RESUMEN
ATRX is an X-linked gene of the SWI/SNF family, mutations in which cause syndromal mental retardation and downregulation of α-globin expression. Here we show that ATRX binds to tandem repeat (TR) sequences in both telomeres and euchromatin. Genes associated with these TRs can be dysregulated when ATRX is mutated, and the change in expression is determined by the size of the TR, producing skewed allelic expression. This reveals the characteristics of the affected genes, explains the variable phenotypes seen with identical ATRX mutations, and illustrates a new mechanism underlying variable penetrance. Many of the TRs are G rich and predicted to form non-B DNA structures (including G-quadruplex) in vivo. We show that ATRX binds G-quadruplex structures in vitro, suggesting a mechanism by which ATRX may play a role in various nuclear processes and how this is perturbed when ATRX is mutated.
Asunto(s)
ADN Helicasas/metabolismo , Proteínas Nucleares/metabolismo , Animales , Células Cultivadas , Inmunoprecipitación de Cromatina , Cromosomas de los Mamíferos/metabolismo , Islas de CpG , ADN Helicasas/genética , ADN Ribosómico/metabolismo , G-Cuádruplex , Expresión Génica , Estudio de Asociación del Genoma Completo , Histonas/metabolismo , Humanos , Ratones , Repeticiones de Minisatélite , Mutación , Proteínas Nucleares/genética , Telómero/metabolismo , Proteína Nuclear Ligada al Cromosoma XRESUMEN
The incorporation of histone H3 variants has been implicated in the epigenetic memory of cellular state. Using genome editing with zinc-finger nucleases to tag endogenous H3.3, we report genome-wide profiles of H3 variants in mammalian embryonic stem cells and neuronal precursor cells. Genome-wide patterns of H3.3 are dependent on amino acid sequence and change with cellular differentiation at developmentally regulated loci. The H3.3 chaperone Hira is required for H3.3 enrichment at active and repressed genes. Strikingly, Hira is not essential for localization of H3.3 at telomeres and many transcription factor binding sites. Immunoaffinity purification and mass spectrometry reveal that the proteins Atrx and Daxx associate with H3.3 in a Hira-independent manner. Atrx is required for Hira-independent localization of H3.3 at telomeres and for the repression of telomeric RNA. Our data demonstrate that multiple and distinct factors are responsible for H3.3 localization at specific genomic locations in mammalian cells.
Asunto(s)
Histonas/análisis , Telómero/química , Animales , Sitios de Unión , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Células Madre Embrionarias/metabolismo , Genoma , Chaperonas de Histonas/genética , Chaperonas de Histonas/metabolismo , Histonas/genética , Histonas/metabolismo , Ratones , Ratones Endogámicos C57BL , Telómero/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Sitio de Iniciación de la TranscripciónRESUMEN
A substantial amount of organismal complexity is thought to be encoded by enhancers which specify the location, timing, and levels of gene expression. In mammals there are more enhancers than promoters which are distributed both between and within genes. Here we show that activated, intragenic enhancers frequently act as alternative tissue-specific promoters producing a class of abundant, spliced, multiexonic poly(A)(+) RNAs (meRNAs) which reflect the host gene's structure. meRNAs make a substantial and unanticipated contribution to the complexity of the transcriptome, appearing as alternative isoforms of the host gene. The low protein-coding potential of meRNAs suggests that many meRNAs may be byproducts of enhancer activation or underlie as-yet-unidentified RNA-encoded functions. Distinguishing between meRNAs and mRNAs will transform our interpretation of dynamic changes in transcription both at the level of individual genes and of the genome as a whole.
Asunto(s)
Elementos de Facilitación Genéticos/fisiología , Regulación de la Expresión Génica , Regiones Promotoras Genéticas/fisiología , Animales , Células Cultivadas , Células Eritroides , Ratones , Poli A , ARN/química , ARN/fisiología , Isoformas de ARN/química , ARN Mensajero/química , ARN Mensajero/fisiología , TranscriptomaRESUMEN
The human genome contains â¼30,000 CpG islands (CGIs). While CGIs associated with promoters nearly always remain unmethylated, many of the â¼9,000 CGIs lying within gene bodies become methylated during development and differentiation. Both promoter and intragenic CGIs may also become abnormally methylated as a result of genome rearrangements and in malignancy. The epigenetic mechanisms by which some CGIs become methylated but others, in the same cell, remain unmethylated in these situations are poorly understood. Analyzing specific loci and using a genome-wide analysis, we show that transcription running across CGIs, associated with specific chromatin modifications, is required for DNA methyltransferase 3B (DNMT3B)-mediated DNA methylation of many naturally occurring intragenic CGIs. Importantly, we also show that a subgroup of intragenic CGIs is not sensitive to this process of transcription-mediated methylation and that this correlates with their individual intrinsic capacity to initiate transcription in vivo. We propose a general model of how transcription could act as a primary determinant of the patterns of CGI methylation in normal development and differentiation, and in human disease.
Asunto(s)
Diferenciación Celular/genética , Islas de CpG/genética , Metilación de ADN/genética , Transcripción Genética/genética , Animales , Línea Celular , Epigénesis Genética/genética , Genoma Humano/genética , Humanos , Ratones , Regiones Promotoras Genéticas/genética , Análisis de Secuencia de ADN/métodosRESUMEN
Remote distal enhancers may be located tens or thousands of kilobases away from their promoters. How they control gene expression is still poorly understood. Here, we analyze the influence of a remote enhancer on the balance between repression (Polycomb-PcG) and activation (Trithorax-TrxG) of a developmentally regulated gene associated with a CpG island. We reveal its essential, nonredundant role in clearing the PcG complex and H3K27me3 from the CpG island. In the absence of the enhancer, the H3K27me3 demethylase (JMJD3) is not recruited to the CpG island. We propose a new role of long-range regulatory elements in removing repressive PcG complexes.
Asunto(s)
Elementos de Facilitación Genéticos/fisiología , Proteínas Represoras/metabolismo , Animales , Línea Celular , Células Cultivadas , Cromatina/metabolismo , Inmunoprecipitación de Cromatina , Islas de CpG , Eritropoyesis/fisiología , Regulación de la Expresión Génica , Humanos , Proteínas del Grupo Polycomb , Unión Proteica , Proteína Metiltransferasas/metabolismoRESUMEN
The role of DNA sequence in determining chromatin state is incompletely understood. We have previously demonstrated that large chromosomal segments from human cells recapitulate their native chromatin state in mouse cells, but the relative contribution of local sequences versus their genomic context remains unknown. In this study, we compare orthologous chromosomal regions for which the human locus establishes prominent sites of Polycomb complex recruitment in pluripotent stem cells, whereas the corresponding mouse locus does not. Using recombination-mediated cassette exchange at the mouse locus, we establish the primacy of local sequences in the encoding of chromatin state. We show that the signal for chromatin bivalency is redundantly encoded across a bivalent domain and that this reflects competition between Polycomb complex recruitment and transcriptional activation. Furthermore, our results suggest that a high density of unmethylated CpG dinucleotides is sufficient for vertebrate Polycomb recruitment. This model is supported by analysis of DNA methyltransferase-deficient embryonic stem cells.
Asunto(s)
Islas de CpG/fisiología , Regulación de la Expresión Génica/genética , Proteínas Represoras/metabolismo , Globinas alfa/genética , Animales , Células Cultivadas/metabolismo , Cromatina/genética , Mapeo Cromosómico , Cromosomas Humanos Par 16 , Metilación de ADN , ADN Recombinante/genética , Células Madre Embrionarias/metabolismo , Humanos , Ratones , Ratones Transgénicos , Células Madre Pluripotentes/metabolismo , Proteínas del Grupo Polycomb , Recombinación Genética , Secuencias Reguladoras de Ácidos Nucleicos , Especificidad de la Especie , Transcripción GenéticaRESUMEN
Chimeric antigen receptor (CAR) T-cell therapy for multiple myeloma targeting B-cell maturation antigen (BCMA) induces high overall response rates. However, relapse still occurs and novel strategies for targeting multiple myeloma cells using CAR T-cell therapy are needed. SLAMF7 (also known as CS1) and CD38 on tumor plasma cells represent potential alternative targets for CAR T-cell therapy in multiple myeloma, but their expression on activated T cells and other hematopoietic cells raises concerns about the efficacy and safety of such treatments. Here, we used CRISPR/Cas9 deletion of the CD38 gene in T cells and developed DCAR, a double CAR system targeting CD38 and CS1 through activation and costimulation receptors, respectively. Inactivation of CD38 enhanced the anti-multiple myeloma activity of DCAR T in vitro. Edited DCAR T cells showed strong in vitro and in vivo responses specifically against target cells expressing both CD38 and CS1. Furthermore, we provide evidence that, unlike anti-CD38 CAR T-cell therapy, which elicited a rapid immune reaction against hematopoietic cells in a humanized mouse model, DCAR T cells showed no signs of toxicity. Thus, DCAR T cells could provide a safe and efficient alternative to anti-BCMA CAR T-cell therapy to treat patients with multiple myeloma.
Asunto(s)
Mieloma Múltiple , Receptores Quiméricos de Antígenos , Animales , Ratones , Humanos , Mieloma Múltiple/patología , Receptores Quiméricos de Antígenos/metabolismo , Receptores de Antígenos de Linfocitos T , Recurrencia Local de Neoplasia , Linfocitos T , Inmunoterapia Adoptiva , Familia de Moléculas Señalizadoras de la Activación LinfocitariaRESUMEN
Although mutations causing monogenic disorders most frequently lie within the affected gene, sequence variation in complex disorders is more commonly found in noncoding regions. Furthermore, recent genome- wide studies have shown that common DNA sequence variants in noncoding regions are associated with "normal" variation in gene expression resulting in cell-specific and/or allele-specific differences. The mechanism by which such sequence variation causes changes in gene expression is largely unknown. We have addressed this by studying natural variation in the binding of key transcription factors (TFs) in the well-defined, purified cell system of erythropoiesis. We have shown that common polymorphisms frequently directly perturb the binding sites of key TFs, and detailed analysis shows how this causes considerable (~10-fold) changes in expression from a single allele in a tissue-specific manner. We also show how a SNP, located at some distance from the recognized TF binding site, may affect the recruitment of a large multiprotein complex and alter the associated chromatin modification of the variant regulatory element. This study illustrates the principles by which common sequence variation may cause changes in tissue-specific gene expression, and suggests that such variation may underlie an individual's propensity to develop complex human genetic diseases.
Asunto(s)
Células Eritroides/metabolismo , Expresión Génica , Péptidos y Proteínas de Señalización Intracelular/genética , Nucleósido Difosfato Quinasa D/genética , Nucleósido Difosfato Quinasa D/metabolismo , Polimorfismo de Nucleótido Simple , Factores de Transcripción/metabolismo , Secuencia de Bases , Sitios de Unión/genética , Variación Genética , Estudio de Asociación del Genoma Completo , Humanos , Péptidos y Proteínas de Señalización Intracelular/química , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Datos de Secuencia Molecular , Unión Proteica , Secuencias Reguladoras de Ácidos NucleicosRESUMEN
Inherited mutations of specific genes have elucidated the normal roles of the proteins they encode by relating specific mutations to particular phenotypes. But many potentially informative mutations in such genes are lethal early in development. Consequently, inherited mutations may not reflect all the functional roles of such proteins. Acquired, somatic defects should reflect a wider spectrum of mutations because they are not prone to negative selection in development. It has been difficult to identify such mutations so far, but microarray analysis provides a new opportunity to do so. Using this approach, we have shown that in individuals with myelodysplasia associated with alpha-thalassemia (ATMDS), somatic mutations of the gene encoding the chromatin remodeling factor ATRX cause an unexpectedly severe hematological phenotype compared with the wide spectrum of inherited mutations affecting this gene. These findings cast new light on this pleiotropic cofactor, which appears to be an essential component rather than a mere facilitator of globin gene expression.
Asunto(s)
ADN Helicasas/genética , Mutación , Síndromes Mielodisplásicos/genética , Proteínas Nucleares/genética , Talasemia alfa/genética , Secuencia de Bases , Células de la Médula Ósea/metabolismo , Estudios de Casos y Controles , ADN/genética , Femenino , Expresión Génica , Perfilación de la Expresión Génica , Globinas/genética , Humanos , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos , Fenotipo , Proteína Nuclear Ligada al Cromosoma XRESUMEN
Nearly all human genetic disorders result from a limited repertoire of mutations in an associated gene or its regulatory elements. We recently described an individual with an inherited form of anemia (alpha-thalassemia) who has a deletion that results in a truncated, widely expressed gene (LUC7L) becoming juxtaposed to a structurally normal alpha-globin gene (HBA2). Although it retains all of its local and remote cis-regulatory elements, expression of HBA2 is silenced and its CpG island becomes completely methylated early during development. Here we show that in the affected individual, in a transgenic model and in differentiating embryonic stem cells, transcription of antisense RNA mediates silencing and methylation of the associated CpG island. These findings identify a new mechanism underlying human genetic disease.
Asunto(s)
Metilación de ADN , Silenciador del Gen , ARN sin Sentido/genética , Talasemia alfa/genética , Animales , Secuencia de Bases , Línea Celular , Cromosomas Humanos Par 16/genética , Islas de CpG , ADN/genética , Globinas/genética , Humanos , Ratones , Ratones Transgénicos , Modelos Genéticos , Regiones Promotoras Genéticas , Transcripción GenéticaRESUMEN
The understanding of nonequilibrium dynamics in many-body quantum systems is a fundamental issue in statistical physics. Experiments that probe universal properties of these systems can address such foundational questions. In this study, we report the measurement of universal dynamics triggered by a quench from the superfluid to normal phase across the Berezinskii-Kosterlitz-Thouless transition in a two-dimensional (2D) Bose gas. We reduced the density by splitting the 2D gas in two, realizing a quench across the critical point. The subsequent relaxation dynamics were probed with matter-wave interferometry to measure the local phase fluctuations. We show that the time evolution of both the phase correlation function and vortex density obeys universal scaling laws. This conclusion is supported by classical-field simulations and interpreted by means of real-time renormalization group theory.
RESUMEN
The developmental cartography of human lymphopoiesis remains incompletely understood. Here, we establish a multimodal map demonstrating that lymphoid specification follows independent direct or stepwise hierarchic routes converging toward the emergence of newly characterized CD117lo multi-lymphoid progenitors (MLPs) that undergo a proliferation arrest before entering the CD127- (NK/ILC/T) or CD127+ (B) lymphoid pathways. While the differentiation of CD127- early lymphoid progenitors is mainly driven by Flt3 signaling, emergence of their CD127+ counterparts is regulated cell-intrinsically and depends exclusively on the divisional history of their upstream precursors, including hematopoietic stem cells. Further, transcriptional mapping of differentiation trajectories reveals that whereas myeloid granulomonocytic lineages follow continuous differentiation pathways, lymphoid trajectories are intrinsically discontinuous and characterized by sequential waves of cell proliferation allowing pre-commitment amplification of lymphoid progenitor pools. Besides identifying new lymphoid specification pathways and regulatory checkpoints, our results demonstrate that NK/ILC/T and B lineages are under fundamentally distinct modes of regulation. (149 words).
RESUMEN
Changes in lymphocyte production patterns occurring across human ontogeny remain poorly defined. In this study, we demonstrate that human lymphopoiesis is supported by three waves of embryonic, fetal, and postnatal multi-lymphoid progenitors (MLPs) differing in CD7 and CD10 expression and their output of CD127-/+ early lymphoid progenitors (ELPs). In addition, our results reveal that, like the fetal-to-adult switch in erythropoiesis, transition to postnatal life coincides with a shift from multilineage to B lineage-biased lymphopoiesis and an increase in production of CD127+ ELPs, which persists until puberty. A further developmental transition is observed in elderly individuals whereby B cell differentiation bypasses the CD127+ compartment and branches directly from CD10+ MLPs. Functional analyses indicate that these changes are determined at the level of hematopoietic stem cells. These findings provide insights for understanding identity and function of human MLPs and the establishment and maintenance of adaptative immunity.
Asunto(s)
Células Madre Hematopoyéticas , Linfopoyesis , Adulto , Humanos , Anciano , Diferenciación Celular , Linaje de la Célula , HematopoyesisRESUMEN
It is well established that all of the cis-acting sequences required for fully regulated human alpha-globin expression are contained within a region of approximately 120 kb of conserved synteny. Here, we show that activation of this cluster in erythroid cells dramatically affects expression of apparently unrelated and noncontiguous genes in the 500 kb surrounding this domain, including a gene (NME4) located 300 kb from the alpha-globin cluster. Changes in NME4 expression are mediated by physical cis-interactions between this gene and the alpha-globin regulatory elements. Polymorphic structural variation within the globin cluster, altering the number of alpha-globin genes, affects the pattern of NME4 expression by altering the competition for the shared alpha-globin regulatory elements. These findings challenge the concept that the genome is organized into discrete, insulated regulatory domains. In addition, this work has important implications for our understanding of genome evolution, the interpretation of genome-wide expression, expression-quantitative trait loci, and copy number variant analyses.
Asunto(s)
Expresión Génica , Polimorfismo de Nucleótido Simple , Regiones Promotoras Genéticas , Cromosomas Humanos Par 16 , Humanos , Secuencias Reguladoras de Ácidos Nucleicos , Telómero , Globinas alfa/genéticaRESUMEN
Chimeric antigen receptor T cells (CAR-T) have provided promising results in multiple myeloma (MM). However, many patients still relapse, pointing toward the need of improving this therapy. Here, we analyzed peripheral blood T cells from MM patients at different stages of the disease and investigated their phenotype and capacity to generate functional CAR-T directed against CS1 or B Cell Maturation antigen. We found a decrease in naive T cells and elevated frequencies of exhaustion markers in T cells from treated MM patients. Interestingly, individuals treated with daratumumab display elevated ratios of central memory T cells. CAR-T derived from patients at relapse show reduced in vitro expansion and cytotoxic capacities in response to MM cells compared to those produced at diagnosis. Of note, CAR-T from daratumumab treated patients display intermediate defects. Reduced anti-myeloma activity of CAR T cells from treated patients was also observed in a mouse model. Our findings suggest that T cell defects in MM patients, specifically during relapse, have a major impact on their capacity to generate efficient therapeutic CAR-T. Selecting naive or central memory T cell subsets to generate therapeutic T cells could improve the CAR-T therapy for MM.
RESUMEN
Erythropoiesis requires a combination of ubiquitous and tissue-specific transcription factors (TFs). Here, through DNA affinity purification followed by mass spectrometry, we have identified the widely expressed protein MAZ (Myc-associated zinc finger) as a TF that binds to the promoter of the erythroid-specific human α-globin gene. Genome-wide mapping in primary human erythroid cells revealed that MAZ also occupies active promoters as well as GATA1-bound enhancer elements of key erythroid genes. Consistent with an important role during erythropoiesis, knockdown of MAZ reduces α-globin expression in K562 cells and impairs differentiation in primary human erythroid cells. Genetic variants in the MAZ locus are associated with changes in clinically important human erythroid traits. Taken together, these findings reveal the zinc-finger TF MAZ to be a previously unrecognized regulator of the erythroid differentiation program.
Asunto(s)
Proteínas de Unión al ADN , Eritropoyesis , Factores de Transcripción , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Células Eritroides/metabolismo , Eritropoyesis/genética , Regulación de la Expresión Génica , Humanos , Células K562 , Regiones Promotoras Genéticas , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Multiple myeloma (MM) is a currently incurable malignancy of antibody-secreting plasma cells. Long non-coding RNAs (lncRNAs) have been recognised as an important class of regulatory molecules which are increasingly implicated in tumorigenesis. While recent studies have demonstrated changes in expression of lncRNAs in MM, the functional significance and molecular pathways downstream of these changes remain poorly characterised. In this study, we have performed CRISPR-mediated deletion of the locus encoding the lncRNA Colorectal Neoplasia Differentially Expressed (CRNDE), a known oncogenic lncRNA that is overexpressed in plasma cells of MM patients and is a marker of poor prognosis. We found that CRISPR-mediated deletion of the CRNDE locus in MM cells decreases proliferation and adhesion properties, increases sensitivity to Dexamethasone and reduces tumour growth in an in vivo xenograft model. Transcriptomic profiling in CRNDE-deleted MM cells demonstrated that CRNDE activates expression of a number of genes previously implicated in the aetiology of MM, including IL6R. We further demonstrate that deletion of the CRNDE locus diminishes IL6 signalling and proliferative responses in MM cells. Altogether this study reveals the IL6 signalling pathway as a novel mechanism by which CRNDE impacts upon MM cell growth and disease progression.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Regulación Neoplásica de la Expresión Génica , Interleucina-6/metabolismo , Mieloma Múltiple/patología , ARN Largo no Codificante/genética , Animales , Apoptosis , Biomarcadores de Tumor/genética , Proliferación Celular , Humanos , Interleucina-6/genética , Ratones , Ratones Endogámicos NOD , Ratones SCID , Mieloma Múltiple/genética , Mieloma Múltiple/metabolismo , Pronóstico , Tasa de Supervivencia , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
We have characterized a newly identified 16.6 kb deletion which removes a significant proportion of the human alpha-globin cluster including the psizeta1, alpha(D), psialpha1 and alpha2-globin genes but leaves the duplicated alpha1 gene intact. This complicated rearrangement results from a combination of slippage and strand switching at sites of microhomology during replication. Functional analysis shows that expression of the remaining alpha1 gene is increased, rather than down-regulated by this deletion. This could be related to its proximity to the remote upstream alpha-globin regulatory elements or reduced competition for these elements in the absence of the dominant alpha2-globin gene. The finding of a very mild phenotype associated with such an extensive deletion in the alpha-globin cluster implies that much of the DNA removed by the deletion is likely to be functionally unimportant. These findings suggest that other than the upstream regulatory elements and promoter proximal elements there are unlikely to be additional positive cis-acting sequences in the alpha-globin cluster.
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Replicación del ADN , Eliminación de Gen , Globinas/genética , Familia de Multigenes , Adulto , Células Cultivadas , Niño , Cromatina/genética , Cromatina/metabolismo , Cromosomas Humanos Par 16/genética , Femenino , Expresión Génica , Globinas/metabolismo , Humanos , Masculino , Fenotipo , ARN Polimerasa II/metabolismoRESUMEN
Although much is known about globin gene activation in erythroid cells, relatively little is known about how these genes are silenced in nonerythroid tissues. Here we show that the human alpha- and beta-globin genes are silenced by fundamentally different mechanisms. The alpha-genes, which are surrounded by widely expressed genes in a gene dense region of the genome, are silenced very early in development via recruitment of the Polycomb (PcG) complex. By contrast, the beta-globin genes, which lie in a relatively gene-poor chromosomal region, are not bound by this complex in nonerythroid cells. The PcG complex seems to be recruited to the alpha-cluster by sequences within the CpG islands associated with their promoters; the beta-globin promoters do not lie within such islands. Chromatin associated with the alpha-globin cluster is modified by histone methylation (H3K27me3), and silencing in vivo is mediated by the localized activity of histone deacetylases (HDACs). The repressive (PcG/HDAC) machinery is removed as hematopoietic progenitors differentiate to form erythroid cells. The alpha- and beta-globin genes thus illustrate important, contrasting mechanisms by which cell-specific hematopoietic genes (and tissue-specific genes in general) may be silenced.
Asunto(s)
Silenciador del Gen , Globinas/genética , Proteínas Represoras/metabolismo , Secuencia de Bases , Línea Celular , Células Cultivadas , Proteínas de Unión al ADN/antagonistas & inhibidores , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Células Madre Embrionarias/metabolismo , Proteína Potenciadora del Homólogo Zeste 2 , Células HeLa , Histona Desacetilasas/metabolismo , Humanos , Células Madre Pluripotentes/metabolismo , Complejo Represivo Polycomb 2 , Proteínas del Grupo Polycomb , Interferencia de ARN , ARN Interferente Pequeño/genética , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
ATRX is an X-encoded member of the SNF2 family of ATPase/helicase proteins thought to regulate gene expression by modifying chromatin at target loci. Mutations in ATRX provided the first example of a human genetic disease associated with defects in such proteins. To better understand the role of ATRX in development and the associated abnormalities in the ATR-X (alpha thalassemia mental retardation, X-linked) syndrome, we conditionally inactivated the homolog in mice, Atrx, at the 8- to 16-cell stage of development. The protein, Atrx, was ubiquitously expressed, and male embryos null for Atrx implanted and gastrulated normally but did not survive beyond 9.5 days postcoitus due to a defect in formation of the extraembryonic trophoblast, one of the first terminally differentiated lineages in the developing embryo. Carrier female mice that inherit a maternal null allele should be affected, since the paternal X chromosome is normally inactivated in extraembryonic tissues. Surprisingly, however, some carrier females established a normal placenta and appeared to escape the usual pattern of imprinted X-inactivation in these tissues. Together these findings demonstrate an unexpected, specific, and essential role for Atrx in the development of the murine trophoblast and present an example of escape from imprinted X chromosome inactivation.