RESUMO
Centromeres are scaffolds for the assembly of kinetochores that ensure chromosome segregation during cell division. How vertebrate centromeres obtain a three-dimensional structure to accomplish their primary function is unclear. Using super-resolution imaging, capture-C, and polymer modeling, we show that vertebrate centromeres are partitioned by condensins into two subdomains during mitosis. The bipartite structure is found in human, mouse, and chicken cells and is therefore a fundamental feature of vertebrate centromeres. Super-resolution imaging and electron tomography reveal that bipartite centromeres assemble bipartite kinetochores, with each subdomain binding a distinct microtubule bundle. Cohesin links the centromere subdomains, limiting their separation in response to spindle forces and avoiding merotelic kinetochore-spindle attachments. Lagging chromosomes during cancer cell divisions frequently have merotelic attachments in which the centromere subdomains are separated and bioriented. Our work reveals a fundamental aspect of vertebrate centromere biology with implications for understanding the mechanisms that guarantee faithful chromosome segregation.
Assuntos
Centrômero , Coesinas , Cinetocoros , Mitose , Animais , Humanos , Camundongos , Proteínas de Ciclo Celular/metabolismo , Centrômero/metabolismo , Galinhas , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Cromossômicas não Histona/química , Segregação de Cromossomos , Cinetocoros/metabolismo , Microtúbulos/metabolismo , Fuso Acromático/metabolismoRESUMO
H3N8 avian influenza viruses (AIVs) in China caused two confirmed human infections in 2022, followed by a fatal case reported in 2023. H3N8 viruses are widespread in chicken flocks; however, the zoonotic features of H3N8 viruses are poorly understood. Here, we demonstrate that H3N8 viruses were able to infect and replicate efficiently in organotypic normal human bronchial epithelial (NHBE) cells and lung epithelial (Calu-3) cells. Human isolates of H3N8 virus were more virulent and caused severe pathology in mice and ferrets, relative to chicken isolates. Importantly, H3N8 virus isolated from a patient with severe pneumonia was transmissible between ferrets through respiratory droplets; it had acquired human-receptor-binding preference and amino acid substitution PB2-E627K necessary for airborne transmission. Human populations, even when vaccinated against human H3N2 virus, appear immunologically naive to emerging mammalian-adapted H3N8 AIVs and could be vulnerable to infection at epidemic or pandemic proportion.
Assuntos
Vírus da Influenza A Subtipo H3N8 , Influenza Humana , Animais , Humanos , Camundongos , Galinhas , Furões , Vírus da Influenza A Subtipo H3N2 , Aerossóis e Gotículas RespiratóriosRESUMO
Membrane remodeling and repair are essential for all cells. Proteins that perform these functions include Vipp1/IM30 in photosynthetic plastids, PspA in bacteria, and ESCRT-III in eukaryotes. Here, using a combination of evolutionary and structural analyses, we show that these protein families are homologous and share a common ancient evolutionary origin that likely predates the last universal common ancestor. This homology is evident in cryo-electron microscopy structures of Vipp1 rings from the cyanobacterium Nostoc punctiforme presented over a range of symmetries. Each ring is assembled from rungs that stack and progressively tilt to form dome-shaped curvature. Assembly is facilitated by hinges in the Vipp1 monomer, similar to those in ESCRT-III proteins, which allow the formation of flexible polymers. Rings have an inner lumen that is able to bind and deform membranes. Collectively, these data suggest conserved mechanistic principles that underlie Vipp1, PspA, and ESCRT-III-dependent membrane remodeling across all domains of life.
Assuntos
Proteínas de Bactérias/metabolismo , Membrana Celular/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Proteínas de Choque Térmico/metabolismo , Família Multigênica , Nostoc/metabolismo , Sequência de Aminoácidos , Animais , Proteínas de Bactérias/química , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/ultraestrutura , Galinhas , Microscopia Crioeletrônica , Complexos Endossomais de Distribuição Requeridos para Transporte/química , Evolução Molecular , Proteínas de Choque Térmico/química , Proteínas de Choque Térmico/ultraestrutura , Humanos , Modelos Moleculares , Estrutura Secundária de Proteína , Homologia de Sequência de Aminoácidos , TermodinâmicaRESUMO
The cholesterol-sensing protein Scap induces cholesterol synthesis by transporting membrane-bound transcription factors called sterol regulatory element-binding proteins (SREBPs) from the endoplasmic reticulum (ER) to the Golgi apparatus for proteolytic activation. Transport requires interaction between Scap's two ER luminal loops (L1 and L7), which flank an intramembrane sterol-sensing domain (SSD). Cholesterol inhibits Scap transport by binding to L1, which triggers Scap's binding to Insig, an ER retention protein. Here we used cryoelectron microscopy (cryo-EM) to elucidate two structures of full-length chicken Scap: (1) a wild-type free of Insigs and (2) mutant Scap bound to chicken Insig without cholesterol. Strikingly, L1 and L7 intertwine tightly to form a globular domain that acts as a luminal platform connecting the SSD to the rest of Scap. In the presence of Insig, this platform undergoes a large rotation accompanied by rearrangement of Scap's transmembrane helices. We postulate that this conformational change halts Scap transport of SREBPs and inhibits cholesterol synthesis.
Assuntos
Colesterol/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Sequência de Aminoácidos , Animais , Anticorpos/metabolismo , Galinhas , Proteínas de Membrana/isolamento & purificação , Proteínas de Membrana/ultraestrutura , Modelos Biológicos , Modelos Moleculares , Ligação Proteica , Domínios Proteicos , Estrutura Secundária de Proteína , Relação Estrutura-AtividadeRESUMO
Organoids capable of forming tissue-like structures have transformed our ability to model human development and disease. With the notable exception of the human heart, lineage-specific self-organizing organoids have been reported for all major organs. Here, we established self-organizing cardioids from human pluripotent stem cells that intrinsically specify, pattern, and morph into chamber-like structures containing a cavity. Cardioid complexity can be controlled by signaling that instructs the separation of cardiomyocyte and endothelial layers and by directing epicardial spreading, inward migration, and differentiation. We find that cavity morphogenesis is governed by a mesodermal WNT-BMP signaling axis and requires its target HAND1, a transcription factor linked to developmental heart chamber defects. Upon cryoinjury, cardioids initiated a cell-type-dependent accumulation of extracellular matrix, an early hallmark of both regeneration and heart disease. Thus, human cardioids represent a powerful platform to mechanistically dissect self-organization, congenital heart defects and serve as a foundation for future translational research.
Assuntos
Coração/embriologia , Organogênese , Organoides/embriologia , Ativinas/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Proteínas Morfogenéticas Ósseas/metabolismo , Cálcio/metabolismo , Linhagem Celular , Linhagem da Célula , Galinhas , Células Endoteliais/citologia , Proteínas da Matriz Extracelular/metabolismo , Feminino , Fibroblastos/citologia , Proteína Homeobox Nkx-2.5/metabolismo , Humanos , Masculino , Mesoderma/embriologia , Modelos Biológicos , Miocárdio/metabolismo , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Proteínas Wnt/metabolismoRESUMO
This year's Lasker Basic Medical Research Award honors Max Cooper and Jacques Miller for discoveries that revealed the organizing principles of adaptive immunity. Their collective contributions have had broad clinical impact in the treatment of immune disease.
Assuntos
Imunidade Adaptativa/imunologia , Comunicação Celular/imunologia , Plasmócitos/imunologia , Linfócitos T/imunologia , Animais , Formação de Anticorpos/imunologia , Apresentação de Antígeno , Medula Óssea/imunologia , Galinhas , Humanos , Hibridomas/imunologia , Switching de Imunoglobulina/imunologia , Linfonodos/imunologia , Camundongos , Prêmio Nobel , Tolerância a Antígenos Próprios/imunologia , Timo/imunologiaRESUMO
Microglia, the brain-resident immune cells, are critically involved in many physiological and pathological brain processes, including neurodegeneration. Here we characterize microglia morphology and transcriptional programs across ten species spanning more than 450 million years of evolution. We find that microglia express a conserved core gene program of orthologous genes from rodents to humans, including ligands and receptors associated with interactions between glia and neurons. In most species, microglia show a single dominant transcriptional state, whereas human microglia display significant heterogeneity. In addition, we observed notable differences in several gene modules of rodents compared with primate microglia, including complement, phagocytic, and susceptibility genes to neurodegeneration, such as Alzheimer's and Parkinson's disease. Our study provides an essential resource of conserved and divergent microglia pathways across evolution, with important implications for future development of microglia-based therapies in humans.
Assuntos
Evolução Molecular , Redes Reguladoras de Genes , Microglia/metabolismo , Doenças Neurodegenerativas/genética , Análise de Célula Única , Transcriptoma , Animais , Galinhas , Perfilação da Expressão Gênica , Predisposição Genética para Doença , Humanos , Primatas , Répteis , Roedores , Ovinos , Suínos , Peixe-ZebraRESUMO
B cells constitute an essential line of defense from pathogenic infections through the generation of class-switched antibody-secreting cells (ASCs) in germinal centers. Although this process is known to be regulated by follicular helper T (TfH) cells, the mechanism by which B cells initially seed germinal center reactions remains elusive. We found that NKT cells, a population of innate-like T lymphocytes, are critical for the induction of B cell immunity upon viral infection. The positioning of NKT cells at the interfollicular areas of lymph nodes facilitates both their direct priming by resident macrophages and the localized delivery of innate signals to antigen-experienced B cells. Indeed, NKT cells secrete an early wave of IL-4 and constitute up to 70% of the total IL-4-producing cells during the initial stages of infection. Importantly, the requirement of this innate immunity arm appears to be evolutionarily conserved because early NKT and IL-4 gene signatures also positively correlate with the levels of neutralizing antibodies in Zika-virus-infected macaques. In conclusion, our data support a model wherein a pre-TfH wave of IL-4 secreted by interfollicular NKT cells triggers the seeding of germinal center cells and serves as an innate link between viral infection and B cell immunity.
Assuntos
Linfócitos B/imunologia , Centro Germinativo/imunologia , Imunidade Inata , Influenza Humana/imunologia , Interleucina-4/genética , Células Matadoras Naturais/imunologia , Infecção por Zika virus/imunologia , Animais , Galinhas , Cães , Centro Germinativo/citologia , Humanos , Interleucina-4/metabolismo , Macaca , Macrófagos/imunologia , Células Madin Darby de Rim Canino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Walking is the predominant locomotor behavior expressed by land-dwelling vertebrates, but it is unknown when the neural circuits that are essential for limb control first appeared. Certain fish species display walking-like behaviors, raising the possibility that the underlying circuitry originated in primitive marine vertebrates. We show that the neural substrates of bipedalism are present in the little skate Leucoraja erinacea, whose common ancestor with tetrapods existed â¼420 million years ago. Leucoraja exhibits core features of tetrapod locomotor gaits, including left-right alternation and reciprocal extension-flexion of the pelvic fins. Leucoraja also deploys a remarkably conserved Hox transcription factor-dependent program that is essential for selective innervation of fin/limb muscle. This network encodes peripheral connectivity modules that are distinct from those used in axial muscle-based swimming and has apparently been diminished in most modern fish. These findings indicate that the circuits that are essential for walking evolved through adaptation of a genetic regulatory network shared by all vertebrates with paired appendages. VIDEO ABSTRACT.
Assuntos
Proteínas Aviárias , Galinhas/fisiologia , Evolução Molecular , Proteínas de Peixes , Proteínas de Homeodomínio , Rede Nervosa/fisiologia , Rajidae/fisiologia , Fatores de Transcrição , Caminhada/fisiologia , Peixe-Zebra/fisiologia , Nadadeiras de Animais/fisiologia , Animais , Proteínas Aviárias/genética , Proteínas Aviárias/metabolismo , Embrião de Galinha , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Músculo Esquelético/fisiologia , Natação/fisiologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
CD4+ T cells orchestrate the adaptive immune response against pathogens and cancer by recognizing epitopes presented on class II major histocompatibility complex (MHC-II) molecules. The high polymorphism of MHC-II genes represents an important hurdle toward accurate prediction and identification of CD4+ T cell epitopes. Here we collected and curated a dataset of 627,013 unique MHC-II ligands identified by mass spectrometry. This enabled us to precisely determine the binding motifs of 88 MHC-II alleles across humans, mice, cattle, and chickens. Analysis of these binding specificities combined with X-ray crystallography refined our understanding of the molecular determinants of MHC-II motifs and revealed a widespread reverse-binding mode in HLA-DP ligands. We then developed a machine-learning framework to accurately predict binding specificities and ligands of any MHC-II allele. This tool improves and expands predictions of CD4+ T cell epitopes and enables us to discover viral and bacterial epitopes following the aforementioned reverse-binding mode.
Assuntos
Epitopos de Linfócito T , Peptídeos , Humanos , Animais , Camundongos , Bovinos , Ligantes , Ligação Proteica , Galinhas/metabolismo , Aprendizado de Máquina , Antígenos de Histocompatibilidade Classe II , AlelosRESUMO
The stable structural conformations that occur along the complete reaction coordinate for ion channel opening have never been observed. In this study, we describe the equilibrium ensemble of structures of Slo2.2, a neuronal Na+-activated K+ channel, as a function of the Na+ concentration. We find that Slo2.2 exists in multiple closed conformations whose relative occupancies are independent of Na+ concentration. An open conformation emerges from an ensemble of closed conformations in a highly Na+-dependent manner, without evidence of Na+-dependent intermediates. In other words, channel opening is a highly concerted, switch-like process. The midpoint of the structural titration matches that of the functional titration. A maximum open conformation probability approaching 1.0 and maximum functional open probability approaching 0.7 imply that, within the class of open channels, there is a subclass that is not permeable to ions.
Assuntos
Proteínas Aviárias/química , Galinhas/metabolismo , Proteínas do Tecido Nervoso/química , Canais de Potássio/química , Animais , Proteínas Aviárias/metabolismo , Microscopia Crioeletrônica , Células HEK293 , Humanos , Proteínas do Tecido Nervoso/metabolismo , Canais de Potássio/metabolismo , Conformação Proteica , Sódio/químicaRESUMO
Recent advances in single-particle cryoelecton microscopy (cryo-EM) are enabling generation of numerous near-atomic resolution structures for well-ordered protein complexes with sizes ≥ â¼200 kDa. Whether cryo-EM methods are equally useful for high-resolution structural analysis of smaller, dynamic protein complexes such as those involved in cellular metabolism remains an important question. Here, we present 3.8 Å resolution cryo-EM structures of the cancer target isocitrate dehydrogenase (93 kDa) and identify the nature of conformational changes induced by binding of the allosteric small-molecule inhibitor ML309. We also report 2.8-Å- and 1.8-Å-resolution structures of lactate dehydrogenase (145 kDa) and glutamate dehydrogenase (334 kDa), respectively. With these results, two perceived barriers in single-particle cryo-EM are overcome: (1) crossing 2 Å resolution and (2) obtaining structures of proteins with sizes < 100 kDa, demonstrating that cryo-EM can be used to investigate a broad spectrum of drug-target interactions and dynamic conformational states.
Assuntos
Descoberta de Drogas , Glutamato Desidrogenase/ultraestrutura , Isocitrato Desidrogenase/ultraestrutura , L-Lactato Desidrogenase/ultraestrutura , Aminoquinolinas/química , Aminoquinolinas/farmacologia , Animais , Bovinos , Galinhas , Microscopia Crioeletrônica , Cristalografia por Raios X , Glutamato Desidrogenase/antagonistas & inibidores , Glutamato Desidrogenase/química , Humanos , Isocitrato Desidrogenase/antagonistas & inibidores , Isocitrato Desidrogenase/química , L-Lactato Desidrogenase/antagonistas & inibidores , L-Lactato Desidrogenase/química , Modelos Moleculares , Conformação Proteica , Sulfonamidas/química , Sulfonamidas/farmacologiaRESUMO
This year marks the 150(th) anniversary of the presentation by Gregor Mendel of his studies of plant hybridization to the Brunn Natural History Society. Their nature and meaning have been discussed many times. However, on this occasion, we reflect on the scientific enterprise and the perception of new discoveries.
Assuntos
Genética/história , Modelos Genéticos , Animais , Galinhas/genética , Cruzamentos Genéticos , História do Século XVIII , Pisum sativum/genética , Zea mays/genéticaRESUMO
Pericentric heterochromatin is a critical component of chromosomes marked by histone H3 K9 (H3K9) methylation1-3. However, what recruits H3K9-specific histone methyltransferases to pericentric regions in vertebrates remains unclear4, as does why pericentric regions in different species share the same H3K9 methylation mark despite lacking highly conserved DNA sequences2,5. Here we show that zinc-finger proteins ZNF512 and ZNF512B specifically localize at pericentric regions through direct DNA binding. Notably, both ZNF512 and ZNF512B are sufficient to initiate de novo heterochromatin formation at ectopically targeted repetitive regions and pericentric regions, as they directly recruit SUV39H1 and SUV39H2 (SUV39H) to catalyse H3K9 methylation. SUV39H2 makes a greater contribution to H3K9 trimethylation, whereas SUV39H1 seems to contribute more to silencing, probably owing to its preferential association with HP1 proteins. ZNF512 and ZNF512B from different species can specifically target pericentric regions of other vertebrates, because the atypical long linker residues between the zinc-fingers of ZNF512 and ZNF512B offer flexibility in recognition of non-consecutively organized three-nucleotide triplets targeted by each zinc-finger. This study addresses two long-standing questions: how constitutive heterochromatin is initiated and how seemingly variable pericentric sequences are targeted by the same set of conserved machinery in vertebrates.
Assuntos
Centrômero , Evolução Molecular , Heterocromatina , Histona-Lisina N-Metiltransferase , Histonas , Motivos de Nucleotídeos , Animais , Humanos , Camundongos , Centrômero/genética , Centrômero/metabolismo , Galinhas , Homólogo 5 da Proteína Cromobox , Inativação Gênica , Heterocromatina/metabolismo , Heterocromatina/química , Heterocromatina/genética , Histona-Lisina N-Metiltransferase/metabolismo , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/química , Histonas/metabolismo , Histonas/química , Anfioxos , Metilação , Petromyzon , Proteínas Repressoras/metabolismo , Proteínas Repressoras/química , Proteínas Repressoras/genética , Serpentes , Xenopus laevis , Peixe-Zebra , Dedos de ZincoRESUMO
We have used a combination of chemical genetics, chromatin proteomics, and imaging to map the earliest chromatin transactions during vertebrate cell entry into mitosis. Chicken DT40 CDK1as cells undergo synchronous mitotic entry within 15 min following release from a 1NM-PP1-induced arrest in late G2. In addition to changes in chromatin association with nuclear pores and the nuclear envelope, earliest prophase is dominated by changes in the association of ribonucleoproteins with chromatin, particularly in the nucleolus, where pre-rRNA processing factors leave chromatin significantly before RNA polymerase I. Nuclear envelope barrier function is lost early in prophase, and cytoplasmic proteins begin to accumulate on the chromatin. As a result, outer kinetochore assembly appears complete by nuclear envelope breakdown (NEBD). Most interphase chromatin proteins remain associated with chromatin until NEBD, after which their levels drop sharply. An interactive proteomic map of chromatin transactions during mitotic entry is available as a resource at https://mitoChEP.bio.ed.ac.uk.
Assuntos
Montagem e Desmontagem da Cromatina , Cromatina/metabolismo , Cromossomos , DNA/metabolismo , Linfoma de Células B/metabolismo , Proteínas Nucleares/metabolismo , Prófase , Proteoma , Proteômica , Animais , Proteína Quinase CDC2/genética , Proteína Quinase CDC2/metabolismo , Linhagem Celular Tumoral , Galinhas , Cromatina/genética , DNA/genética , Lamina Tipo B/genética , Lamina Tipo B/metabolismo , Linfoma de Células B/genética , Linfoma de Células B/patologia , Proteínas Nucleares/genética , Ligação Proteica , RNA Ribossômico/genética , RNA Ribossômico/metabolismo , Fatores de TempoRESUMO
Acid-sensing ion channels (ASICs) detect extracellular protons produced during inflammation or ischemic injury and belong to the superfamily of degenerin/epithelial sodium channels. Here, we determine the cocrystal structure of chicken ASIC1a with MitTx, a pain-inducing toxin from the Texas coral snake, to define the structure of the open state of ASIC1a. In the MitTx-bound open state and in the previously determined low-pH desensitized state, TM2 is a discontinuous α helix in which the Gly-Ala-Ser selectivity filter adopts an extended, belt-like conformation, swapping the cytoplasmic one-third of TM2 with an adjacent subunit. Gly 443 residues of the selectivity filter provide a ring of three carbonyl oxygen atoms with a radius of â¼3.6 Å, presenting an energetic barrier for hydrated ions. The ASIC1a-MitTx complex illuminates the mechanism of MitTx action, defines the structure of the selectivity filter of voltage-independent, sodium-selective ion channels, and captures the open state of an ASIC.
Assuntos
Canais Iônicos Sensíveis a Ácido/química , Proteínas Aviárias/química , Galinhas , Venenos Elapídicos/química , Elapidae , Canais Iônicos Sensíveis a Ácido/metabolismo , Sequência de Aminoácidos , Animais , Proteínas Aviárias/metabolismo , Cristalografia por Raios X , Venenos Elapídicos/metabolismo , Modelos Moleculares , Dados de Sequência Molecular , Alinhamento de Sequência , Canais de Sódio/químicaRESUMO
I am a developmental biologist, but I started off as a civil engineer. I did some research on soil mechanics but decided to change to biology. A friend changed my life when he told me about the mechanics of cell division, on which I did my PhD at Kings College. I then worked on the morphogenesis of the sea urchin embryo and became interested in how embryos are patterned, and I proposed positional information as a basic mechanism. I was a professor at the Middlesex Hospital Medical School, where we concentrated on how the chick limb developed.
Assuntos
Morfogênese/fisiologia , Animais , Galinhas/crescimento & desenvolvimento , Biologia do Desenvolvimento/métodos , Ouriços-do-Mar/embriologiaRESUMO
Centromere protein A (CENP-A) nucleosomes containing the centromere-specific histone H3 variant CENP-A represent an epigenetic mark that specifies centromere position. The Mis18 complex is a licensing factor for new CENP-A deposition via the CENP-A chaperone, Holliday junction recognition protein (HJURP), on the centromere chromatin. Chicken KINETOCHORE NULL2 (KNL2) (ggKNL2), a Mis18 complex component, has a CENP-C-like motif, and our previous study suggested that ggKNL2 directly binds to the CENP-A nucleosome to recruit HJURP/CENP-A to the centromere. However, the molecular basis for CENP-A nucleosome recognition by ggKNL2 has remained unclear. Here, we present the cryo-EM structure of the chicken CENP-A nucleosome in complex with a ggKNL2 fragment containing the CENP-C-like motif. Chicken KNL2 distinguishes between CENP-A and histone H3 in the nucleosome using the CENP-C-like motif and its downstream region. Both the C-terminal tail and the RG-loop of CENP-A are simultaneously recognized as CENP-A characteristics. The CENP-A nucleosome-ggKNL2 interaction is thus essential for KNL2 functions. Furthermore, our structural, biochemical, and cell biology data indicate that ggKNL2 changes its binding partner at the centromere during chicken cell cycle progression.
Assuntos
Histonas , Nucleossomos , Autoantígenos/genética , Autoantígenos/metabolismo , Proteínas de Ciclo Celular/metabolismo , Centrômero/metabolismo , Proteína Centromérica A/metabolismo , Microscopia Crioeletrônica , Histonas/metabolismo , Proteínas de Ligação a DNA/química , Animais , GalinhasRESUMO
The cellular and genetic networks that contribute to the development of the zeugopod (radius and ulna of the forearm, tibia and fibula of the leg) are not well understood, although these bones are susceptible to loss in congenital human syndromes and to the action of teratogens such as thalidomide. Using a new fate-mapping approach with the Chameleon transgenic chicken line, we show that there is a small contribution of SHH-expressing cells to the posterior ulna, posterior carpals and digit 3. We establish that although the majority of the ulna develops in response to paracrine SHH signalling in both the chicken and mouse, there are differences in the contribution of SHH-expressing cells between mouse and chicken as well as between the chicken ulna and fibula. This is evidence that, although zeugopod bones are clearly homologous according to the fossil record, the gene regulatory networks that contribute to their development and evolution are not fixed.
Assuntos
Animais Geneticamente Modificados , Galinhas , Proteínas Hedgehog , Animais , Proteínas Hedgehog/metabolismo , Proteínas Hedgehog/genética , Galinhas/genética , Camundongos , Evolução Biológica , Embrião de Galinha , Ulna , Regulação da Expressão Gênica no Desenvolvimento , Fíbula/metabolismo , Rádio (Anatomia)/metabolismo , Humanos , Extremidades/embriologiaRESUMO
During tissue regeneration, proliferation, dedifferentiation and reprogramming are necessary to restore lost structures. However, it is not fully understood how metabolism intersects with these processes. Chicken embryos can regenerate their retina through retinal pigment epithelium (RPE) reprogramming when treated with fibroblast factor 2 (FGF2). Using transcriptome profiling, we uncovered extensive regulation of gene sets pertaining to proliferation, neurogenesis and glycolysis throughout RPE-to-neural retina reprogramming. By manipulating cell media composition, we determined that glucose, glutamine or pyruvate are individually sufficient to support RPE reprogramming, identifying glycolysis as a requisite. Conversely, the activation of pyruvate dehydrogenase by inhibition of pyruvate dehydrogenase kinases, induces epithelial-to-mesenchymal transition, while simultaneously blocking the activation of neural retina fate. We also identified that epithelial-to-mesenchymal transition fate is partially driven by an oxidative environment. Our findings provide evidence that metabolism controls RPE cell fate decisions and provide insights into the metabolic state of RPE cells, which are prone to fate changes in regeneration and pathologies, such as proliferative vitreoretinopathy.