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1.
Int J Mol Sci ; 22(17)2021 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-34502050

RESUMO

To ensure the formation of a properly patterned embryo, multiple processes must operate harmoniously at sequential phases of development. This is implemented by mutual interactions between cells and tissues that together regulate the segregation and specification of cells, their growth and morphogenesis. The formation of the spinal cord and paraxial mesoderm derivatives exquisitely illustrate these processes. Following early gastrulation, while the vertebrate body elongates, a population of bipotent neuromesodermal progenitors resident in the posterior region of the embryo generate both neural and mesodermal lineages. At later stages, the somitic mesoderm regulates aspects of neural patterning and differentiation of both central and peripheral neural progenitors. Reciprocally, neural precursors influence the paraxial mesoderm to regulate somite-derived myogenesis and additional processes by distinct mechanisms. Central to this crosstalk is the activity of the axial notochord, which, via sonic hedgehog signaling, plays pivotal roles in neural, skeletal muscle and cartilage ontogeny. Here, we discuss the cellular and molecular basis underlying this complex developmental plan, with a focus on the logic of sonic hedgehog activities in the coordination of the neural-mesodermal axis.


Assuntos
Diferenciação Celular , Mesoderma/citologia , Tubo Neural/citologia , Animais , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Células-Tronco Embrionárias/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Humanos , Mesoderma/embriologia , Mesoderma/metabolismo , Tubo Neural/embriologia , Tubo Neural/metabolismo
2.
Nat Commun ; 12(1): 4797, 2021 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34376651

RESUMO

Sutures separate the flat bones of the skull and enable coordinated growth of the brain and overlying cranium. The coronal suture is most commonly fused in monogenic craniosynostosis, yet the unique aspects of its development remain incompletely understood. To uncover the cellular diversity within the murine embryonic coronal suture, we generated single-cell transcriptomes and performed extensive expression validation. We find distinct pre-osteoblast signatures between the bone fronts and periosteum, a ligament-like population above the suture that persists into adulthood, and a chondrogenic-like population in the dura mater underlying the suture. Lineage tracing reveals an embryonic Six2+ osteoprogenitor population that contributes to the postnatal suture mesenchyme, with these progenitors being preferentially affected in a Twist1+/-; Tcf12+/- mouse model of Saethre-Chotzen Syndrome. This single-cell atlas provides a resource for understanding the development of the coronal suture and the mechanisms for its loss in craniosynostosis.


Assuntos
Suturas Cranianas/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Osteogênese/genética , Análise de Célula Única/métodos , Transcriptoma/genética , Acrocefalossindactilia/embriologia , Acrocefalossindactilia/genética , Acrocefalossindactilia/patologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Suturas Cranianas/citologia , Suturas Cranianas/embriologia , Dura-Máter/citologia , Dura-Máter/embriologia , Dura-Máter/metabolismo , Mesoderma/citologia , Mesoderma/embriologia , Mesoderma/metabolismo , Camundongos Knockout , Camundongos Transgênicos , Osteoblastos/citologia , Osteoblastos/metabolismo , RNA-Seq/métodos , Crânio/citologia , Crânio/embriologia , Crânio/metabolismo , Proteína 1 Relacionada a Twist/genética , Proteína 1 Relacionada a Twist/metabolismo
3.
Development ; 148(15)2021 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-34338289

RESUMO

Transcriptional regulatory networks refine gene expression boundaries to define the dimensions of organ progenitor territories. Kidney progenitors originate within the intermediate mesoderm (IM), but the pathways that establish the boundary between the IM and neighboring vessel progenitors are poorly understood. Here, we delineate roles for the zinc-finger transcription factor Osr1 in kidney and vessel progenitor development. Zebrafish osr1 mutants display decreased IM formation and premature emergence of lateral vessel progenitors (LVPs). These phenotypes contrast with the increased IM and absent LVPs observed with loss of the bHLH transcription factor Hand2, and loss of hand2 partially suppresses osr1 mutant phenotypes. hand2 and osr1 are expressed together in the posterior mesoderm, but osr1 expression decreases dramatically prior to LVP emergence. Overexpressing osr1 during this timeframe inhibits LVP development while enhancing IM formation, and can rescue the osr1 mutant phenotype. Together, our data demonstrate that osr1 modulates the extent of IM formation and the temporal dynamics of LVP development, suggesting that a balance between levels of osr1 and hand2 expression is essential to demarcate the kidney and vessel progenitor territories.


Assuntos
Diferenciação Celular/fisiologia , Mesoderma/metabolismo , Mesoderma/fisiologia , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/metabolismo , Peixe-Zebra/fisiologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Rim/metabolismo , Rim/fisiologia , Organogênese/fisiologia , Fatores de Transcrição/metabolismo
4.
Nat Commun ; 12(1): 5126, 2021 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-34446705

RESUMO

Embryonic development is largely conserved among mammals. However, certain genes show divergent functions. By generating a transcriptional atlas containing >30,000 cells from post-implantation non-human primate embryos, we uncover that ISL1, a gene with a well-established role in cardiogenesis, controls a gene regulatory network in primate amnion. CRISPR/Cas9-targeting of ISL1 results in non-human primate embryos which do not yield viable offspring, demonstrating that ISL1 is critically required in primate embryogenesis. On a cellular level, mutant ISL1 embryos display a failure in mesoderm formation due to reduced BMP4 signaling from the amnion. Via loss of function and rescue studies in human embryonic stem cells we confirm a similar role of ISL1 in human in vitro derived amnion. This study highlights the importance of the amnion as a signaling center during primate mesoderm formation and demonstrates the potential of in vitro primate model systems to dissect the genetics of early human embryonic development.


Assuntos
Âmnio/metabolismo , Macaca fascicularis/embriologia , Mesoderma/embriologia , Âmnio/embriologia , Animais , Proteína Morfogenética Óssea 4/metabolismo , Desenvolvimento Embrionário , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Proteínas com Homeodomínio LIM/genética , Proteínas com Homeodomínio LIM/metabolismo , Macaca fascicularis/genética , Macaca fascicularis/metabolismo , Mesoderma/metabolismo , Gravidez , Transdução de Sinais
5.
Am J Physiol Renal Physiol ; 321(3): F378-F388, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34338032

RESUMO

Developmentally heterogeneous renin-expressing cells serve as progenitors for mural, glomerular, and tubular cells during nephrogenesis and are collectively termed renin lineage cells (RLCs). In this study, we quantified different renal vascular and tubular cell types based on specific markers and assessed proliferation and de novo differentiation in the RLC population. We used kidney sections of mRenCre-mT/mG mice throughout nephrogenesis. Marker positivity was evaluated in whole digitalized sections. At embryonic day 16, RLCs appeared in the developing kidney, and the expression of all stained markers in RLCs was observed. The proliferation rate of RLCs did not differ from the proliferation rate of non-RLCs. RLCs expanded mainly by de novo differentiation (neogenesis). Fractions of RLCs originating from the stromal progenitors of the metanephric mesenchyme (renin-producing cells, vascular smooth muscle cells, and mesangial cells) decreased during nephrogenesis. In contrast, aquaporin-2-positive RLCs in the collecting duct system, which embryonically emerges almost exclusively from the ureteric bud, expanded postpartum. The cubilin-positive RLC fraction in the proximal tubule, deriving from the cap mesenchyme, remained constant. In summary, RLCs were continuously detectable in the vascular and tubular compartments of the kidney during nephrogenesis. Therein, various patterns of RLC differentiation that depend on the embryonic origin of the cells were identified.NEW & NOTEWORTHY The unifying feature of the renal renin lineage cells (RLCs) is their origin from renin-expressing progenitors. RLCs evolve to an embryologically heterogeneous large population in structures with different ancestry. RLCs are also targets for the widely used renin-angiotensin-system blockers, which modulate their phenotype. Unveiling the different differentiation patterns of RLCs in the developing kidney contributes to understanding changes in their cell fate in response to homeostatic challenges and the use of antihypertensive drugs.


Assuntos
Diferenciação Celular/fisiologia , Glomérulos Renais/metabolismo , Rim/metabolismo , Células Mesangiais/metabolismo , Renina/metabolismo , Animais , Linhagem da Célula/fisiologia , Mesoderma/metabolismo , Camundongos , Células-Tronco/metabolismo
6.
Nat Commun ; 12(1): 5136, 2021 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-34446717

RESUMO

One fundamental yet unresolved question in biology remains how cells interpret the same signalling cues in a context-dependent manner resulting in lineage specification. A key step for decoding signalling cues is the establishment of a permissive chromatin environment at lineage-specific genes triggering transcriptional responses to inductive signals. For instance, bipotent neuromesodermal progenitors (NMPs) are equipped with a WNT-decoding module, which relies on TCFs/LEF activity to sustain both NMP expansion and paraxial mesoderm differentiation. However, how WNT signalling activates lineage specific genes in a temporal manner remains unclear. Here, we demonstrate that paraxial mesoderm induction relies on the TALE/HOX combinatorial activity that simultaneously represses NMP genes and activates the differentiation program. We identify the BRACHYURY-TALE/HOX code that destabilizes the nucleosomes at WNT-responsive regions and establishes the permissive chromatin landscape for de novo recruitment of the WNT-effector LEF1, unlocking the WNT-mediated transcriptional program that drives NMPs towards the paraxial mesodermal fate.


Assuntos
Proteínas Fetais/metabolismo , Mesoderma/metabolismo , Família Multigênica , Células-Tronco Neurais/metabolismo , Proteínas com Domínio T/metabolismo , Via de Sinalização Wnt , Animais , Diferenciação Celular , Linhagem da Célula , Proteínas Fetais/genética , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Mesoderma/embriologia , Camundongos , Camundongos Knockout , Células-Tronco Neurais/citologia , Nucleossomos/genética , Nucleossomos/metabolismo , Proteínas com Domínio T/genética
7.
Nature ; 597(7876): 410-414, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34408322

RESUMO

Signals from sympathetic neurons and immune cells regulate adipocytes and thereby contribute to fat tissue biology. Interactions between the nervous and immune systems have recently emerged as important regulators of host defence and inflammation1-4. Nevertheless, it is unclear whether neuronal and immune cells co-operate in brain-body axes to orchestrate metabolism and obesity. Here we describe a neuro-mesenchymal unit that controls group 2 innate lymphoid cells (ILC2s), adipose tissue physiology, metabolism and obesity via a brain-adipose circuit. We found that sympathetic nerve terminals act on neighbouring adipose mesenchymal cells via the ß2-adrenergic receptor to control the expression of glial-derived neurotrophic factor (GDNF) and the activity of ILC2s in gonadal fat. Accordingly, ILC2-autonomous manipulation of the GDNF receptor machinery led to alterations in ILC2 function, energy expenditure, insulin resistance and propensity to obesity. Retrograde tracing and chemical, surgical and chemogenetic manipulations identified a sympathetic aorticorenal circuit that modulates ILC2s in gonadal fat and connects to higher-order brain areas, including the paraventricular nucleus of the hypothalamus. Our results identify a neuro-mesenchymal unit that translates cues from long-range neuronal circuitry into adipose-resident ILC2 function, thereby shaping host metabolism and obesity.


Assuntos
Tecido Adiposo/inervação , Tecido Adiposo/metabolismo , Encéfalo/metabolismo , Imunidade Inata/imunologia , Mesoderma/citologia , Vias Neurais , Neurônios/citologia , Obesidade/metabolismo , Tecido Adiposo/citologia , Animais , Encéfalo/citologia , Sinais (Psicologia) , Citocinas/metabolismo , Metabolismo Energético , Feminino , Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Gônadas/metabolismo , Mesoderma/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Núcleo Hipotalâmico Paraventricular/metabolismo , Proteínas Proto-Oncogênicas c-ret/metabolismo , Receptores Adrenérgicos beta 2/metabolismo , Sistema Nervoso Simpático/citologia , Sistema Nervoso Simpático/metabolismo
8.
Development ; 148(14)2021 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-34195802

RESUMO

Tooth formation requires complex signaling interactions both within the oral epithelium and between the epithelium and the underlying mesenchyme. Previous studies of the Wnt/ß-catenin pathway have shown that tooth formation is partly inhibited in loss-of-function mutants, and gain-of-function mutants have perturbed tooth morphology. However, the stage at which Wnt signaling is first important in tooth formation remains unclear. Here, using an Fgf8-promoter-driven, and therefore early, deletion of ß-catenin in mouse molar epithelium, we found that loss of Wnt/ß-catenin signaling completely deletes the molar tooth, demonstrating that this pathway is central to the earliest stages of tooth formation. Early expression of a dominant-active ß-catenin protein also perturbs tooth formation, producing a large domed evagination at early stages and supernumerary teeth later on. The early evaginations are associated with premature mesenchymal condensation marker, and are reduced by inhibition of condensation-associated collagen synthesis. We propose that invagination versus evagination morphogenesis is regulated by the relative timing of epithelial versus mesenchymal cell convergence regulated by canonical Wnt signaling. Together, these studies reveal new aspects of Wnt/ß-catenin signaling in tooth formation and in epithelial morphogenesis more broadly.


Assuntos
Dente Molar/crescimento & desenvolvimento , Dente Molar/metabolismo , Odontogênese/fisiologia , Via de Sinalização Wnt/fisiologia , Animais , Proliferação de Células , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Epitélio/metabolismo , Mesoderma/metabolismo , Camundongos , Dente Molar/citologia , Morfogênese/fisiologia , Odontogênese/genética , beta Catenina/metabolismo
9.
Nutrients ; 13(7)2021 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-34209455

RESUMO

Glucose-based solutions remain the most used osmotic agents in peritoneal dialysis (PD), but unavoidably they contribute to the loss of peritoneal filtration capacity. Here, we evaluated at a molecular level the effects of XyloCore, a new PD solution with a low glucose content, in mesothelial and endothelial cells. Cell viability, integrity of mesothelial and endothelial cell membrane, activation of mesothelial and endothelial to mesenchymal transition programs, inflammation, and angiogenesis were evaluated by several techniques. Results showed that XyloCore preserves mesothelial and endothelial cell viability and membrane integrity. Moreover XyloCore, unlike glucose-based solutions, does not exert pro-fibrotic, -inflammatory, and -angiogenic effects. Overall, the in vitro evidence suggests that XyloCore could represent a potential biocompatible solution promising better outcomes in clinical practice.


Assuntos
Soluções para Diálise/farmacologia , Células Epiteliais/metabolismo , Epitélio/metabolismo , Glucose/farmacologia , Inflamação/patologia , Mesoderma/metabolismo , Neovascularização Fisiológica , Diálise Peritoneal , Biomarcadores/metabolismo , Linhagem Celular , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Transdiferenciação Celular/efeitos dos fármacos , Impedância Elétrica , Células Epiteliais/efeitos dos fármacos , Epitélio/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Mesoderma/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Neovascularização Fisiológica/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Permeabilidade , Fatores de Transcrição da Família Snail/genética , Fatores de Transcrição da Família Snail/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo
10.
Elife ; 102021 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-34227938

RESUMO

In classical descriptions of vertebrate development, the segregation of the three embryonic germ layers completes by the end of gastrulation. Body formation then proceeds in a head to tail fashion by progressive deposition of lineage-committed progenitors during regression of the primitive streak (PS) and tail bud (TB). The identification by retrospective clonal analysis of a population of neuromesodermal progenitors (NMPs) contributing to both musculoskeletal precursors (paraxial mesoderm) and spinal cord during axis formation challenged these notions. However, classical fate mapping studies of the PS region in amniotes have so far failed to provide direct evidence for such bipotential cells at the single-cell level. Here, using lineage tracing and single-cell RNA sequencing in the chicken embryo, we identify a resident cell population of the anterior PS epiblast, which contributes to neural and mesodermal lineages in trunk and tail. These cells initially behave as monopotent progenitors as classically described and only acquire a bipotential fate later, in more posterior regions. We show that NMPs exhibit a conserved transcriptomic signature during axis elongation but lose their epithelial characteristicsin the TB. Posterior to anterior gradients of convergence speed and ingression along the PS lead to asymmetric exhaustion of PS mesodermal precursor territories. Through limited ingression and increased proliferation, NMPs are maintained and amplified as a cell population which constitute the main progenitors in the TB. Together, our studies provide a novel understanding of the PS and TB contribution through the NMPs to the formation of the body of amniote embryos.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Mesoderma/embriologia , Células-Tronco Neurais/citologia , Linha Primitiva/embriologia , Animais , Padronização Corporal/genética , Diferenciação Celular/genética , Embrião de Galinha/embriologia , Mesoderma/metabolismo , Células-Tronco Neurais/fisiologia , Linha Primitiva/metabolismo
11.
Cell Death Dis ; 12(7): 677, 2021 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-34226515

RESUMO

Muscular dystrophies are debilitating neuromuscular disorders for which no cure exists. As this disorder affects both cardiac and skeletal muscle, patients would benefit from a cellular therapy that can simultaneously regenerate both tissues. The current protocol to derive bipotent mesodermal progenitors which can differentiate into cardiac and skeletal muscle relies on the spontaneous formation of embryoid bodies, thereby hampering further clinical translation. Additionally, as skeletal muscle is the largest organ in the human body, a high myogenic potential is necessary for successful regeneration. Here, we have optimized a protocol to generate chemically defined human induced pluripotent stem cell-derived mesodermal progenitors (cdMiPs). We demonstrate that these cells contribute to myotube formation and differentiate into cardiomyocytes, both in vitro and in vivo. Furthermore, the addition of valproic acid, a clinically approved small molecule, increases the potential of the cdMiPs to contribute to myotube formation that can be prevented by NOTCH signaling inhibitors. Moreover, valproic acid pre-treated cdMiPs injected in dystrophic muscles increase physical strength and ameliorate the functional performances of transplanted mice. Taken together, these results constitute a novel approach to generate mesodermal progenitors with enhanced myogenic potential using clinically approved reagents.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Mesoderma/efeitos dos fármacos , Desenvolvimento Muscular/efeitos dos fármacos , Fibras Musculares Esqueléticas/efeitos dos fármacos , Miócitos Cardíacos/efeitos dos fármacos , Receptores Notch/metabolismo , Ácido Valproico/farmacologia , Animais , Linhagem da Célula , Células Cultivadas , Técnicas de Cocultura , Modelos Animais de Doenças , Feminino , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/transplante , Masculino , Mesoderma/citologia , Mesoderma/metabolismo , Mesoderma/transplante , Camundongos , Camundongos Knockout , Contração Muscular , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/transplante , Força Muscular , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiopatologia , Distrofias Musculares/genética , Distrofias Musculares/metabolismo , Distrofias Musculares/fisiopatologia , Distrofias Musculares/cirurgia , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/transplante , Fenótipo , Ratos , Transdução de Sinais
12.
Development ; 148(12)2021 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-34142711

RESUMO

Axial elongation of the neural tube is crucial during mammalian embryogenesis for anterior-posterior body axis establishment and subsequent spinal cord development, but these processes cannot be interrogated directly in humans as they occur post-implantation. Here, we report an organoid model of neural tube extension derived from human pluripotent stem cell (hPSC) aggregates that have been caudalized with Wnt agonism, enabling them to recapitulate aspects of the morphological and temporal gene expression patterns of neural tube development. Elongating organoids consist largely of neuroepithelial compartments and contain TBXT+SOX2+ neuro-mesodermal progenitors in addition to PAX6+NES+ neural progenitors. A critical threshold of Wnt agonism stimulated singular axial extensions while maintaining multiple cell lineages, such that organoids displayed regionalized anterior-to-posterior HOX gene expression with hindbrain (HOXB1) regions spatially distinct from brachial (HOXC6) and thoracic (HOXB9) regions. CRISPR interference-mediated silencing of TBXT, a Wnt pathway target, increased neuroepithelial compartmentalization, abrogated HOX expression and disrupted uniaxial elongation. Together, these results demonstrate the potent capacity of caudalized hPSC organoids to undergo axial elongation in a manner that can be used to dissect the cellular organization and patterning decisions that dictate early human nervous system development.


Assuntos
Padronização Corporal , Tubo Neural/embriologia , Organogênese , Organoides , Padronização Corporal/efeitos dos fármacos , Diferenciação Celular , Desenvolvimento Embrionário , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Mesoderma/embriologia , Mesoderma/metabolismo , Neurogênese/efeitos dos fármacos , Organogênese/efeitos dos fármacos , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos
13.
Nat Commun ; 12(1): 3709, 2021 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-34140509

RESUMO

Fibrotic skin disease represents a major global healthcare burden, characterized by fibroblast hyperproliferation and excessive accumulation of extracellular matrix. Fibroblasts are found to be heterogeneous in multiple fibrotic diseases, but fibroblast heterogeneity in fibrotic skin diseases is not well characterized. In this study, we explore fibroblast heterogeneity in keloid, a paradigm of fibrotic skin diseases, by using single-cell RNA-seq. Our results indicate that keloid fibroblasts can be divided into 4 subpopulations: secretory-papillary, secretory-reticular, mesenchymal and pro-inflammatory. Interestingly, the percentage of mesenchymal fibroblast subpopulation is significantly increased in keloid compared to normal scar. Functional studies indicate that mesenchymal fibroblasts are crucial for collagen overexpression in keloid. Increased mesenchymal fibroblast subpopulation is also found in another fibrotic skin disease, scleroderma, suggesting this is a broad mechanism for skin fibrosis. These findings will help us better understand skin fibrotic pathogenesis, and provide potential targets for fibrotic disease therapies.


Assuntos
Colágeno/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Queloide/metabolismo , Mesoderma/citologia , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Colágeno/genética , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Matriz Extracelular/patologia , Fibroblastos/patologia , Regulação da Expressão Gênica/genética , Ontologia Genética , Humanos , Queloide/genética , Queloide/patologia , Ligantes , Mesoderma/metabolismo , Mesoderma/patologia , RNA-Seq , Escleroderma Sistêmico/genética , Escleroderma Sistêmico/metabolismo , Escleroderma Sistêmico/patologia , Análise de Célula Única , Dermatopatias/genética , Dermatopatias/metabolismo , Dermatopatias/patologia
14.
Nat Commun ; 12(1): 3851, 2021 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-34158501

RESUMO

Positional information driving limb muscle patterning is contained in connective tissue fibroblasts but not in myogenic cells. Limb muscles originate from somites, while connective tissues originate from lateral plate mesoderm. With cell and genetic lineage tracing we challenge this model and identify an unexpected contribution of lateral plate-derived fibroblasts to the myogenic lineage, preferentially at the myotendinous junction. Analysis of single-cell RNA-sequencing data from whole limbs at successive developmental stages identifies a population displaying a dual muscle and connective tissue signature. BMP signalling is active in this dual population and at the tendon/muscle interface. In vivo and in vitro gain- and loss-of-function experiments show that BMP signalling regulates a fibroblast-to-myoblast conversion. These results suggest a scenario in which BMP signalling converts a subset of lateral plate mesoderm-derived cells to a myogenic fate in order to create a boundary of fibroblast-derived myonuclei at the myotendinous junction that controls limb muscle patterning.


Assuntos
Padronização Corporal/genética , Fibroblastos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Músculo Esquelético/metabolismo , Somitos/metabolismo , Animais , Linhagem da Célula/genética , Células Cultivadas , Embrião de Galinha , Extremidades/embriologia , Fibroblastos/citologia , Mesoderma/citologia , Mesoderma/embriologia , Mesoderma/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Camundongos Transgênicos , Desenvolvimento Muscular/genética , Músculo Esquelético/citologia , Músculo Esquelético/embriologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Somitos/citologia , Somitos/embriologia
15.
Nucleic Acids Res ; 49(11): 6281-6295, 2021 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-34107030

RESUMO

Epigenetics, especially histone marks, functions beyond the DNA sequences to regulate gene expression. Depletion of NSD1, which catalyzes H3K36me2, leads to both up- and down-regulation of gene expression, indicating NSD1 is associated with both active and repressed gene expression. It's known that NSD1 regulates the deposition and expansion of H3K27me3, a repressive mark for gene expression, to keep active gene transcription. However, how NSD1 functions to repress gene expression is largely unknown. Here, we find that, when NSD1 is knocked out in mouse embryonic stem cells (mESCs), H3K27ac increases correlatively with the decrease of H3K36me2 at active enhancers, which is associated with mesoderm differentiation genes, leading to elevated gene expression. Mechanistically, NSD1 recruits HDAC1, the deacetylase of H3K27ac, to chromatin. Moreover, HDAC1 knockout (KO) recapitulates the increase of H3K27ac at active enhancers as the NSD1 depletion. Together, we propose that NSD1 deposits H3K36me2 and recruits HDAC1 at active enhancers to serve as a 'safeguard', preventing further activation of active enhancer-associated genes.


Assuntos
Elementos Facilitadores Genéticos , Regulação da Expressão Gênica , Código das Histonas , Histona-Lisina N-Metiltransferase/metabolismo , Animais , Linhagem Celular , Células Cultivadas , Cromatina/metabolismo , Células-Tronco Embrionárias/metabolismo , Técnicas de Inativação de Genes , Histona Desacetilase 1/genética , Histona Desacetilase 1/metabolismo , Histona-Lisina N-Metiltransferase/genética , Histonas/metabolismo , Mesoderma/metabolismo , Camundongos
16.
Development ; 148(11)2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-34061172

RESUMO

Organs stop growing to achieve a characteristic size and shape in scale with the body of an animal. Likewise, regenerating organs sense injury extents to instruct appropriate replacement growth. Fish fins exemplify both phenomena through their tremendous diversity of form and remarkably robust regeneration. The classic zebrafish mutant longfint2 develops and regenerates dramatically elongated fins and underlying ray skeleton. We show longfint2 chromosome 2 overexpresses the ether-a-go-go-related voltage-gated potassium channel kcnh2a. Genetic disruption of kcnh2a in cis rescues longfint2, indicating longfint2 is a regulatory kcnh2a allele. We find longfint2 fin overgrowth originates from prolonged outgrowth periods by showing Kcnh2a chemical inhibition during late stage regeneration fully suppresses overgrowth. Cell transplantations demonstrate longfint2-ectopic kcnh2a acts tissue autonomously within the fin intra-ray mesenchymal lineage. Temporal inhibition of the Ca2+-dependent phosphatase calcineurin indicates it likewise entirely acts late in regeneration to attenuate fin outgrowth. Epistasis experiments suggest longfint2-expressed Kcnh2a inhibits calcineurin output to supersede growth cessation signals. We conclude ion signaling within the growth-determining mesenchyme lineage controls fin size by tuning outgrowth periods rather than altering positional information or cell-level growth potency.


Assuntos
Nadadeiras de Animais/fisiologia , Expressão Ectópica do Gene/fisiologia , Canais de Potássio Éter-A-Go-Go/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Nadadeiras de Animais/anatomia & histologia , Animais , Sistemas CRISPR-Cas , Calcineurina/metabolismo , Proliferação de Células , Expressão Ectópica do Gene/genética , Éter , Canais de Potássio Éter-A-Go-Go/genética , Regulação da Expressão Gênica no Desenvolvimento , Mesoderma/metabolismo , Tamanho do Órgão , Regeneração/fisiologia , Transdução de Sinais/genética , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética
17.
Development ; 148(11)2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-34061173

RESUMO

During early embryogenesis, the vertebrate embryo extends from anterior to posterior because of the progressive addition of cells from a posteriorly localized neuromesodermal progenitor (NMp) population. An autoregulatory loop between Wnt and Brachyury/Tbxt is required for NMps to retain mesodermal potential and, hence, normal axis development. We recently showed that Hox13 genes help to support body axis formation and to maintain the autoregulatory loop, although the direct Hox13 target genes were unknown. Here, using a new method for identifying in vivo transcription factor-binding sites, we identified more than 500 potential Hox13 target genes in zebrafish. Importantly, we found two highly conserved Hox13-binding elements far from the tbxta transcription start site that also contain a conserved Tcf7/Lef1 (Wnt response) site. We show that the proximal of the two elements is sufficient to confer somitogenesis-stage expression to a tbxta promoter that, on its own, only drives NMp expression during gastrulation. Importantly, elimination of this proximal element produces shortened embryos due to aberrant formation of the most posterior somites. Our study provides a potential direct connection between Hox13 and regulation of the Wnt/Brachyury loop.


Assuntos
Proteínas Fetais/genética , Proteínas Fetais/metabolismo , Proteínas com Domínio T/genética , Proteínas com Domínio T/metabolismo , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Animais , Sítios de Ligação , Padronização Corporal , Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário , Regulação da Expressão Gênica no Desenvolvimento , Mesoderma/metabolismo , Somitos/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Via de Sinalização Wnt , Peixe-Zebra/embriologia
18.
Development ; 148(11)2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-34061174

RESUMO

During embryonic development, the otic epithelium and surrounding periotic mesenchymal cells originate from distinct lineages and coordinate to form the mammalian cochlea. Epithelial sensory precursors within the cochlear duct first undergo terminal mitosis before differentiating into sensory and non-sensory cells. In parallel, periotic mesenchymal cells differentiate to shape the lateral wall, modiolus and pericochlear spaces. Previously, Wnt activation was shown to promote proliferation and differentiation of both otic epithelial and mesenchymal cells. Here, we fate-mapped Wnt-responsive epithelial and mesenchymal cells in mice and found that Wnt activation resulted in opposing cell fates. In the post-mitotic cochlear epithelium, Wnt activation via ß-catenin stabilization induced clusters of proliferative cells that dedifferentiated and lost epithelial characteristics. In contrast, Wnt-activated periotic mesenchyme formed ectopic pericochlear spaces and cell clusters showing a loss of mesenchymal and gain of epithelial features. Finally, clonal analyses via multi-colored fate-mapping showed that Wnt-activated epithelial cells proliferated and formed clonal colonies, whereas Wnt-activated mesenchymal cells assembled as aggregates of mitotically quiescent cells. Together, we show that Wnt activation drives transition between epithelial and mesenchymal states in a cell type-dependent manner.


Assuntos
Cóclea/embriologia , Epitélio/metabolismo , Células-Tronco Mesenquimais/metabolismo , Via de Sinalização Wnt/fisiologia , Animais , Desdiferenciação Celular , Diferenciação Celular , Proliferação de Células , Cóclea/citologia , Cóclea/crescimento & desenvolvimento , Mesoderma/metabolismo , Camundongos , Camundongos Transgênicos , Proteínas Wnt , beta Catenina/metabolismo
19.
Development ; 148(14)2021 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-34184034

RESUMO

Signaling through the platelet-derived growth factor receptor alpha (PDGFRα) is crucial for mammalian craniofacial development, although the mechanisms by which the activity of downstream intracellular effectors is regulated to mediate gene expression changes have not been defined. We find that the RNA-binding protein Srsf3 is phosphorylated at Akt consensus sites downstream of PI3K-mediated PDGFRα signaling in mouse palatal mesenchyme cells, leading to its nuclear translocation. We further demonstrate that ablation of Srsf3 in the mouse neural crest lineage leads to facial clefting due to defective cranial neural crest cell proliferation and survival. Finally, we show that Srsf3 regulates the alternative RNA splicing of transcripts encoding protein kinases in the mouse facial process mesenchyme to regulate PDGFRα-dependent intracellular signaling. Collectively, our findings reveal that alternative RNA splicing is an important mechanism of gene expression regulation downstream of PI3K/Akt-mediated PDGFRα signaling in the facial mesenchyme and identify Srsf3 as a critical regulator of craniofacial development.


Assuntos
Processamento Alternativo , Mesoderma/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Fatores de Processamento de Serina-Arginina/metabolismo , Transdução de Sinais , Animais , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Ligantes , Masculino , Camundongos , Camundongos Knockout , Crista Neural/embriologia , Crista Neural/metabolismo , Fenótipo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas de Ligação a RNA/metabolismo , Fatores de Processamento de Serina-Arginina/genética
20.
Nature ; 595(7865): 85-90, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33981037

RESUMO

The ontogeny of the human haematopoietic system during fetal development has previously been characterized mainly through careful microscopic observations1. Here we reconstruct a phylogenetic tree of blood development using whole-genome sequencing of 511 single-cell-derived haematopoietic colonies from healthy human fetuses at 8 and 18 weeks after conception, coupled with deep targeted sequencing of tissues of known embryonic origin. We found that, in healthy fetuses, individual haematopoietic progenitors acquire tens of somatic mutations by 18 weeks after conception. We used these mutations as barcodes and timed the divergence of embryonic and extra-embryonic tissues during development, and estimated the number of blood antecedents at different stages of embryonic development. Our data support a hypoblast origin of the extra-embryonic mesoderm and primitive blood in humans.


Assuntos
Linhagem da Célula/genética , Desenvolvimento Embrionário/genética , Sistema Hematopoético/embriologia , Sistema Hematopoético/metabolismo , Mutação , Células Sanguíneas/citologia , Células Sanguíneas/metabolismo , Células Clonais/citologia , Células Clonais/metabolismo , Análise Mutacional de DNA , Feto/citologia , Feto/embriologia , Feto/metabolismo , Camadas Germinativas/citologia , Camadas Germinativas/metabolismo , Saúde , Sistema Hematopoético/citologia , Humanos , Cariotipagem , Masculino , Mesoderma/citologia , Mesoderma/embriologia , Mesoderma/metabolismo , Taxa de Mutação , Especificidade de Órgãos/genética , Fatores de Tempo , Sequenciamento Completo do Genoma , Fluxo de Trabalho
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