ABSTRACT
Bioconductor is an open-source, open-development software project for the analysis and comprehension of high-throughput data in genomics and molecular biology. The project aims to enable interdisciplinary research, collaboration and rapid development of scientific software. Based on the statistical programming language R, Bioconductor comprises 934 interoperable packages contributed by a large, diverse community of scientists. Packages cover a range of bioinformatic and statistical applications. They undergo formal initial review and continuous automated testing. We present an overview for prospective users and contributors.
Subject(s)
Computational Biology , Gene Expression Profiling , Genomics/methods , High-Throughput Screening Assays/methods , Software , Programming Languages , User-Computer InterfaceABSTRACT
We describe Bioconductor infrastructure for representing and computing on annotated genomic ranges and integrating genomic data with the statistical computing features of R and its extensions. At the core of the infrastructure are three packages: IRanges, GenomicRanges, and GenomicFeatures. These packages provide scalable data structures for representing annotated ranges on the genome, with special support for transcript structures, read alignments and coverage vectors. Computational facilities include efficient algorithms for overlap and nearest neighbor detection, coverage calculation and other range operations. This infrastructure directly supports more than 80 other Bioconductor packages, including those for sequence analysis, differential expression analysis and visualization.
Subject(s)
Databases, Genetic , Genomics/methods , Software , Algorithms , Animals , Genomics/standards , Humans , Mice , Sequence Alignment , Sequence Analysis, DNAABSTRACT
Infants exposed to HIV but uninfected (iHEU) display altered cellular immunity and are at increased risk of infection through poorly understood mechanisms. We previously reported that iHEU have lower levels of maternal microchimerism (MMc), maternal cells transferred to the offspring in utero/during breastfeeding. We evaluated MMc levels in T cell subsets in iHEU and HIV unexposed infants (iHU) to determine whether a selective deficiency in MMc may contribute to altered cellular immunity. Across all infants, MMc levels were highest in CD8+ T cells; however, the level of MMc in the CD8 T cell subset was significantly lower in iHEU compared to iHU.
ABSTRACT
T cells in the human female genital tract (FGT) 2 are key mediators of susceptibility to and protection from infection, including HIV and other sexually transmitted infections. There is a critical need for increased understanding of the distribution and activation of T cell populations in the FGT, but current sampling methods require a healthcare provider and are expensive, limiting the ability to study these populations longitudinally. To address these challenges, we have developed a method to sample immune cells from the FGT utilizing disposable menstrual discs which are non-invasive, self-applied, and low-cost. To demonstrate reproducibility, we sampled the cervicovaginal fluid (CVF) 3 of healthy, reproductive-aged individuals using menstrual discs over three sequential days. CVF was processed for cervicovaginal cells, and high parameter flow cytometry was used to characterize immune populations. We identified large numbers of live, CD45+ leukocytes, as well as distinct populations of T cells and B cells. Within the T cell compartment, activation and suppression status of T cell subsets were consistent with previous studies of the FGT utilizing current approaches, including identification of both tissue resident and migratory populations. In addition, the T cell population structure was highly conserved across days within individuals but divergent across individuals. Our approach to sample immune cells in the FGT with menstrual discs will decrease barriers to participation and empower longitudinal sampling in future research studies.
ABSTRACT
T cells in the human female genital tract (FGT) are key mediators of susceptibility to and protection from infection, including HIV and other sexually transmitted infections. There is a critical need for increased understanding of the distribution and activation of T cell populations in the FGT, but current sampling methods require a healthcare provider and are expensive, limiting the ability to study these populations longitudinally. To address these challenges, we have developed a method to sample immune cells from the FGT utilizing disposable menstrual discs which are noninvasive, self-applied, and low in cost. To demonstrate reproducibility, we sampled the cervicovaginal fluid of healthy, reproductive-aged individuals using menstrual discs across 3 sequential days. Cervicovaginal fluid was processed for cervicovaginal cells, and high-parameter flow cytometry was used to characterize immune populations. We identified large numbers of live, CD45+ leukocytes, as well as distinct populations of T cells and B cells. Within the T cell compartment, activation and suppression status of T cell subsets were consistent with previous studies of the FGT utilizing current approaches, including identification of both tissue-resident and migratory populations. In addition, the T cell population structure was highly conserved across days within individuals but divergent across individuals. Our approach to sample immune cells in the FGT with menstrual discs will decrease barriers to participation and empower longitudinal sampling in future research studies.
Subject(s)
HIV Infections , Female , Humans , Adult , Reproducibility of Results , Genitalia, Female , T-Lymphocyte SubsetsABSTRACT
Genetic engineering of human lymphocytes for therapeutic applications is constrained by a lack of transgene transcriptional control, resulting in a compromised therapeutic index. Incomplete understanding of transcriptional logic limits the rational design of contextually responsive genetic modules1. Here, we juxtaposed rationally curated transcriptional response element (TRE) oligonucleotides by random concatemerization to generate a library from which we selected context-specific inducible synthetic promoters (iSynPros). Through functional selection, we screened an iSynPro library for "IF-THEN" logic-gated transcriptional responses in human CD8+ T cells expressing a 4-1BB second generation chimeric antigen receptor (CAR). iSynPros exhibiting stringent off-states in quiescent T cells and CAR activation-dependent transcriptional responsiveness were cloned and subjected to TRE composition and pattern analysis, as well as performance in regulating candidate antitumor potency enhancement modules. These data reveal synthetic TRE grammar can mediate logic-gated transgene transcription in human T cells that, when applied to CAR T cell engineering, enhance potency and improve therapeutic indices.
ABSTRACT
Human breastmilk is rich in T cells; however, their specificity and function are largely unknown. We compared the phenotype, diversity, and antigen specificity of T cells in breastmilk and peripheral blood of lactating individuals who received SARS-CoV-2 messenger RNA (mRNA) vaccination. Relative to blood, breastmilk contained higher frequencies of T effector and central memory populations that expressed mucosal-homing markers. T cell receptor sequence overlap was limited between blood and breastmilk. Overabundant breastmilk clones were observed in all individuals, were diverse, and contained complementarity-determining regions in three sequences with known epitope specificity, including to SARS-CoV-2 spike. SARS-CoV-2 spike-specific T cell receptors were more frequent in breastmilk compared to blood and expanded in breastmilk following a 3rd mRNA vaccine dose. Our observations indicate that the lactating breast contains a distinct T cell population that can be modulated by maternal vaccination with potential implications for passive infant protection.
Subject(s)
COVID-19 , Milk, Human , Infant , Female , Humans , SARS-CoV-2 , T-Lymphocytes , Lactation , Vaccination , RNA, Messenger , Antibodies, ViralABSTRACT
Human breastmilk is rich in T cells; however, their specificity and function are largely unknown. We compared the phenotype, diversity, and antigen specificity of T cells in the breastmilk and peripheral blood of lactating individuals who received SARS-CoV-2 mRNA vaccination. Relative to blood, breastmilk contained higher frequencies of T effector and central memory populations that expressed mucosal-homing markers. T cell receptor (TCR) sequence overlap was limited between blood and breastmilk. Overabundan t breastmilk clones were observed in all individuals, were diverse, and contained CDR3 sequences with known epitope specificity including to SARS-CoV-2 Spike. Spike-specific TCRs were more frequent in breastmilk compared to blood and expanded in breastmilk following a third mRNA vaccine dose. Our observations indicate that the lactating breast contains a distinct T cell population that can be modulated by maternal vaccination with potential implications for infant passive protection. One-Sentence Summary: The breastmilk T cell repertoire is distinct and enriched for SARS-CoV-2 Spike-specificity after maternal mRNA vaccination.
ABSTRACT
In addition to thymus-derived or natural T regulatory (nT(reg)) cells, a second subset of induced T regulatory (iT(reg)) cells arises de novo from conventional CD4(+) T cells in the periphery. The function of iT(reg) cells in tolerance was examined in a CD45RB(high)CD4(+) T cell transfer model of colitis. In situ-generated iT(reg) cells were similar to nT(reg) cells in their capacity to suppress T cell proliferation in vitro and their absence in vivo accelerated bowel disease. Treatment with nT(reg) cells resolved the colitis, but only when iT(reg) cells were also present. Although iT(reg) cells required Foxp3 for suppressive activity and phenotypic stability, their gene expression profile was distinct from the established nT(reg) "genetic signature," indicative of developmental and possibly mechanistic differences. These results identified a functional role for iT(reg) cells in vivo and demonstrated that both iT(reg) and nT(reg) cells can act in concert to maintain tolerance.
Subject(s)
Colitis/immunology , Colitis/pathology , Immune Tolerance , T-Lymphocytes, Regulatory/immunology , T-Lymphocytes, Regulatory/pathology , Adoptive Transfer , Animals , Cell Differentiation/genetics , Cell Differentiation/immunology , Cell Survival/genetics , Cell Survival/immunology , Cells, Cultured , Colitis/genetics , Colitis/therapy , Disease Models, Animal , Forkhead Transcription Factors/genetics , Forkhead Transcription Factors/physiology , Green Fluorescent Proteins/biosynthesis , Green Fluorescent Proteins/genetics , Male , Mice , Mice, Inbred BALB C , Mice, Knockout , Mice, Transgenic , Promoter Regions, Genetic/immunologyABSTRACT
Ewing's family tumors (EFTs) are characterized by recurrent chromosomal translocations that produce chimeric fusions between the EWS gene and one of five ETS transcription factors. The expression of EWS/FLI1, the predominant fusion product in EFTs, is believed to deregulate downstream target genes in an undefined tissue type and leads to development of EFTs. Attempts to generate model systems that represent EFTs have been hampered by an unexpected toxicity of the fusion gene. In the present study, we used gene expression analysis to identify tissue types based on the similarity of their expression profiles to those of EWS/FLI1-modulated genes. The data obtained from this screen helped to identify IMR-90 cells, a human fetal fibroblast, that upon further manipulation can maintain stable EWS/FLI1 expression without the reported toxicity. In addition, gene expression profiling of these cells revealed a significant overlap of genes that have been previously reported to be targets of EWS/FLI1. Furthermore, we show, for the first time, a partial transformation of these human primary fibroblasts with EWS/FLI1 expression. The experiments presented here provide a solid foundation for generation of a new model system for studying Ewing's sarcoma biology.
Subject(s)
Mesoderm/pathology , Models, Biological , Oncogene Proteins, Fusion/genetics , Sarcoma, Ewing/genetics , Transcription Factors/genetics , Apoptosis , Blotting, Western , Cell Line , Gene Expression Profiling , Gene Silencing , Humans , Mesenchymal Stem Cells/pathology , Polymerase Chain Reaction , Proto-Oncogene Protein c-fli-1 , RNA-Binding Protein EWSABSTRACT
PURPOSE: To determine the difference in gene expression between completely versus incompletely enhancing glioblastoma multiforme (GBM). MATERIALS AND METHODS: Gene expression was determined for 52 newly diagnosed GBMs by using DNA microarrays, and the relationship to enhancement pattern and survival was analyzed. This study was approved by the institutional review board and was HIPAA compliant; informed consent was obtained. RESULTS: Thirty-eight percent (20 of 52) of GBMs were incompletely enhancing (IE). The expression of eight genes was increased more than twofold in IE GBM when compared with completely enhancing (CE) GBM. Among these were tight junction protein-2 (2.2-fold increase, P = .019), and the oligodendroglioma markers oligodendrocyte lineage transcription factor 2 (2.4-fold increase, P = .029) and Achaete-scute complex-like 1 (ASCL1; 2.7-fold increase, P = .023). The expression of 71 genes showed relative overexpression in CE when compared with IE GBM. These included several proangiogenic and edema-related genes, including vascular endothelial growth factor (2.1-fold, P = .005) and neuronal pentraxin-2 (3.0-fold, P = .029). Several genes associated with primary GBM were overexpressed in CE tumors, whereas ASCL1, which is associated with secondary GBM, was overexpressed in IE tumors. Many genes overexpressed in IE GBM were associated with longer survival, whereas several genes overexpressed in CE GBM correlated with shortened survival. CONCLUSION: The enhancement pattern divides GBM in two groups with differing prognoses. By comparing gene expression between IE and CE GBMs, it was possible to identify genes that may affect magnetic resonance imaging features of edema and enhancement, and genes whose expression levels are predictive of both improved and shortened survival.
Subject(s)
Brain Neoplasms/genetics , Gene Expression Regulation, Neoplastic , Glioblastoma/genetics , Magnetic Resonance Imaging , Oligonucleotide Array Sequence Analysis , Basic Helix-Loop-Helix Transcription Factors/genetics , Brain Neoplasms/mortality , C-Reactive Protein/genetics , Glioblastoma/mortality , Humans , Membrane Proteins/genetics , Nerve Tissue Proteins/genetics , Oligodendrocyte Transcription Factor 2 , Vascular Endothelial Growth Factor A/genetics , Zonula Occludens-2 ProteinABSTRACT
PURPOSE: Vascular endothelial growth factor (VEGF) is a potent mediator of vascular permeability. VEGF inhibition reduces edema and tumor burden in some patients with malignant glioma, whereas others show no response. The role of VEGF expression in edema production and the relationship to survival is not well understood. EXPERIMENTAL DESIGN: Using DNA microarray analysis, we examined VEGF and related gene expression in 71 newly diagnosed malignant gliomas and analyzed the relationship to edema and survival. RESULTS AND CONCLUSIONS: VEGF expression was predictive of survival in tumors with little or no edema [Cox proportional hazard model, 6.88; 95% confidence interval (95% CI), 2.61-18.1; P<0.0001], but not in tumors with extensive edema. The expression of several proangiogenic genes, including adrenomedullin (correlation coefficient, 0.80), hypoxia-inducible factor-1A (0.51), and angiopoietin-2 (0.44), was correlated with VEGF expression (all with P<0.0001), whereas that of several antiangiogenic genes was inversely correlated. The expression of six genes was increased greater than 3-fold in edematous versus nonedematous tumors in the absence of increased VEGF expression. The most increased, neuronal pentraxin 2 (NPTX2, 7-fold change), was predictive of survival in tumors with the highest levels of edema, in contrast to VEGF (hazard ratio, 2.73; 95% CI, 1.49-5.02; P=0.049). NPTX2 was tightly correlated with expression of the water channel aquaporin-3 (0.74, P<0.0001). These results suggest that there are both VEGF-dependent and VEGF-independent pathways of edema production in gliomas and may explain why edema is not reduced in some patients following anti-VEGF treatment.
Subject(s)
Brain Edema/diagnosis , Brain Neoplasms/mortality , C-Reactive Protein/genetics , Glioma/mortality , Nerve Tissue Proteins/genetics , Vascular Endothelial Growth Factor A/genetics , Adrenomedullin/genetics , Angiopoietin-2/genetics , Aquaporin 3/genetics , Brain Edema/genetics , Brain Neoplasms/complications , Gene Expression , Glioma/complications , Humans , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Oligonucleotide Array Sequence Analysis , Prognosis , SurvivalABSTRACT
BACKGROUND: Genes and proteins are organized into functional modular networks in which the network context of a gene or protein has implications for cellular function. Highly connected hub proteins, largely responsible for maintaining network connectivity, have been found to be much more likely to be essential for yeast survival. RESULTS: Here we investigate the properties of weighted gene co-expression networks formed from multiple microarray datasets. The constructed networks approximate scale-free topology, but this is not universal across all datasets. We show strong positive correlations between gene connectivity within the whole network and gene essentiality as well as gene sequence conservation. We demonstrate the preservation of a modular structure of the networks formed, and demonstrate that, within some of these modules, it is possible to observe a strong correlation between connectivity and essentiality or between connectivity and conservation within the modules particularly within modules containing larger numbers of essential genes. CONCLUSION: Application of these techniques can allow a finer scale prediction of relative gene importance for a particular process within a group of similarly expressed genes.
Subject(s)
Fungal Proteins/genetics , Gene Expression Regulation, Fungal , Models, Genetic , Saccharomyces/genetics , Base Sequence , Cell Cycle/genetics , Conserved Sequence , DNA Damage , DNA, Fungal/chemistry , Fungal Proteins/physiology , Gene Expression Profiling , Genomics , Oligonucleotide Array Sequence Analysis , Saccharomyces/metabolismABSTRACT
Annotation resources make up a significant proportion of the Bioconductor project (Huber et al., Nat Methods 12:115-121, 2015). And there are also a diverse set of online resources available which are accessed using specific packages. Here we describe the most popular of these resources and give some high level examples on how to use them.
Subject(s)
Computational Biology/methods , Genomics/methods , Molecular Sequence Annotation/methods , SoftwareABSTRACT
PURPOSE: To identify proteinases and growth factors abnormally expressed in human corneas of donors with diabetic retinopathy (DR), additional to previously described matrix metalloproteinase (MMP)-10 and -3 and insulin-like growth factor (IGF)-I. METHODS: RNA was isolated from 35 normal, diabetic, and DR autopsy human corneas ex vivo or after organ culture. Amplified cRNA was analyzed using 22,000-gene microarrays (Agilent Technologies, Palo Alto, CA). Gene expression in each diabetic corneal cRNA was assessed against pooled cRNA from 7 to 9 normal corneas. Select differentially expressed genes were validated by quantitative real-time RT-PCR (QPCR) and immunohistochemistry. Organ cultures were treated with a cathepsin inhibitor, cystatin C, or MMP-10. RESULTS: More than 100 genes were upregulated and 2200 were downregulated in DR corneas. Expression of cathepsin F and hepatocyte growth factor (HGF) genes was increased in ex vivo and organ-cultured DR corneas compared with normal corneas. HGF receptor c-met, fibroblast growth factor (FGF)-3, its receptor FGFR3, tissue inhibitor of metalloproteinase (TIMP)-4, laminin alpha4 chain, and thymosin beta(4) genes were downregulated. The data were corroborated by QPCR and immunohistochemistry analyses; main changes of these components occurred in corneal epithelium. In organ-cultured DR corneas, cystatin C increased laminin-10 and integrin alpha(3)beta(1), whereas in normal corneas MMP-10 decreased laminin-10 and integrin alpha(3)beta(1) expression. CONCLUSIONS: Elevated cathepsin F and the ability of its inhibitor to produce a more normal phenotype in diabetic corneas suggest increased proteolysis in these corneas. Proteinase changes may result from abnormalities of growth factors, such as HGF and FGF-3, in DR corneas. Specific modulation of proteinases and growth factors could reduce diabetic corneal epitheliopathy.
Subject(s)
Cornea/metabolism , Corneal Diseases/genetics , Diabetic Retinopathy/genetics , Gene Expression Regulation , Growth Substances/genetics , Peptide Hydrolases/genetics , Aged , Corneal Diseases/enzymology , Diabetic Retinopathy/enzymology , Fluorescent Antibody Technique, Indirect , Gene Expression Profiling , Growth Substances/metabolism , Humans , Oligonucleotide Array Sequence Analysis , Organ Culture Techniques , Peptide Hydrolases/metabolism , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Tissue Donors , Up-RegulationABSTRACT
The Tyrolean Iceman, a 5,300-year-old Copper age individual, was discovered in 1991 on the Tisenjoch Pass in the Italian part of the Ötztal Alps. Here we report the complete genome sequence of the Iceman and show 100% concordance between the previously reported mitochondrial genome sequence and the consensus sequence generated from our genomic data. We present indications for recent common ancestry between the Iceman and present-day inhabitants of the Tyrrhenian Sea, that the Iceman probably had brown eyes, belonged to blood group O and was lactose intolerant. His genetic predisposition shows an increased risk for coronary heart disease and may have contributed to the development of previously reported vascular calcifications. Sequences corresponding to ~60% of the genome of Borrelia burgdorferi are indicative of the earliest human case of infection with the pathogen for Lyme borreliosis.
Subject(s)
Genome, Human , Genome, Mitochondrial , Mummies , Base Sequence , Borrelia burgdorferi/genetics , Chromosome Mapping , DNA, Mitochondrial/genetics , Genetic Predisposition to Disease , History, Ancient , Humans , Lyme Disease/history , Mitochondria/genetics , Mummies/microbiology , Paleontology , Phenotype , Sequence Analysis, DNA , Vascular CalcificationABSTRACT
We previously reported that specific oxysterols stimulate osteogenic differentiation of pluripotent bone marrow stromal cells (MSCs) through activation of hedgehog (Hh) signaling and may serve as potential future therapies for intervention in osteopenia and osteoporosis. In this study we report that the osteogenic oxysterol 20(S)-hydroxycholesterol (20S) induces the expression of genes associated with Notch signaling. Using M2-10B4 (M2) MSCs, we found that 20S significantly induced HES-1, HEY-1, and HEY-2 mRNA expression compared with untreated cells, with maximal induction after 48 hours, whereas the nonosteogenic oxysterols did not. Similar observations were made when M2 cells were treated with sonic hedgehog (Shh), and the specific Hh pathway inhibitor cyclopamine blocked 20S-induced Notch target gene expression. 20S did not induce Notch target genes in Smo(-/-) mouse embryonic fibroblasts, further confirming the role of Hh signaling in 20S-induced expression of Notch target genes. Despite the inability of liver X-receptor (LXR) synthetic ligand TO901317 to induce Notch target genes in M2 cells, LXR knockdown studies using siRNA showed inhibition of 20S-induced HEY-1 but not HES-1 expression, suggesting the partial role of LXR signaling in MSC responses to 20S. Moreover, 20S-induced Notch target gene expression was independent of canonical Notch signaling because neither 20S nor Shh induced CBF1 luciferase reporter activity or NICD protein accumulation in the nucleus, which are hallmarks of canonical Notch signaling activation. Finally, HES-1 and HEY-1 siRNA transfection significantly inhibited 20S-induced osteogenic genes, suggesting that the pro-osteogenic effects of 20S are regulated in part by HES-1 and HEY-1.
Subject(s)
Bone Marrow Cells/drug effects , Gene Expression/drug effects , Hydroxycholesterols/pharmacology , Osteogenesis/drug effects , Receptors, Notch/genetics , Animals , Basic Helix-Loop-Helix Transcription Factors/analysis , Basic Helix-Loop-Helix Transcription Factors/metabolism , Bone Marrow Cells/metabolism , Cell Culture Techniques , Cell Cycle Proteins/analysis , Cell Cycle Proteins/metabolism , Fibroblasts/drug effects , Fibroblasts/metabolism , Hedgehog Proteins/metabolism , Homeodomain Proteins/analysis , Homeodomain Proteins/metabolism , Liver X Receptors , Mice , Orphan Nuclear Receptors/analysis , Orphan Nuclear Receptors/metabolism , RNA, Messenger/analysis , RNA, Messenger/metabolism , Stromal Cells/drug effects , Stromal Cells/metabolism , Transcription Factor HES-1 , Veratrum Alkaloids/pharmacologyABSTRACT
Celsius is a data warehousing system to aggregate Affymetrix CEL files and associated metadata. It provides mechanisms for importing, storing, querying, and exporting large volumes of primary and pre-processed microarray data. Celsius contains ten billion assay measurements and affiliated metadata. It is the largest publicly available source of Affymetrix microarray data, and through sheer volume it allows a sophisticated, broad view of transcription that has not previously been possible.
Subject(s)
Database Management Systems , Databases, Genetic , Oligonucleotide Array Sequence Analysis , Animals , Gene Expression Profiling , HumansABSTRACT
Pluripotent mesenchymal cells form a population of precursors to a variety of cell types, including osteoblasts and adipocytes. Aging tilts the balance in favor of adipocyte differentiation at the expense of osteoblast differentiation, resulting in reduced bone formation and osteopenic disorders, including osteoporosis, in humans and animals. Understanding the mechanisms involved in causing this apparent shift in differentiation and identifying factors that stimulate osteoblast formation while inhibiting adipogenesis are of great therapeutic interest. In this study we report that specific, naturally occurring oxysterols, previously shown to direct pluripotent mesenchymal cells toward an osteoblast lineage, exert their osteoinductive effects through activation of Hedgehog signaling pathway. This was demonstrated by 1) oxysterol-induced expression of the Hh target genes Gli-1 and Patched, 2) oxysterol-induced activation of a luciferase reporter driven by a multimerized Gli-responsive element, 3) inhibition of oxysterol effects by the hedgehog pathway inhibitor, cyclopamine, and 4) unresponsiveness of Smoothened-/- mouse embryonic fibroblasts to oxysterols. Using Patched-/- cells that possess high baseline Gli activity, we found that oxysterols did not dramatically shift the IC50 concentration of cyclopamine needed to inhibit Gli activity in these cells. Furthermore, binding studies showed that oxysterols did not compete with fluorescently labeled cyclopamine, BODIPY-cyclopamine, for direct binding to Smoothened. These findings demonstrate that oxysterols stimulate hedgehog pathway activity by indirectly activating the seven-transmembrane pathway component Smoothened. Osteoinductive oxysterols are, therefore, novel activators of the hedgehog pathway in pluripotent mesenchymal cells, and they may be important modulators of this critical signaling pathway that regulates numerous developmental and post-developmental processes.
Subject(s)
Hedgehog Proteins/metabolism , Mesenchymal Stem Cells/metabolism , Sterols/metabolism , Adipocytes/metabolism , Animals , Dose-Response Relationship, Drug , Fibroblasts/metabolism , Mice , Mice, Inbred C3H , Osteoblasts/metabolism , Protein Binding , Protein Kinase C/metabolism , Receptors, G-Protein-Coupled/metabolism , Signal Transduction , Smoothened Receptor , Veratrum Alkaloids/metabolismABSTRACT
Although the development of regulatory T cells (T(reg) cells) in the thymus is defined by expression of the lineage marker Foxp3, the precise function of Foxp3 in T(reg) cell lineage commitment is unknown. Here we examined T(reg) cell development and function in mice with a Foxp3 allele that directs expression of a nonfunctional fusion protein of Foxp3 and enhanced green fluorescent protein (Foxp3DeltaEGFP). Thymocyte development in Foxp3DeltaEGFP male mice and Foxp3DeltaEGFP/+ female mice recapitulated that of wild-type mice. Although mature EGFP(+) CD4(+) T cells from Foxp3DeltaEGFP mice lacked suppressor function, they maintained the characteristic T(reg) cell 'genetic signature' and failed to develop from EGFP(-) CD4(+) T cells when transferred into lymphopenic hosts, indicative of their common ontogeny with T(reg) cells. Our results indicate that T(reg) cell effector function but not lineage commitment requires the expression of functional Foxp3 protein.