13): 1582-1597.e6, 2022 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-35709765

RESUMO

Myoblast fusion is an indispensable process in skeletal muscle development and regeneration. Studies in Drosophila led to the discovery of the asymmetric fusogenic synapse, in which one cell invades its fusion partner with actin-propelled membrane protrusions to promote fusion. However, the timing and sites of vertebrate myoblast fusion remain elusive. Here, we show that fusion between zebrafish fast muscle cells is mediated by an F-actin-enriched invasive structure. Two cell adhesion molecules, Jam2a and Jam3b, are associated with the actin structure, with Jam2a being the major organizer. The Arp2/3 actin nucleation-promoting factors, WAVE and WASP-but not the bipartite fusogenic proteins, Myomaker or Myomixer-promote the formation of the invasive structure. Moreover, the convergence of fusogen-containing microdomains and the invasive protrusions is a prerequisite for cell membrane fusion. Thus, our study provides unprecedented insights into the cellular architecture and molecular determinants of the asymmetric fusogenic synapse in an intact vertebrate animal.


Assuntos
Actinas , Peixe-Zebra , Actinas/metabolismo , Animais , Fusão Celular , Drosophila/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Desenvolvimento Muscular , Proteínas Musculares , Sinapses/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/metabolismo
16.
J Vis Exp ; (189)2022 11 11.
Artigo em Inglês | MEDLINE | ID: mdl-36440890

RESUMO

Organoids are a reliable method for modeling organ tissue due to their self-organizing properties and retention of function and architecture after propagation from primary tissue or stem cells. This method of organoid generation forgoes single-cell differentiation through multiple passages and instead uses differential centrifugation to isolate mammary epithelial organoids from mechanically and enzymatically dissociated tissues. This protocol provides a streamlined technique for rapidly producing small and large epithelial organoids from both mouse and human mammary tissue in addition to techniques for organoid embedding in collagen and basement extracellular matrix. Furthermore, instructions for in-gel fixation and immunofluorescent staining are provided for the purpose of visualizing organoid morphology and density. These methodologies are suitable for myriad downstream analyses, such as co-culturing with immune cells and ex vivo metastasis modeling via collagen invasion assay. These analyses serve to better elucidate cell-cell behavior and create a more complete understanding of interactions within the tumor microenvironment.


Assuntos
Neoplasias , Organoides , Humanos , Camundongos , Animais , Diagnóstico por Imagem , Mama , Colágeno , Microambiente Tumoral
17.
Nat Cell Biol ; 22(6): 674-688, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32451441

RESUMO

The dynamin GTPase is known to bundle actin filaments, but the underlying molecular mechanism and physiological relevance remain unclear. Our genetic analyses revealed a function of dynamin in propelling invasive membrane protrusions during myoblast fusion in vivo. Using biochemistry, total internal reflection fluorescence microscopy, electron microscopy and cryo-electron tomography, we show that dynamin bundles actin while forming a helical structure. At its full capacity, each dynamin helix captures 12-16 actin filaments on the outer rim of the helix. GTP hydrolysis by dynamin triggers disassembly of fully assembled dynamin helices, releasing free dynamin dimers/tetramers and facilitating Arp2/3-mediated branched actin polymerization. The assembly/disassembly cycles of dynamin promote continuous actin bundling to generate mechanically stiff actin super-bundles. Super-resolution and immunogold platinum replica electron microscopy revealed dynamin along actin bundles at the fusogenic synapse. These findings implicate dynamin as a unique multifilament actin-bundling protein that regulates the dynamics and mechanical strength of the actin cytoskeletal network.


Assuntos
Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Comunicação Celular , Drosophila melanogaster/metabolismo , Dinaminas/metabolismo , Endocitose , Complexo 2-3 de Proteínas Relacionadas à Actina/metabolismo , Actinas/genética , Sequência de Aminoácidos , Animais , Drosophila melanogaster/genética , Dinaminas/genética , Feminino , Guanosina Trifosfato/metabolismo , Masculino , Mioblastos/citologia , Mioblastos/metabolismo , Ligação Proteica , Homologia de Sequência
18.
Trends Cell Biol ; 14(8): 452-60, 2004 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-15308212

RESUMO

Intercellular fusion among myoblasts is required for the generation of multinucleated muscle fibers during skeletal muscle development. Recent studies in Drosophila have shed light on the molecular mechanisms that underlie this process, and a signaling pathway that relays fusion signals from the cell membrane to the cytoskeleton has emerged. In this article, we review these recent advances and discuss how Drosophila offers a powerful model system to study myoblast fusion in vivo.


Assuntos
Proteínas de Drosophila/metabolismo , Drosophila/metabolismo , Mioblastos/fisiologia , Transdução de Sinais/fisiologia , Animais , Fusão Celular , Drosophila/genética , Proteínas de Drosophila/genética
19.
Genetics ; 212(2): 365-376, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31167898

RESUMO

Bruce Baker, a preeminent Drosophila geneticist who made fundamental contributions to our understanding of the molecular genetic basis of sex differences, passed away July 1, 2018 at the age of 72. Members of Bruce's laboratory remember him as an intensely dedicated, rigorous, creative, deep-thinking, and fearless scientist. His trainees also remember his strong commitment to teaching students at every level. Bruce's career studying sex differences had three major epochs, where the laboratory was focused on: (1) sex determination and dosage compensation, (2) the development of sex-specific structures, and (3) the molecular genetic basis for sex differences in behavior. Several members of the Baker laboratory have come together to honor Bruce by highlighting some of the laboratory's major scientific contributions in these areas.


Assuntos
Drosophila/genética , Genética/história , Processos de Determinação Sexual/genética , Animais , Mecanismo Genético de Compensação de Dose , Evolução Molecular , História do Século XX , História do Século XXI , Mentores , Comportamento Sexual
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA