RESUMO
CD4(+) follicular helper T cells (T(FH) cells) are essential for germinal center (GC) responses and long-lived antibody responses. Here we report that naive CD4(+) T cells deficient in the transcription factor Foxp1 'preferentially' differentiated into T(FH) cells, which resulted in substantially enhanced GC and antibody responses. We found that Foxp1 used both constitutive Foxp1A and Foxp1D induced by stimulation of the T cell antigen receptor (TCR) to inhibit the generation of T(FH) cells. Mechanistically, Foxp1 directly and negatively regulated interleukin 21 (IL-21); Foxp1 also dampened expression of the costimulatory molecule ICOS and its downstream signaling at early stages of T cell activation, which rendered Foxp1-deficient CD4(+) T cells partially resistant to blockade of the ICOS ligand (ICOSL) during T(FH) cell development. Our findings demonstrate that Foxp1 is a critical negative regulator of T(FH) cell differentiation.
Assuntos
Diferenciação Celular , Fatores de Transcrição Forkhead/fisiologia , Proteínas Repressoras/fisiologia , Linfócitos T Auxiliares-Indutores/citologia , Animais , Linfócitos T CD4-Positivos/imunologia , Proteína Coestimuladora de Linfócitos T Induzíveis/genética , Interleucinas/genética , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Antígenos de Linfócitos T/fisiologiaRESUMO
Mutations in the transcription factor Forkhead box p1 (FOXP1) are causative for neurodevelopmental disorders such as autism. However, the function of FOXP1 within the brain remains largely uncharacterized. Here, we identify the gene expression program regulated by FoxP1 in both human neural cells and patient-relevant heterozygous Foxp1 mouse brains. We demonstrate a role for FoxP1 in the transcriptional regulation of autism-related pathways as well as genes involved in neuronal activity. We show that Foxp1 regulates the excitability of striatal medium spiny neurons and that reduction of Foxp1 correlates with defects in ultrasonic vocalizations. Finally, we demonstrate that FoxP1 has an evolutionarily conserved role in regulating pathways involved in striatal neuron identity through gene expression studies in human neural progenitors with altered FOXP1 levels. These data support an integral role for FoxP1 in regulating signaling pathways vulnerable in autism and the specific regulation of striatal pathways important for vocal communication.
Assuntos
Transtorno do Espectro Autista/fisiopatologia , Corpo Estriado/fisiopatologia , Fatores de Transcrição Forkhead/metabolismo , Proteínas Repressoras/metabolismo , Transdução de Sinais/genética , Animais , Transtorno do Espectro Autista/genética , Células Cultivadas , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/genética , Regulação da Expressão Gênica/genética , Haploinsuficiência , Hipocampo/fisiopatologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Mutação , Neurônios/patologia , Proteínas Repressoras/genética , Comportamento Verbal/fisiologiaRESUMO
STYK1/NOK functions in a ligand independent and constitutive fashion to provoke tumor formation and to be up-regulated in many types of cancer cells. However, how STYK1/NOK functions at the whole animal level is completely unknown. Here, we found that STYK1/NOK-transgenic (tg) mice spontaneously developed immunosuppressive B-CLL-like disease with generally shorter life spans. The phenotype of STYK1/NOK-induced B-CLL was typically heterogeneous, and most often, presented lymphadenectasis accompanied with hepatomegaly and/or splenomegaly. STYK1/NOK-tg mice also suffered reduced immune responses. The expanded CD5+CD19+ (B1) lymphocyte pool was detected within peripheral lymphoid organs. Analysis on GEO profile revealed that expression of STYK1/NOK were significantly up-regulated in primary human B-CLL. Inoculation of blood cells from sick STYK1/NOK-tg mice into immune-deficient recipients recaptured the B1 malignant phenotype. Our study demonstrated that STYK1/NOK transgenic mouse may serve as a useful model system for the developments of novel diagnosis and treatment of B-CLL.
Assuntos
Leucemia Linfocítica Crônica de Células B , Receptores Proteína Tirosina Quinases , Animais , Humanos , Leucemia Linfocítica Crônica de Células B/genética , Camundongos , Camundongos Transgênicos , Receptores Proteína Tirosina Quinases/metabolismoRESUMO
Despite their origin from the inner cell mass, embryonic stem (ES) cells undergo differentiation to the trophectoderm (TE) lineage by repression of the ES cell master regulator Oct4 or activation of the TE master regulator Caudal-type homeobox 2 (Cdx2). In contrast to the in-depth studies of ES cell self-renewal and pluripotency, few TE-specific regulators have been identified, thereby limiting our understanding of mechanisms underlying the first cell fate decision. Here we show that up-regulation and nuclear entry of AT-rich interactive domain 3a (Arid3a) drives TE-like transcriptional programs in ES cells, maintains trophoblast stem (TS) cell self-renewal, and promotes further trophoblastic differentiation both upstream and independent of Cdx2. Accordingly, Arid3a(-/-) mouse post-implantation placental development is severely impaired, resulting in early embryonic death. We provide evidence that Arid3a directly activates TE-specific and trophoblast lineage-specific genes while directly repressing pluripotency genes via differential regulation of epigenetic acetylation or deacetylation. Our results identify Arid3a as a critical regulator of TE and placental development through execution of the commitment and differentiation phases of the first cell fate decision.
Assuntos
Diferenciação Celular/genética , Proteínas de Ligação a DNA/metabolismo , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Fatores de Transcrição/metabolismo , Transporte Ativo do Núcleo Celular , Animais , Linhagem da Célula/genética , Proteínas de Ligação a DNA/genética , Feminino , Células HEK293 , Humanos , Camundongos , Fator 3 de Transcrição de Octâmero/metabolismo , Placentação , Gravidez , Fatores de Transcrição/genéticaRESUMO
Fundamental human traits, such as language and bipedalism, are associated with a range of anatomical adaptations in craniofacial shaping and skeletal remodeling. However, it is unclear how such morphological features arose during hominin evolution. FOXP2 is a brain-expressed transcription factor implicated in a rare disorder involving speech apraxia and language impairments. Analysis of its evolutionary history suggests that this gene may have contributed to the emergence of proficient spoken language. In the present study, through analyses of skeleton-specific knockout mice, we identified roles of Foxp2 in skull shaping and bone remodeling. Selective ablation of Foxp2 in cartilage disrupted pup vocalizations in a similar way to that of global Foxp2 mutants, which may be due to pleiotropic effects on craniofacial morphogenesis. Our findings also indicate that Foxp2 helps to regulate strength and length of hind limbs and maintenance of joint cartilage and intervertebral discs, which are all anatomical features that are susceptible to adaptations for bipedal locomotion. In light of the known roles of Foxp2 in brain circuits that are important for motor skills and spoken language, we suggest that this gene may have been well placed to contribute to coevolution of neural and anatomical adaptations related to speech and bipedal locomotion.
Assuntos
Remodelação Óssea/genética , Fatores de Transcrição Forkhead , Locomoção/genética , Mutação , Proteínas Repressoras , Crânio/metabolismo , Vocalização Animal , Animais , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Membro Posterior/metabolismo , Humanos , Camundongos , Camundongos Knockout , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismoRESUMO
Smyd1, a muscle-specific histone methyltransferase, has established roles in skeletal and cardiac muscle development, but its role in the adult heart remains poorly understood. Our prior work demonstrated that cardiac-specific deletion of Smyd1 in adult mice (Smyd1-KO) leads to hypertrophy and heart failure. Here we show that down-regulation of mitochondrial energetics is an early event in these Smyd1-KO mice preceding the onset of structural abnormalities. This early impairment of mitochondrial energetics in Smyd1-KO mice is associated with a significant reduction in gene and protein expression of PGC-1α, PPARα, and RXRα, the master regulators of cardiac energetics. The effect of Smyd1 on PGC-1α was recapitulated in primary cultured rat ventricular myocytes, in which acute siRNA-mediated silencing of Smyd1 resulted in a greater than twofold decrease in PGC-1α expression without affecting that of PPARα or RXRα. In addition, enrichment of histone H3 lysine 4 trimethylation (a mark of gene activation) at the PGC-1α locus was markedly reduced in Smyd1-KO mice, and Smyd1-induced transcriptional activation of PGC-1α was confirmed by luciferase reporter assays. Functional confirmation of Smyd1's involvement showed an increase in mitochondrial respiration capacity induced by overexpression of Smyd1, which was abolished by siRNA-mediated PGC-1α knockdown. Conversely, overexpression of PGC-1α rescued transcript expression and mitochondrial respiration caused by silencing Smyd1 in cardiomyocytes. These findings provide functional evidence for a role of Smyd1, or any member of the Smyd family, in regulating cardiac energetics in the adult heart, which is mediated, at least in part, via modulating PGC-1α.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Metabolismo Energético/fisiologia , Histona-Lisina N-Metiltransferase/metabolismo , Mitocôndrias Cardíacas/metabolismo , Proteínas Musculares/metabolismo , Miocárdio/enzimologia , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/biossíntese , Fatores de Transcrição/metabolismo , Animais , Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica , Histona Metiltransferases , Histona-Lisina N-Metiltransferase/genética , Camundongos , Camundongos Knockout , Mitocôndrias Cardíacas/genética , Proteínas Musculares/genética , PPAR alfa/biossíntese , PPAR alfa/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Receptor X Retinoide alfa/biossíntese , Receptor X Retinoide alfa/genética , Fatores de Transcrição/genéticaRESUMO
The five membered SET and MYND Domain-containing lysine methyltransferase (SMYD) family plays pivotal roles in development and proliferation. Initially characterized within the cardiovascular system, one such member, SMYD2, has been implicated as an oncogene in leukemias deriving from flawed hematopoietic stem cell (HSC) differentiation. We show here that conditional SMYD2 loss disrupts hematopoiesis at and downstream of the HSC via both apoptotic loss and transcriptional deregulation of HSC proliferation and disruption of Wnt-ß-Catenin signaling. Yet, previously documented SMYD2 cell cycle targets were unscathed. Turning our analysis to human leukemias, we observed that SMYD2 is highly expressed in CML, MLLr-B-ALL, AML, T-ALL, and B-ALL leukemias and its levels in B-ALL correlate with poor survival. SMYD2 knockdown results in apoptotic death and loss of anchorage-independent transformation of each of these hematopoietic leukemias. These data provide an underlying mechanism by which SMYD2 acts during normal hematopoiesis and as a proto-oncogene in leukemia.
Assuntos
Histona-Lisina N-Metiltransferase/metabolismo , Leucemia/metabolismo , Animais , Linhagem Celular Tumoral , Transplante de Células-Tronco Hematopoéticas/métodos , Histona-Lisina N-Metiltransferase/genética , Histona-Lisina N-Metiltransferase/fisiologia , Humanos , Leucemia/genética , Linfócitos/metabolismo , Linfócitos/fisiologia , Lisina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Oxigenases de Função Mista/genética , Oxigenases de Função Mista/metabolismo , Proto-Oncogene MasRESUMO
BACKGROUND: Current methods to repair ablation-type peripheral nerve injuries (PNIs) using peripheral nerve allografts (PNAs) often result in poor functional recovery due to immunological rejection as well as to slow and inaccurate outgrowth of regenerating axonal sprouts. In contrast, ablation-type PNIs repaired by PNAs, using a multistep protocol in which one step employs the membrane fusogen polyethylene glycol (PEG), permanently restore sciatic-mediated behaviors within weeks. Axons and cells within PEG-fused PNAs remain viable, even though outbred host and donor tissues are neither immunosuppressed nor tissue matched. PEG-fused PNAs exhibit significantly reduced T cell and macrophage infiltration, expression of major histocompatibility complex I/II and consistently low apoptosis. In this study, we analyzed the coding transcriptome of PEG-fused PNAs to examine possible mechanisms underlying immunosuppression. METHODS: Ablation-type sciatic PNIs in adult Sprague-Dawley rats were repaired using PNAs and a PEG-fusion protocol combined with neurorrhaphy. Electrophysiological and behavioral tests confirmed successful PEG-fusion of PNAs. RNA sequencing analyzed differential expression profiles of protein-coding genes between PEG-fused PNAs and negative control PNAs (not treated with PEG) at 14 days PO, along with unoperated control nerves. Sequencing results were validated by quantitative reverse transcription PCR (RT-qPCR), and in some cases, immunohistochemistry. RESULTS: PEG-fused PNAs display significant downregulation of many gene transcripts associated with innate and adaptive allorejection responses. Schwann cell-associated transcripts are often upregulated, and cellular processes such as extracellular matrix remodeling and cell/tissue development are particularly enriched. Transcripts encoding several potentially immunosuppressive proteins (e.g., thrombospondins 1 and 2) also are upregulated in PEG-fused PNAs. CONCLUSIONS: This study is the first to characterize the coding transcriptome of PEG-fused PNAs and to identify possible links between alterations of the extracellular matrix and suppression of the allorejection response. The results establish an initial molecular basis to understand mechanisms underlying PEG-mediated immunosuppression.
Assuntos
Aloenxertos/fisiologia , Perfilação da Expressão Gênica/métodos , Tolerância Imunológica/fisiologia , Polietilenoglicóis/administração & dosagem , Nervo Isquiático/fisiologia , Nervo Isquiático/transplante , Animais , Feminino , Mapas de Interação de Proteínas/fisiologia , Ratos , Ratos Sprague-DawleyRESUMO
Ablation/segmental loss peripheral nerve injuries (PNIs) exhibit poor functional recovery due to slow and inaccurate outgrowth of regenerating axons. Viable peripheral nerve allografts (PNAs) as growth-guide conduits are immunologically rejected and all anucleated donor/host axonal segments undergo Wallerian degeneration. In contrast, we report that ablation-type sciatic PNIs repaired by neurorrhaphy of viable sciatic PNAs and a polyethylene glycol (PEG)-fusion protocol using PEG immediately restored axonal continuity for many axons, reinnervated/maintained their neuromuscular junctions, and prevented much Wallerian degeneration. PEG-fused PNAs permanently restored many sciatic-mediated behaviors within 2-6 weeks. PEG-fused PNAs were not rejected even though host/donors were neither immunosuppressed nor tissue-matched in outbred female Sprague Dawley rats. Innate and adaptive immune responses to PEG-fused sciatic PNAs were analyzed using electron microscopy, immunohistochemistry, and quantitative reverse transcription polymerase chain reaction for morphological features, T cell and macrophage infiltration, major histocompatibility complex (MHC) expression, apoptosis, expression of cytokines, chemokines, and cytotoxic effectors. PEG-fused PNAs exhibited attenuated innate and adaptive immune responses by 14-21 days postoperatively, as evidenced by (a) many axons and cells remaining viable, (b) significantly reduced infiltration of cytotoxic and total T cells and macrophages, (c) significantly reduced expression of inflammatory cytokines, chemokines, and MHC proteins, (d) consistently low apoptotic response. Morphologically and/or biochemically, PEG-fused sciatic PNAs often resembled sciatic autografts or intact sciatic nerves. In brief, PEG-fused PNAs are an unstudied, perhaps unique, example of immune tolerance of viable allograft tissue in a nonimmune-privileged environment and could greatly improve the clinical outcomes for PNIs relative to current protocols.
Assuntos
Imunidade Adaptativa/fisiologia , Imunidade Inata/fisiologia , Polietilenoglicóis/administração & dosagem , Nervo Isquiático/imunologia , Nervo Isquiático/transplante , Neuropatia Ciática/terapia , Imunidade Adaptativa/efeitos dos fármacos , Aloenxertos/imunologia , Aloenxertos/transplante , Animais , Feminino , Imunidade Inata/efeitos dos fármacos , Masculino , Ratos , Ratos Sprague-Dawley , Neuropatia Ciática/imunologia , Transplante Homólogo/métodosRESUMO
Direct reprogramming can be achieved by forced expression of master transcription factors. Yet how such factors mediate repression of initial cell-type-specific genes while activating target cell-type-specific genes is unclear. Through embryonic stem (ES) to trophoblast stem (TS)-like cell reprogramming by introducing individual TS cell-specific 'CAG' factors (Cdx2, Arid3a and Gata3), we interrogate their chromosomal target occupancies, modulation of global transcription and chromatin accessibility at the initial stage of reprogramming. From the studies, we uncover a sequential, two-step mechanism of cellular reprogramming in which repression of pre-existing ES cell-associated gene expression program is followed by activation of TS cell-specific genes by CAG factors. Therefore, we reveal that CAG factors function as both decommission and pioneer factors during ES to TS-like cell fate conversion.
Assuntos
Fator de Transcrição CDX2/fisiologia , Técnicas de Reprogramação Celular , Proteínas de Ligação a DNA/fisiologia , Células-Tronco Embrionárias/citologia , Fator de Transcrição GATA3/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Fatores de Transcrição/fisiologia , Trofoblastos/citologia , Animais , Fator de Transcrição CDX2/genética , Células Cultivadas , Cromatina/genética , Cromatina/metabolismo , Meios de Cultivo Condicionados , Proteínas de Ligação a DNA/genética , Elementos Facilitadores Genéticos , Fibroblastos , Fator de Transcrição GATA3/genética , Ontologia Genética , Código das Histonas , Camundongos , Fatores de Transcrição/genética , Transcrição GênicaRESUMO
High expression of the forkhead box P1 (FOXP1) transcription factor distinguishes the aggressive activated B cell (ABC) diffuse large B-cell lymphoma (DLBCL) subtype from the better prognosis germinal center B-cell (GCB)-DLBCL subtype and is highly correlated with poor outcomes. A genetic or functional role for FOXP1 in lymphomagenesis, however, remains unknown. Here, we report that sustained FOXP1 expression is vital for ABC-DLBCL cell-line survival. Genome-wide analyses revealed direct and indirect FOXP1 transcriptional enforcement of ABC-DLBCL hallmarks, including the classical NF-κB and MYD88 (myeloid differentiation primary response gene 88) pathways. FOXP1 promoted gene expression underlying transition of the GCB cell to the plasmablast--the transient B-cell stage targeted in ABC-DLBCL transformation--by antagonizing pathways distinctive of GCB-DLBCL, including that of the GCB "master regulator," BCL6 (B-cell lymphoma 6). Cell-line derived FOXP1 target genes that were highly correlated with FOXP1 expression in primary DLBCL accurately segregated the corresponding clinical subtypes of a large cohort of primary DLBCL isolates and identified conserved pathways associated with ABC-DLBCL pathology.
Assuntos
Linfócitos B/imunologia , Fatores de Transcrição Forkhead/fisiologia , Linfoma Difuso de Grandes Células B/imunologia , Proteínas Repressoras/fisiologia , Diferenciação Celular , Linhagem Celular Tumoral , Humanos , Ativação Linfocitária , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/patologia , Transcrição GênicaRESUMO
Genetic perturbations of the transcription factor Forkhead Box P1 (FOXP1) are causative for severe forms of autism spectrum disorder that are often comorbid with intellectual disability. Recent work has begun to reveal an important role for FoxP1 in brain development, but the brain-region-specific contributions of Foxp1 to autism and intellectual disability phenotypes have yet to be determined fully. Here, we describe Foxp1 conditional knock-out (Foxp1cKO) male and female mice with loss of Foxp1 in the pyramidal neurons of the neocortex and the CA1/CA2 subfields of the hippocampus. Foxp1cKO mice exhibit behavioral phenotypes that are of potential relevance to autism spectrum disorder, including hyperactivity, increased anxiety, communication impairments, and decreased sociability. In addition, Foxp1cKO mice have gross deficits in learning and memory tasks of relevance to intellectual disability. Using a genome-wide approach, we identified differentially expressed genes in the hippocampus of Foxp1cKO mice associated with synaptic function and development. Furthermore, using magnetic resonance imaging, we uncovered a significant reduction in the volumes of both the entire hippocampus as well as individual hippocampal subfields of Foxp1cKO mice. Finally, we observed reduced maintenance of LTP in area CA1 of the hippocampus in these mutant mice. Together, these data suggest that proper expression of Foxp1 in the pyramidal neurons of the forebrain is important for regulating gene expression pathways that contribute to specific behaviors reminiscent of those seen in autism and intellectual disability. In particular, Foxp1 regulation of gene expression appears to be crucial for normal hippocampal development, CA1 plasticity, and spatial learning.SIGNIFICANCE STATEMENT Loss-of-function mutations in the transcription factor Forkhead Box P1 (FOXP1) lead to autism spectrum disorder and intellectual disability. Understanding the potential brain-region-specific contributions of FOXP1 to disease-relevant phenotypes could be a critical first step in the management of patients with these mutations. Here, we report that Foxp1 conditional knock-out (Foxp1cKO) mice with loss of Foxp1 in the neocortex and hippocampus display autism and intellectual-disability-relevant behaviors. We also show that these phenotypes correlate with changes in both the genomic and physiological profiles of the hippocampus in Foxp1cKO mice. Our work demonstrates that brain-region-specific FOXP1 expression may relate to distinct, clinically relevant phenotypes.
Assuntos
Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/fisiologia , Expressão Gênica/genética , Expressão Gênica/fisiologia , Plasticidade Neuronal/genética , Plasticidade Neuronal/fisiologia , Células Piramidais/fisiologia , Proteínas Repressoras/genética , Proteínas Repressoras/fisiologia , Aprendizagem Espacial/fisiologia , Sinapses/fisiologia , Animais , Transtorno do Espectro Autista , Comportamento Animal/fisiologia , Região CA1 Hipocampal/fisiologia , Feminino , Deficiências da Aprendizagem/genética , Deficiências da Aprendizagem/psicologia , Masculino , Transtornos da Memória/genética , Transtornos da Memória/psicologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neocórtex/citologia , Neocórtex/fisiologia , Células Piramidais/metabolismo , Vocalização Animal/fisiologiaRESUMO
Previous studies in the mouse indicated that ARID3A plays a critical role in the first cell fate decision required for generation of trophectoderm (TE). Here, we demonstrate that ARID3A is widely expressed during mouse and human placentation and essential for early embryonic viability. ARID3A localizes to trophoblast giant cells and other trophoblast-derived cell subtypes in the junctional and labyrinth zones of the placenta. Conventional Arid3a knockout embryos suffer restricted intrauterine growth with severe defects in placental structural organization. Arid3a null placentas show aberrant expression of subtype-specific markers as well as significant alteration in cytokines, chemokines and inflammatory response-related genes, including previously established markers of human placentation disorders. BMP4-mediated induction of trophoblast stem (TS)-like cells from human induced pluripotent stem cells results in ARID3A up-regulation and cytoplasmic to nuclear translocation. Overexpression of ARID3A in BMP4-mediated TS-like cells up-regulates TE markers, whereas pluripotency markers are down-regulated. Our results reveal an essential, conserved function for ARID3A in mammalian placental development through regulation of both intrinsic and extrinsic developmental programs.
Assuntos
Proteína Morfogenética Óssea 4/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Placenta/metabolismo , Placentação/fisiologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transporte Ativo do Núcleo Celular/fisiologia , Animais , Diferenciação Celular , Células Cultivadas , Citocinas/metabolismo , Embrião de Mamíferos/metabolismo , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Células Gigantes/metabolismo , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Morfogênese/fisiologia , Gravidez , Trofoblastos/citologiaRESUMO
Many publications report that ablations of segments of peripheral nerves produce the following unfortunate results: (1) Immediate loss of sensory signaling and motor control; (2) rapid Wallerian degeneration of severed distal axons within days; (3) muscle atrophy within weeks; (4) poor behavioral (functional) recovery after many months, if ever, by slowly-regenerating (â¼1mm/d) axon outgrowths from surviving proximal nerve stumps; and (5) Nerve allografts to repair gap injuries are rejected, often even if tissue matched and immunosuppressed. In contrast, using a female rat sciatic nerve model system, we report that neurorrhaphy of allografts plus a well-specified-sequence of solutions (one containing polyethylene glycol: PEG) successfully addresses each of these problems by: (a) Reestablishing axonal continuity/signaling within minutes by nonspecific ally PEG-fusing (connecting) severed motor and sensory axons across each anastomosis; (b) preventing Wallerian degeneration by maintaining many distal segments of inappropriately-reconnected, PEG-fused axons that continuously activate nerve-muscle junctions; (c) maintaining innervation of muscle fibers that undergo much less atrophy than otherwise-denervated muscle fibers; (d) inducing remarkable behavioral recovery to near-unoperated levels within days to weeks, almost certainly by CNS and PNS plasticities well-beyond what most neuroscientists currently imagine; and (e) preventing rejection of PEG-fused donor nerve allografts with no tissue matching or immunosuppression. Similar behavioral results are produced by PEG-fused autografts. All results for Negative Control allografts agree with current neuroscience data 1-5 given above. Hence, PEG-fusion of allografts for repair of ablated peripheral nerve segments expand on previous observations in single-cut injuries, provoke reconsideration of some current neuroscience dogma, and further extend the potential of PEG-fusion in clinical practice.
Assuntos
Regeneração Nervosa/efeitos dos fármacos , Nervo Fibular/efeitos dos fármacos , Nervo Fibular/transplante , Polietilenoglicóis/farmacologia , Nervo Isquiático/efeitos dos fármacos , Neuropatia Ciática/terapia , Aloenxertos/efeitos dos fármacos , Animais , Axônios/efeitos dos fármacos , Axônios/fisiologia , Axotomia , Modelos Animais de Doenças , Feminino , Músculo Esquelético , Fibras Nervosas/efeitos dos fármacos , Condução Nervosa/efeitos dos fármacos , Junção Neuromuscular/efeitos dos fármacos , Traumatismos dos Nervos Periféricos/patologia , Traumatismos dos Nervos Periféricos/terapia , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica/efeitos dos fármacos , Nervo Isquiático/patologia , Nervo Isquiático/fisiologia , Nervo Isquiático/cirurgia , Neuropatia Ciática/induzido quimicamente , Transplante Homólogo , Degeneração Walleriana/prevenção & controleRESUMO
In the developing mouse embryo, the first hematopoietic stem cells (HSCs) arise in the aorta-gonad-mesonephros (AGM) and mature as they transit through the fetal liver (FL). Compared with FL and adult HSCs, AGM HSCs have reduced repopulation potential in irradiated adult transplant recipients but mechanisms underlying this deficiency in AGM HSCs are poorly understood. By co-expression gene network analysis, we deduced that AGM HSCs show lower levels of interferon-α (IFN-α)/Jak-Stat1-associated gene expression than FL HSCs. Treatment of AGM HSCs with IFN-α enhanced long-term hematopoietic engraftment and donor chimerism. Conversely, IFN-α receptor-deficient AGMs (Ifnαr1(-/-)), had significantly reduced donor chimerism. We identify adenine-thymine-rich interactive domain-3a (Arid3a), a factor essential for FL and B lymphopoiesis, as a key transcriptional co-regulator of IFN-α/Stat1 signaling. Arid3a occupies the genomic loci of Stat1 as well as several IFN-α effector genes, acting to regulate their expression. Accordingly, Arid3a(-/-) AGM HSCs had significantly reduced transplant potential, which was rescued by IFN-α treatment. Our results implicate the inflammatory IFN-α/Jak-Stat pathway in the developmental maturation of embryonic HSCs, whose manipulation may lead to increased potency of reprogrammed HSCs for transplantation.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Embrião de Mamíferos/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Interferon-alfa/metabolismo , Receptor de Interferon alfa e beta/metabolismo , Transdução de Sinais/fisiologia , Fatores de Transcrição/metabolismo , Animais , Proteínas de Ligação a DNA/genética , Embrião de Mamíferos/citologia , Células-Tronco Hematopoéticas/citologia , Interferon-alfa/genética , Camundongos , Camundongos Knockout , Receptor de Interferon alfa e beta/genética , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/metabolismo , Fatores de Transcrição/genéticaRESUMO
The SMYD (SET and MYND domain) family of lysine methyltransferases harbor a unique structure in which the methyltransferase (SET) domain is intervened by a zinc finger protein-protein interaction MYND domain. SMYD proteins methylate both histone and non-histone substrates and participate in diverse biological processes including transcriptional regulation, DNA repair, proliferation and apoptosis. Smyd1 is unique among the five family members in that it is specifically expressed in striated muscles. Smyd1 is critical for development of the right ventricle in mice. In zebrafish, Smyd1 is necessary for sarcomerogenesis in fast-twitch muscles. Smyd1 is expressed in the skeletal muscle lineage throughout myogenesis and in mature myofibers, shuttling from nucleus to cytosol during myoblast differentiation. Because of this expression pattern, we hypothesized that Smyd1 plays multiple roles at different stages of myogenesis. To determine the role of Smyd1 in mammalian myogenesis, we conditionally eliminated Smyd1 from the skeletal muscle lineage at the myoblast stage using Myf5(cre). Deletion of Smyd1 impaired myoblast differentiation, resulted in fewer myofibers and decreased expression of muscle-specific genes. Muscular defects were temporally restricted to the second wave of myogenesis. Thus, in addition to the previously described functions for Smyd1 in heart development and skeletal muscle sarcomerogenesis, these results point to a novel role for Smyd1 in myoblast differentiation.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Desenvolvimento Muscular , Proteínas Musculares/fisiologia , Fatores de Transcrição/fisiologia , Animais , Diferenciação Celular , Células Cultivadas , Proteínas de Ligação a DNA/análise , Camundongos , Fibras Musculares Esqueléticas , Proteínas Musculares/análise , Mioblastos/citologia , Fatores de Transcrição/análiseRESUMO
The inhibitory mechanisms that prevent gene expression programs from one tissue to be expressed in another are poorly understood. Foxp1/2/4 are forkhead transcription factors that repress gene expression and are individually important for endoderm development. We show that combined loss of all three Foxp1/2/4 family members in the developing anterior foregut endoderm leads to a loss of lung endoderm lineage commitment and subsequent development. Foxp1/2/4 deficient lungs express high levels of transcriptional regulators not normally expressed in the developing lung, including Pax2, Pax8, Pax9 and the Hoxa9-13 cluster. Ectopic expression of these transcriptional regulators is accompanied by decreased expression of lung restricted transcription factors including Nkx2-1, Sox2, and Sox9. Foxp1 binds to conserved forkhead DNA binding sites within the Hoxa9-13 cluster, indicating a direct repression mechanism. Thus, Foxp1/2/4 are essential for promoting lung endoderm development by repressing expression of non-pulmonary transcription factors.
Assuntos
Fatores de Transcrição Forkhead/genética , Regulação da Expressão Gênica no Desenvolvimento , Pulmão/embriologia , Proteínas Repressoras/genética , Animais , Sítios de Ligação , Sequência Conservada , DNA/genética , DNA/metabolismo , Endoderma/citologia , Endoderma/embriologia , Fatores de Transcrição Forkhead/deficiência , Fatores de Transcrição Forkhead/metabolismo , Genes Homeobox , Camundongos , Especificidade de Órgãos , Organogênese , Proteínas Repressoras/deficiência , Proteínas Repressoras/metabolismo , Alinhamento de Sequência , Homologia de Sequência do Ácido Nucleico , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genéticaRESUMO
The regulatory logic underlying global transcriptional programs controlling development of visceral organs like the pancreas remains undiscovered. Here, we profiled gene expression in 12 purified populations of fetal and adult pancreatic epithelial cells representing crucial progenitor cell subsets, and their endocrine or exocrine progeny. Using probabilistic models to decode the general programs organizing gene expression, we identified co-expressed gene sets in cell subsets that revealed patterns and processes governing progenitor cell development, lineage specification, and endocrine cell maturation. Purification of Neurog3 mutant cells and module network analysis linked established regulators such as Neurog3 to unrecognized gene targets and roles in pancreas development. Iterative module network analysis nominated and prioritized transcriptional regulators, including diabetes risk genes. Functional validation of a subset of candidate regulators with corresponding mutant mice revealed that the transcription factors Etv1, Prdm16, Runx1t1 and Bcl11a are essential for pancreas development. Our integrated approach provides a unique framework for identifying regulatory genes and functional gene sets underlying pancreas development and associated diseases such as diabetes mellitus.
Assuntos
Separação Celular/métodos , Regulação da Expressão Gênica no Desenvolvimento , Pâncreas/citologia , Pâncreas/embriologia , Pâncreas/crescimento & desenvolvimento , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Genômica/métodos , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/fisiologia , Camundongos Mutantes , Camundongos Transgênicos , Modelos Estatísticos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Reprodutibilidade dos Testes , Fatores de Transcrição SOX9/genética , Células-Tronco/citologia , Células-Tronco/fisiologiaRESUMO
The plasmacytoid dendritic cell (pDC) is vital to the coordinated action of innate and adaptive immunity. pDC development has not been unequivocally traced, nor has its transcriptional regulatory network been fully clarified. Here we confirm an essential requirement for the BCL11A transcription factor in fetal pDC development, and demonstrate this lineage-specific requirement in the adult organism. Furthermore, we identify BCL11A gene targets and provide a molecular mechanism for its action in pDC commitment. Embryonic germ-line deletion of Bcl11a revealed an absolute cellular, molecular, and functional absence of pDCs in fetal mice. In adults, deletion of Bcl11a in hematopoietic stem cells resulted in perturbed yet continued generation of progenitors, loss of downstream pDC and B-cell lineages, and persisting myeloid, conventional dendritic, and T-cell lineages. Challenge with virus resulted in a marked reduction of antiviral response in conditionally deleted adults. Genome-wide analyses of BCL11A DNA binding and expression revealed that BCL11A regulates transcription of E2-2 and other pDC differentiation modulators, including ID2 and MTG16. Our results identify BCL11A as an essential, lineage-specific factor that regulates pDC development, supporting a model wherein differentiation into pDCs represents a primed "default" pathway for common dendritic cell progenitors.