RESUMEN
During development innate lymphoid cells and specialized lymphocyte subsets colonize peripheral tissues, where they contribute to organogenesis and later constitute the first line of protection while maintaining tissue homeostasis. A few of these subsets are produced only during embryonic development and remain in the tissues throughout life. They are generated through a unique developmental program initiated in lympho-myeloid-primed progenitors, which lose myeloid and B cell potential. They either differentiate into innate lymphoid cells or migrate to the thymus to give rise to embryonic T cell receptor-invariant T cells. At later developmental stages, adaptive T lymphocytes are derived from lympho-myeloid progenitors that colonize the thymus, while lymphoid progenitors become specialized in the production of B cells. This sequence of events highlights the requirement for stratification in the establishment of immune functions that determine efficient seeding of peripheral tissues by a limited number of cells.
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Linfocitos B/inmunología , Linfocitos/fisiología , Células Progenitoras Linfoides/fisiología , Células T Asesinas Naturales/inmunología , Timo/inmunología , Animales , Diferenciación Celular , Linaje de la Célula , Microambiente Celular , Citocinas/metabolismo , Humanos , Inmunidad Innata , Activación de Linfocitos , Comunicación Paracrina , TranscriptomaRESUMEN
A fundamental question in developmental immunology is how bipotential thymocyte precursors generate both CD4+ helper and CD8+ cytotoxic T cell lineages. The MHC specificity of αß T cell receptors (TCRs) on precursors is closely correlated with cell fate-determining processes, prompting studies to characterize how variations in TCR signaling are linked with genetic programs establishing lineage-specific gene expression signatures, such as exclusive CD4 or CD8 expression. The key transcription factors ThPOK and Runx3 have been identified as mediating development of helper and cytotoxic T cell lineages, respectively. Together with increasing knowledge of epigenetic regulators, these findings have advanced our understanding of the transcription factor network regulating the CD4/CD8 dichotomy. It has also become apparent that CD4+ T cells retain developmental plasticity, allowing them to acquire cytotoxic activity in the periphery. Despite such advances, further studies are necessary to identify the molecular links between TCR signaling and the nuclear machinery regulating expression of ThPOK and Runx3.
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Diferenciación Celular/inmunología , Linfocitos T Citotóxicos/citología , Linfocitos T Citotóxicos/inmunología , Linfocitos T Colaboradores-Inductores/citología , Linfocitos T Colaboradores-Inductores/inmunología , Animales , Antígenos CD4/genética , Antígenos CD4/metabolismo , Antígenos CD8/genética , Antígenos CD8/metabolismo , Diferenciación Celular/genética , Linaje de la Célula/genética , Linaje de la Célula/inmunología , Subunidad alfa 3 del Factor de Unión al Sitio Principal/genética , Proteínas de Unión al ADN/genética , Regulación de la Expresión Génica , Humanos , Inmunomodulación/genética , Inmunomodulación/inmunología , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T/metabolismo , Secuencias Reguladoras de Ácidos Nucleicos , Linfocitos T Citotóxicos/metabolismo , Linfocitos T Colaboradores-Inductores/metabolismo , Factores de Transcripción/genética , Transcripción GenéticaRESUMEN
Platelet dysregulation is drastically increased with advanced age and contributes to making cardiovascular disorders the leading cause of death of elderly humans. Here, we reveal a direct differentiation pathway from hematopoietic stem cells into platelets that is progressively propagated upon aging. Remarkably, the aging-enriched platelet path is decoupled from all other hematopoietic lineages, including erythropoiesis, and operates as an additional layer in parallel with canonical platelet production. This results in two molecularly and functionally distinct populations of megakaryocyte progenitors. The age-induced megakaryocyte progenitors have a profoundly enhanced capacity to engraft, expand, restore, and reconstitute platelets in situ and upon transplantation and produce an additional platelet population in old mice. The two pools of co-existing platelets cause age-related thrombocytosis and dramatically increased thrombosis in vivo. Strikingly, aging-enriched platelets are functionally hyper-reactive compared with the canonical platelet populations. These findings reveal stem cell-based aging as a mechanism for platelet dysregulation and age-induced thrombosis.
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Envejecimiento , Plaquetas , Diferenciación Celular , Células Madre Hematopoyéticas , Trombosis , Animales , Células Madre Hematopoyéticas/metabolismo , Plaquetas/metabolismo , Trombosis/patología , Trombosis/metabolismo , Ratones , Humanos , Megacariocitos/metabolismo , Ratones Endogámicos C57BL , Células Progenitoras de Megacariocitos/metabolismo , MasculinoRESUMEN
Retrospective lineage reconstruction of humans predicts that dramatic clonal imbalances in the body can be traced to the 2-cell stage embryo. However, whether and how such clonal asymmetries arise in the embryo is unclear. Here, we performed prospective lineage tracing of human embryos using live imaging, non-invasive cell labeling, and computational predictions to determine the contribution of each 2-cell stage blastomere to the epiblast (body), hypoblast (yolk sac), and trophectoderm (placenta). We show that the majority of epiblast cells originate from only one blastomere of the 2-cell stage embryo. We observe that only one to three cells become internalized at the 8-to-16-cell stage transition. Moreover, these internalized cells are more frequently derived from the first cell to divide at the 2-cell stage. We propose that cell division dynamics and a cell internalization bottleneck in the early embryo establish asymmetry in the clonal composition of the future human body.
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Blastómeros , Linaje de la Célula , Embrión de Mamíferos , Femenino , Humanos , Blastómeros/citología , Blastómeros/metabolismo , División Celular , Embrión de Mamíferos/citología , Embrión de Mamíferos/metabolismo , Desarrollo Embrionario , Estratos Germinativos/citología , Estratos Germinativos/metabolismo , Masculino , Animales , RatonesRESUMEN
During wound healing, different pools of stem cells (SCs) contribute to skin repair. However, how SCs become activated and drive the tissue remodeling essential for skin repair is still poorly understood. Here, by developing a mouse model allowing lineage tracing and basal cell lineage ablation, we monitor SC fate and tissue dynamics during regeneration using confocal and intravital imaging. Analysis of basal cell rearrangements shows dynamic transitions from a solid-like homeostatic state to a fluid-like state allowing tissue remodeling during repair, as predicted by a minimal mathematical modeling of the spatiotemporal dynamics and fate behavior of basal cells. The basal cell layer progressively returns to a solid-like state with re-epithelialization. Bulk, single-cell RNA, and epigenetic profiling of SCs, together with functional experiments, uncover a common regenerative state regulated by the EGFR/AP1 axis activated during tissue fluidization that is essential for skin SC activation and tissue repair.
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In the prevailing model, Lgr5+ cells are the only intestinal stem cells (ISCs) that sustain homeostatic epithelial regeneration by upward migration of progeny through elusive upper crypt transit-amplifying (TA) intermediates. Here, we identify a proliferative upper crypt population marked by Fgfbp1, in the location of putative TA cells, that is transcriptionally distinct from Lgr5+ cells. Using a kinetic reporter for time-resolved fate mapping and Fgfbp1-CreERT2 lineage tracing, we establish that Fgfbp1+ cells are multi-potent and give rise to Lgr5+ cells, consistent with their ISC function. Fgfbp1+ cells also sustain epithelial regeneration following Lgr5+ cell depletion. We demonstrate that FGFBP1, produced by the upper crypt cells, is an essential factor for crypt proliferation and epithelial homeostasis. Our findings support a model in which tissue regeneration originates from upper crypt Fgfbp1+ cells that generate progeny propagating bi-directionally along the crypt-villus axis and serve as a source of Lgr5+ cells in the crypt base.
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Mucosa Intestinal , Receptores Acoplados a Proteínas G , Receptores Acoplados a Proteínas G/metabolismo , Animales , Ratones , Mucosa Intestinal/metabolismo , Mucosa Intestinal/citología , Células Madre/metabolismo , Células Madre/citología , Linaje de la Célula , Regeneración , Proliferación Celular , Células Epiteliales/metabolismo , Células Epiteliales/citología , Ratones Endogámicos C57BL , HomeostasisRESUMEN
The dendritic cells (DCs) of the immune system function in innate and adaptive responses by directing activity of various effector cells rather than serving as effectors themselves. DCs and closely related myeloid lineages share expression of many surface receptors, presenting a challenge in distinguishing their unique in vivo functions. Recent work has taken advantage of unique transcriptional programs to identify and manipulate murine DCs in vivo. This work has assigned several nonredundant in vivo functions to distinct DC lineages, consisting of plasmacytoid DCs and several subsets of classical DCs that promote different immune effector modules in response to pathogens. In parallel, a correspondence between human and murine DC subsets has emerged, underlying structural similarities for the DC lineages between these species. Recent work has begun to unravel the transcriptional circuitry that controls the development and diversification of DCs from common progenitors in the bone marrow.
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Células de la Médula Ósea/fisiología , Células Dendríticas/fisiología , Regulación de la Expresión Génica , Inmunidad Celular , Animales , Diferenciación Celular , Linaje de la Célula , Perfilación de la Expresión Génica , Redes Reguladoras de Genes , Humanos , Inmunidad Celular/genética , Ratones , Activación TranscripcionalRESUMEN
Cellular lineage histories and their molecular states encode fundamental principles of tissue development and homeostasis. Current lineage-recording mouse models have insufficient barcode diversity and single-cell lineage coverage for profiling tissues composed of millions of cells. Here, we developed DARLIN, an inducible Cas9 barcoding mouse line that utilizes terminal deoxynucleotidyl transferase (TdT) and 30 CRISPR target sites. DARLIN is inducible, generates massive lineage barcodes across tissues, and enables the detection of edited barcodes in â¼70% of profiled single cells. Using DARLIN, we examined fate bias within developing hematopoietic stem cells (HSCs) and revealed unique features of HSC migration. Additionally, we established a protocol for joint transcriptomic and epigenomic single-cell measurements with DARLIN and found that cellular clonal memory is associated with genome-wide DNA methylation rather than gene expression or chromatin accessibility. DARLIN will enable the high-resolution study of lineage relationships and their molecular signatures in diverse tissues and physiological contexts.
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Epigenómica , Transcriptoma , Animales , Ratones , Transcriptoma/genética , Linaje de la Célula/genética , Perfilación de la Expresión Génica , Modelos Animales de Enfermedad , ADNRESUMEN
Embryogenesis necessitates harmonious coordination between embryonic and extraembryonic tissues. Although stem cells of both embryonic and extraembryonic origins have been generated, they are grown in different culture conditions. In this study, utilizing a unified culture condition that activates the FGF, TGF-ß, and WNT pathways, we have successfully derived embryonic stem cells (FTW-ESCs), extraembryonic endoderm stem cells (FTW-XENs), and trophoblast stem cells (FTW-TSCs) from the three foundational tissues of mouse and cynomolgus monkey (Macaca fascicularis) blastocysts. This approach facilitates the co-culture of embryonic and extraembryonic stem cells, revealing a growth inhibition effect exerted by extraembryonic endoderm cells on pluripotent cells, partially through extracellular matrix signaling. Additionally, our cross-species analysis identified both shared and unique transcription factors and pathways regulating FTW-XENs. The embryonic and extraembryonic stem cell co-culture strategy offers promising avenues for developing more faithful embryo models and devising more developmentally pertinent differentiation protocols.
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Embrión de Mamíferos , Células Madre Embrionarias , Animales , Técnicas de Cocultivo , Macaca fascicularis , Células Madre Embrionarias/metabolismo , Diferenciación Celular , Endodermo/metabolismo , Linaje de la CélulaRESUMEN
A generic level of chromatin organization generated by the interplay between cohesin and CTCF suffices to limit promiscuous interactions between regulatory elements, but a lineage-specific chromatin assembly that supersedes these constraints is required to configure the genome to guide gene expression changes that drive faithful lineage progression. Loss-of-function approaches in B cell precursors show that IKAROS assembles interactions across megabase distances in preparation for lymphoid development. Interactions emanating from IKAROS-bound enhancers override CTCF-imposed boundaries to assemble lineage-specific regulatory units built on a backbone of smaller invariant topological domains. Gain of function in epithelial cells confirms IKAROS' ability to reconfigure chromatin architecture at multiple scales. Although the compaction of the Igκ locus required for genome editing represents a function of IKAROS unique to lymphocytes, the more general function to preconfigure the genome to support lineage-specific gene expression and suppress activation of extra-lineage genes provides a paradigm for lineage restriction.
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Cromatina , Genoma , Linfocitos B/metabolismo , Factor de Unión a CCCTC/metabolismo , Cromatina/metabolismo , Ensamble y Desensamble de Cromatina , Humanos , Animales , RatonesRESUMEN
The lymphocyte family has expanded significantly in recent years to include not only the adaptive lymphocytes (T cells, B cells) and NK cells, but also several additional innate lymphoid cell (ILC) types. ILCs lack clonally distributed antigen receptors characteristic of adaptive lymphocytes and instead respond exclusively to signaling via germline-encoded receptors. ILCs resemble T cells more closely than any other leukocyte lineage at the transcriptome level and express many elements of the core T cell transcriptional program, including Notch, Gata3, Tcf7, and Bcl11b. We present our current understanding of the shared and distinct transcriptional regulatory mechanisms involved in the development of adaptive T lymphocytes and closely related ILCs. We discuss the possibility that a core set of transcriptional regulators common to ILCs and T cells establish enhancers that enable implementation of closely aligned effector pathways. Studies of the transcriptional regulation of lymphopoiesis will support the development of novel therapeutic approaches to correct early lymphoid developmental defects and aberrant lymphocyte function.
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Inmunidad Adaptativa/genética , Linaje de la Célula/genética , Regulación de la Expresión Génica , Inmunidad Innata/genética , Linfocitos/inmunología , Linfocitos/metabolismo , Transcripción Genética , Animales , Diferenciación Celular , Humanos , Linfocitos/citología , Células Progenitoras Linfoides/citología , Células Progenitoras Linfoides/metabolismoRESUMEN
Selective breeding of domestic dogs has generated diverse breeds often optimized for performing specialized tasks. Despite the heritability of breed-typical behavioral traits, identification of causal loci has proven challenging due to the complexity of canine population structure. We overcome longstanding difficulties in identifying genetic drivers of canine behavior by developing a framework for understanding relationships between breeds and the behaviors that define them, utilizing genetic data for over 4,000 domestic, semi-feral, and wild canids and behavioral survey data for over 46,000 dogs. We identify ten major canine genetic lineages and their behavioral correlates and show that breed diversification is predominantly driven by non-coding regulatory variation. We determine that lineage-associated genes converge in neurodevelopmental co-expression networks, identifying a sheepdog-associated enrichment for interrelated axon guidance functions. This work presents a scaffold for canine diversification that positions the domestic dog as an unparalleled system for revealing the genetic origins of behavioral diversity.
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Conducta Animal , Perros , Animales , Perros/genética , Perros/fisiología , Variación Genética , Fenotipo , LinajeRESUMEN
In development, lineage segregation is coordinated in time and space. An important example is the mammalian inner cell mass, in which the primitive endoderm (PrE, founder of the yolk sac) physically segregates from the epiblast (EPI, founder of the fetus). While the molecular requirements have been well studied, the physical mechanisms determining spatial segregation between EPI and PrE remain elusive. Here, we investigate the mechanical basis of EPI and PrE sorting. We find that rather than the differences in static cell surface mechanical parameters as in classical sorting models, it is the differences in surface fluctuations that robustly ensure physical lineage sorting. These differential surface fluctuations systematically correlate with differential cellular fluidity, which we propose together constitute a non-equilibrium sorting mechanism for EPI and PrE lineages. By combining experiments and modeling, we identify cell surface dynamics as a key factor orchestrating the correct spatial segregation of the founder embryonic lineages.
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Blastocisto , Embrión de Mamíferos , Endodermo , Animales , Blastocisto/metabolismo , Diferenciación Celular/fisiología , Linaje de la Célula/fisiología , Membrana Celular/metabolismo , Embrión de Mamíferos/metabolismo , Desarrollo Embrionario , Endodermo/metabolismo , Mamíferos , Ratones , Transporte de ProteínasRESUMEN
Incomplete lineage sorting (ILS) makes ancestral genetic polymorphisms persist during rapid speciation events, inducing incongruences between gene trees and species trees. ILS has complicated phylogenetic inference in many lineages, including hominids. However, we lack empirical evidence that ILS leads to incongruent phenotypic variation. Here, we performed phylogenomic analyses to show that the South American monito del monte is the sister lineage of all Australian marsupials, although over 31% of its genome is closer to the Diprotodontia than to other Australian groups due to ILS during ancient radiation. Pervasive conflicting phylogenetic signals across the whole genome are consistent with some of the morphological variation among extant marsupials. We detected hundreds of genes that experienced stochastic fixation during ILS, encoding the same amino acids in non-sister species. Using functional experiments, we confirm how ILS may have directly contributed to hemiplasy in morphological traits that were established during rapid marsupial speciation ca. 60 mya.
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Marsupiales , Animales , Australia , Evolución Molecular , Especiación Genética , Genoma , Marsupiales/genética , Fenotipo , FilogeniaRESUMEN
Although adult pluripotent stem cells (aPSCs) are found in many animal lineages, mechanisms for their formation during embryogenesis are unknown. Here, we leveraged Hofstenia miamia, a regenerative worm that possesses collectively pluripotent aPSCs called neoblasts and produces manipulable embryos. Lineage tracing and functional experiments revealed that one pair of blastomeres gives rise to cells that resemble neoblasts in distribution, behavior, and gene expression. In Hofstenia, aPSCs include transcriptionally distinct subpopulations that express markers associated with differentiated tissues; our data suggest that despite their heterogeneity, aPSCs are derived from one lineage, not from multiple tissue-specific lineages during development. Next, we combined single-cell transcriptome profiling across development with neoblast cell-lineage tracing and identified a molecular trajectory for neoblast formation that includes transcription factors Hes, FoxO, and Tbx. This identification of a cellular mechanism and molecular trajectory for aPSC formation opens the door for in vivo studies of aPSC regulation and evolution.
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Células Madre Adultas , Eucariontes , Células Madre Pluripotentes , Animales , Diferenciación Celular , Linaje de la Célula , Células Madre Pluripotentes/fisiología , Eucariontes/clasificación , Eucariontes/citologíaRESUMEN
Tumor evolution is driven by the progressive acquisition of genetic and epigenetic alterations that enable uncontrolled growth and expansion to neighboring and distal tissues. The study of phylogenetic relationships between cancer cells provides key insights into these processes. Here, we introduced an evolving lineage-tracing system with a single-cell RNA-seq readout into a mouse model of Kras;Trp53(KP)-driven lung adenocarcinoma and tracked tumor evolution from single-transformed cells to metastatic tumors at unprecedented resolution. We found that the loss of the initial, stable alveolar-type2-like state was accompanied by a transient increase in plasticity. This was followed by the adoption of distinct transcriptional programs that enable rapid expansion and, ultimately, clonal sweep of stable subclones capable of metastasizing. Finally, tumors develop through stereotypical evolutionary trajectories, and perturbing additional tumor suppressors accelerates progression by creating novel trajectories. Our study elucidates the hierarchical nature of tumor evolution and, more broadly, enables in-depth studies of tumor progression.
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Neoplasias , Animales , Genes ras , Ratones , Neoplasias/genética , Filogenia , Secuenciación del ExomaRESUMEN
The emergence and spread of SARS-CoV-2 lineage B.1.1.7, first detected in the United Kingdom, has become a global public health concern because of its increased transmissibility. Over 2,500 COVID-19 cases associated with this variant have been detected in the United States (US) since December 2020, but the extent of establishment is relatively unknown. Using travel, genomic, and diagnostic data, we highlight that the primary ports of entry for B.1.1.7 in the US were in New York, California, and Florida. Furthermore, we found evidence for many independent B.1.1.7 establishments starting in early December 2020, followed by interstate spread by the end of the month. Finally, we project that B.1.1.7 will be the dominant lineage in many states by mid- to late March. Thus, genomic surveillance for B.1.1.7 and other variants urgently needs to be enhanced to better inform the public health response.
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Prueba de COVID-19 , COVID-19 , Modelos Biológicos , SARS-CoV-2 , COVID-19/genética , COVID-19/mortalidad , COVID-19/transmisión , Femenino , Humanos , Masculino , SARS-CoV-2/genética , SARS-CoV-2/metabolismo , SARS-CoV-2/patogenicidad , Estados Unidos/epidemiologíaRESUMEN
The independent emergence late in 2020 of the B.1.1.7, B.1.351, and P.1 lineages of SARS-CoV-2 prompted renewed concerns about the evolutionary capacity of this virus to overcome public health interventions and rising population immunity. Here, by examining patterns of synonymous and non-synonymous mutations that have accumulated in SARS-CoV-2 genomes since the pandemic began, we find that the emergence of these three "501Y lineages" coincided with a major global shift in the selective forces acting on various SARS-CoV-2 genes. Following their emergence, the adaptive evolution of 501Y lineage viruses has involved repeated selectively favored convergent mutations at 35 genome sites, mutations we refer to as the 501Y meta-signature. The ongoing convergence of viruses in many other lineages on this meta-signature suggests that it includes multiple mutation combinations capable of promoting the persistence of diverse SARS-CoV-2 lineages in the face of mounting host immune recognition.
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COVID-19/epidemiología , Evolución Molecular , Mutación , Pandemias , SARS-CoV-2/genética , Secuencia de Aminoácidos/genética , COVID-19/inmunología , COVID-19/transmisión , COVID-19/virología , Codón/genética , Genes Virales , Flujo Genético , Adaptación al Huésped/genética , Humanos , Evasión Inmune , Filogenia , Salud PúblicaRESUMEN
In vivo cell fate conversions have emerged as potential regeneration-based therapeutics for injury and disease. Recent studies reported that ectopic expression or knockdown of certain factors can convert resident astrocytes into functional neurons with high efficiency, region specificity, and precise connectivity. However, using stringent lineage tracing in the mouse brain, we show that the presumed astrocyte-converted neurons are actually endogenous neurons. AAV-mediated co-expression of NEUROD1 and a reporter specifically and efficiently induces reporter-labeled neurons. However, these neurons cannot be traced retrospectively to quiescent or reactive astrocytes using lineage-mapping strategies. Instead, through a retrograde labeling approach, our results reveal that endogenous neurons are the source for these viral-reporter-labeled neurons. Similarly, despite efficient knockdown of PTBP1 in vivo, genetically traced resident astrocytes were not converted into neurons. Together, our results highlight the requirement of lineage-tracing strategies, which should be broadly applied to studies of cell fate conversions in vivo.
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Astrocitos/citología , Diferenciación Celular , Linaje de la Célula , Neuronas/citología , Animales , Astrocitos/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Encéfalo/patología , Lesiones Encefálicas/patología , Línea Celular Tumoral , Reprogramación Celular , Dependovirus/metabolismo , Regulación hacia Abajo , Regulación de la Expresión Génica , Genes Reporteros , Proteína Ácida Fibrilar de la Glía/genética , Ribonucleoproteínas Nucleares Heterogéneas/metabolismo , Proteínas de Homeodominio/metabolismo , Humanos , Integrasas/metabolismo , Ratones Endogámicos C57BL , Ratones Transgénicos , Neuronas/metabolismo , Proteína de Unión al Tracto de Polipirimidina/metabolismo , Regiones Promotoras Genéticas/genética , Factores de Transcripción/metabolismoRESUMEN
Tracing the lineage history of cells is key to answering diverse and fundamental questions in biology. Coupling of cell ancestry information with other molecular readouts represents an important goal in the field. Here, we describe the CRISPR array repair lineage tracing (CARLIN) mouse line and corresponding analysis tools that can be used to simultaneously interrogate the lineage and transcriptomic information of single cells in vivo. This model exploits CRISPR technology to generate up to 44,000 transcribed barcodes in an inducible fashion at any point during development or adulthood, is compatible with sequential barcoding, and is fully genetically defined. We have used CARLIN to identify intrinsic biases in the activity of fetal liver hematopoietic stem cell (HSC) clones and to uncover a previously unappreciated clonal bottleneck in the response of HSCs to injury. CARLIN also allows the unbiased identification of transcriptional signatures associated with HSC activity without cell sorting.