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SRP039970
Mus musculus
6 Downloadable Samples
Illumina HiSeq 2500
Description
Neuroanatomical methods enable high-resolution mapping of neural circuitry, but do not allow systematic molecular profiling of neurons based on their connectivity. Here, we report the development of a novel approach for molecularly profiling projective neurons. We show that ribosomes can be labeled with a camelid nanobody raised against GFP and that this system can be engineered to selectively capture translating mRNAs from cells expressing GFP. We generated a transgenic mouse encoding a nanobody-ribosomal protein fusion (Syn-NBL10) and used a retrograde virus (CAV) encoding GFP to immunoprecipitate ribosomes from projection neurons. This enabled us to profile neurons projecting to the nucleus accumbens. The current method provides a new means for profiling neurons based on their projections. Overall design: Translating mRNAs immunoprecipitated from neurons projecting to the nucleus accumbens. Each Input and IP sample corrspond to a pooled group of 6 mice.
Publication Title
Molecular profiling of neurons based on connectivity.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 10 Downloadable Samples
- Illumina HiSeq 2500
Description
Hunger, driven by negative energy balance, elicits the search for and consumption of food. In mammals, this is orchestrated principally through the activity of neurons in the hypothalamus, direct manipulation of which can potently drive food intake. However, the neural circuits outside of the hypothalamus that control feeding are poorly understood. Here, we identify two functionally opponent cell types within the dorsal raphe nucleus (DRN), marked by the vesicular transporters for GABA (Vgat) or glutamate (VGLUT3), that project to many known feeding centers and rapidly control feeding. We find that DRNVgat neurons drive, while DRNVGLUT3 neurons suppress, food intake. Furthermore, through the development and application of cell type-specific molecular profiling technologies, we identify many differentially expressed transmembrane receptors, which may represent unique druggable targets. Local application of agonists for these receptors potently modulates feeding, recapitulating the effects of cell-specific manipulations. Together, these data establish a key role for the DRN in controlling food intake and add an important anatomic site that controls energy balance. Overall design: Paired - Inputs and IPs; Unpaired for Vgat/VGLUT3 comparison
Publication Title
Identification of a Brainstem Circuit Controlling Feeding.
Sample Metadata Fields
Specimen part, Subject
View Samples- Mus musculus
- 28 Downloadable Samples
- Illumina HiSeq 2500
Description
Translational profiling methodologies enable the systematic characterization of cell types in complex tissues such as the mammalian brain, where neuronal isolation is exceptionally difficult. Here, we report a versatile strategy to profile CNS cell types in a spatiotemporally-restricted fashion by engineering a Cre-dependent adeno-associated virus expressing an EGFP-tagged ribosomal protein (AAV-FLEX-EGFPL10a) to access translating mRNAs by TRAP. We demonstrate the utility of this AAV to target a variety of genetically and anatomically defined neural populations expressing Cre recombinase and illustrate the ability of this viral TRAP (vTRAP) approach to recapitulate the molecular profiles obtained by bacTRAP in corticothalamic neurons across multiple serotypes. Furthermore, spatially restricting AAV injections enabled the elucidation of regional differences in gene expression within this cell type. Taken together, these results establish the broad applicability of the vTRAP strategy for the molecular dissection of any CNS or peripheral cell type that can be engineered to express Cre. Overall design: Polysome-bound mRNAs from TRAP IPs were compared to whole tissue mRNAs. Data was collected from MCH neurons in hypothalamus using vTRAP, cortical layer 6 Ntsr1 neurons using vTRAP, and cortical layer 6 Ntsr1 neurons using bacTRAP. We include vTRAP data from three AAV serotypes for the cortical Ntsr1 cells. We collected three replicates for IP and inputs for vTRAP experiments, while bacTRAP data was collected in duplicate.
Publication Title
Rapid Molecular Profiling of Defined Cell Types Using Viral TRAP.
Sample Metadata Fields
Specimen part, Subject
View Samples- Homo sapiens
- 43 Downloadable Samples
- Illumina HiSeq 2000
Description
Hepatitis C virus (HCV) is widely used to investigate host-virus interactions and cellular responses to infection have been extensively studied in vitro. In human liver, interferon (IFN) stimulated gene expression can mask direct transcriptional responses to virus infection. To better characterize the direct effects of HCV infection in vivo, we analyze the transcriptomes of HCV-infected patients lacking an activated endogenous IFN system. We show that the expression changes observed in these patients predominantly reflect immune cell infiltrates rather than changes in cell-intrinsic metabolic pathways. We also investigate the transcriptomes of patients with endogenous IFN activation, which paradoxically cannot eradicate viral infection. We find that most IFN-stimulated genes (ISGs) are induced by both the endogenous IFN system and by recombinant IFN therapy, but with significantly higher induction levels in the latter. We conclude that the innate host immune response in chronic hepatitis C is too weak to clear the virus. Overall design: In this study, we aimed to disentangle the direct and indirect effects of HCV infection on cellular transcriptional profiles, by performing a detailed characterization of the gene expression changes associated with HCV infection, endogenous IFN system activation and pegIFNa treatment in the human liver. With this objective, we generated and analyzed high-throughput transcriptome sequencing profiles from liver biopsies derived from different categories of HCV-infected and non-infected patients, prior to and during treatment. First, to unveil HCV-induced cell-autonomous effects and to separate them from IFN-induced changes in the transcriptome, we selected liver biopsies from patients with chronic hepatitis C (CHC) without hepatic ISG induction, and compared them with un-infected control biopsies. Second, we examined the transcriptomic changes associated with the endogenous activation of the IFN system. Finally, we analyzed the gene expression changes resulting from pegIFNa/ribavirin treatment, by comparing transcriptome data from liver biopsies obtained before treatment and at different time points during the first week of therapy.
Publication Title
Transcriptional response to hepatitis C virus infection and interferon-alpha treatment in the human liver.
Sample Metadata Fields
Specimen part, Treatment, Subject
View Samples- Mus musculus
- 24 Downloadable Samples
- Illumina HiSeq 2500
Description
Despite decades of interest, the mechanisms that control Hox gene expression are not yet fully understood. It was recently proposed that Hotair, a lncRNA transcribed from the HoxC cluster, regulates HoxD gene expression via Polycomb targeting and thus is important for correct skeletal development. However, genetic manipulations of the locus led to conflicting results regarding the roles of Hotair. Here, we analyze the molecular and phenotypic consequences of deleting the Hotair locus in vivo. In contradiction with previous findings, we show that deleting Hotair has no detectable effect on HoxD gene expression in vivo. We could not observe any morphological alteration in mice lacking the Hotair locus. However, we find a significant impact of deleting Hotair on the expression of neighboring genes Hoxc11 and Hoxc12. Our results do not support an RNA-dependent role for Hotair in vivo, but argue in favor of a DNA-dependent effect of Hotair deletion on the transcriptional landscape in cis. Overall design: We micro-dissected wild type and Del(Hotair)-/- E12.5 embryos into 6 segments: forelimbs (FL), hindlimbs (HL), genital tubercle (GT), trunk section corresponding to the lumbar/sacral region (T1); trunk section corresponding to the sacral/caudal region (T2) and trunk section corresponding to the caudal region (T3). We generated strand-specific RNA-seq data for each segment, in two biological replicates and we performed differential expression analyses for each tissue. Furthermore, we analyzed the impact of deleting the Hotair locus on the local transcriptional landscape, in the HoxC cluster.
Publication Title
Hotair Is Dispensible for Mouse Development.
Sample Metadata Fields
Specimen part, Cell line, Subject
View Samples- Mus musculus
- 4 Downloadable Samples
- Illumina HiSeq 2500
Description
In this work we have analyzed the transcriptomic profiles of E9 mouse embryos. We show that Hoxd1 and Haglr transcripts are absent after targeted deletion of the CpG: 114 island. Overall design: RNA-seq analysis of trunk from the anterior limit of the forelimb bud to the tailbud, aiming to exclude all extra-embryonic, head, cervical and heart tissues. Individuals 443 (wt) and 445 (Del(CpG114) homozygous), were siblings from the same dam, while biological replicates 456 (wt) and 455 (Del(CpG114) homozygous) were siblings from another dam.
Publication Title
Control of growth and gut maturation by <i>HoxD</i> genes and the associated lncRNA <i>Haglr</i>.
Sample Metadata Fields
Specimen part, Cell line, Subject
View Samples- Mus musculus
- 4 Downloadable Samples
- Illumina HiSeq 2500
Description
During vertebrate limb development, Hoxd genes are regulated following a bimodal strategy involving two topologically associating domains (TADs) located on either side of the gene cluster. These regulatory landscapes alternatively control different subsets of Hoxd targets, first into the arm and, subsequently, into the digits. We studied the transition between these two global regulations, a switch that correlates with the positioning of the wrist, which articulates these two main limb segments. We show that the HOX13 proteins themselves help switch off the telomeric TAD, likely through a global repressive mechanism. At the same time, they directly interact with distal enhancers to sustain the activity of the centromeric TAD, thus explaining both the sequential and exclusive operating processes of these two regulatory domains. We propose a model whereby the activation of Hox13 gene expression in distal limb cells both interrupts the proximal Hox gene regulation and re-enforces the distal regulation. In the absence of HOX13 proteins, a proximal limb structure grows without any sign of wrist articulation, likely related to an ancestral fish-like condition. Overall design: RNA-seq analysis of proximal and distal forelimbs from E12.5 wt or Hoxa13-/-;Hoxd13-/- mutant embryos
Publication Title
A role for HOX13 proteins in the regulatory switch between TADs at the HoxD locus.
Sample Metadata Fields
Specimen part, Cell line, Subject
View Samples- Mus musculus
- 6 Downloadable Samples
- Illumina HiSeq 2500
Description
In this work we have analyzed the transcriptomic profiles of E13.5 mouse embryonic mammary buds. We show that Hoxd8 and Hoxd9, two gene members of the HoxD cluster, are transcribed during mammary bud development. Yet, unlike in other developmental contexts, their co-expression does not rely upon the same regulatory mechanism. Hoxd8 is regulated by the combined activity of closely located sequences and the most distant telomeric gene desert. On the other hand, Hoxd9 is controlled by an enhancer sequence also located within the telomeric gene desert, but which has no impact on Hoxd8 transcription, thus constituting an exception to the global regulations systematically observed at this locus. The latter DNA region is also involved in Hoxd gene regulation in other contexts and strongly interacts with Hoxd9 in all tissues analyzed so far as well as in other vertebrate species, indicating that its regulatory activity was already operational before the appearance of mammary glands. Within this DNA region and neighboring the CS39 limb enhancer, we further identified a short sequence conserved in therian mammals and capable of enhancer activity in the mammary buds. We propose that Hoxd gene regulation in embryonic mammary buds evolved by hijacking a preexisting regulatory landscape, which was already at work before the emergence of mammals in structures such like the limbs or the intestinal tract. Overall design: RNA-seq analysis of e13.5 mammary buds and adjacent embryonic skin
Publication Title
Control of Hoxd gene transcription in the mammary bud by hijacking a preexisting regulatory landscape.
Sample Metadata Fields
Specimen part, Cell line, Subject
View Samples- Mus musculus
- 6 Downloadable Samples
Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
Postnatal neural progenitors of the enteric nervous system are a potential source for future cell replacement therapies of developmental dysplasia like Hirschsprung's disease. However, little is known about the molecular mechanisms driving the homeostasis and differentiation of this cell pool. In this work, we conducted Affymetrix gene chip experiments to identify differences in gene regulation between proliferation and early differentiation of enteric neural progenitors. We detected a total of 1333 regulated genes that were linked to different groups of cellular mechanisms involved in cell cycle, apoptosis, neural proliferation, and differentiation. As expected, we found a strong inhibition of cell cycle progression as well as an enhanced expression of neuronal and glial markers. We further found a marked inactivation of the canonical Wnt pathway during the beginning of cellular differentiation. Taken together, this data illustrated the various mechanisms taking place during the proliferation and early differentiation of enteric neural progenitor cells.
Publication Title
Comparative Microarray Analysis of Proliferating and Differentiating Murine ENS Progenitor Cells.
Sample Metadata Fields
Specimen part
View Samples- Drosophila melanogaster
- 9 Downloadable Samples
- Affymetrix Drosophila Genome 2.0 Array (drosophila2)
Description
To determine the physiological targets of the NELF complex, and provide insight into the mechanism of NELF activity in vivo.
Publication Title
NELF-mediated stalling of Pol II can enhance gene expression by blocking promoter-proximal nucleosome assembly.
Sample Metadata Fields
No sample metadata fields
View Samples