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| Status |
Public on Jun 17, 2024 |
| Title |
STARR-seq library profiling 100 defined open chromatin regions (‘focused library’) in livers of male and female mice treated with the nuclear receptor CAR (Nr1i3) agonist ligand TCPOBOP. |
| Organisms |
Mus musculus; synthetic construct |
| Experiment type |
Other
|
| Summary |
Massively parallel reporter assays are widely used to discover functional enhancers but have largely been limited to transfected cell models, which are confounded by vector-induced innate immune responses and lack the physiologically relevant cellular and endogenous hormonal context and chromatin environment of complex mammalian tissues. Here, we combine hydrodynamic injection with a modified STARR-seq-based MPRA to determine condition-specific enhancer activity in mouse liver at scale. Strong liver enhancer activity was observed with STARR-seq libraries containing an Albumin minimal promoter but not when using a Super Core promoter or an origin of replication promoter. We prepared a focused STARR-seq library comprised of 100 PCR-amplified open chromatin regions nearby genes showing sex-biased expression or responsiveness to TCPOBOP, a xenobiotic and agonist ligand of the nuclear receptor CAR (Nr1i3). We assayed STARR-seq activity for the 100 genomic regions in male liver, in female liver and in TCPOBOP-treated male liver to quantitatively measure their intrinsic transcriptional activity under the 3 indicated biological conditions, and thereby identified enhancers whose activity is sex-dependent or xenobiotic-responsive.
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| Overall design |
The focused plasmid library was delivered to mouse liver by hydrodynamic tail vein injection (HDI). Three types of Illumina sequencing libraries were prepared and are included in this GEO submission: the original STARR-seq plasmid libraries used for HDI, STARR-seq DNA libraries extracted from the transfected livers, and STARR-seq RNA libraries prepared from STARR-seq reporter RNA extracted from each transfected liver. These libraries were all generated by PCR amplification carried out in parallel and under the same PCR conditions. The STARR-seq plasmid library is the same material used for HDI and indicates the relative abundance of each reporter sequence mapping to the 100 open chromatin (DNase I-hypersensitive; DHS) regions used for library preparation. The STARR-seq DNA plasmid library was prepared using DNA reporter-specific primers that amplify plasmid DNA extracted from the transfected livers. STARR-seq enhancer activity was calculated by dividing the Illumina sequencing read counts of each RNA reporter (numerator) by that of the corresponding STARR-seq plasmid or DNA library sequence reads (denominator). The STARR-seq RNA libraries were prepared from poly-adenylated RNA extracted from the HDI livers were reverse-transcribed followed by DNase treatment, and then amplified using RNA reporter-specific primers; these RNA libraries represent the self-transcribing activity of each DHS region included in the STARR-seq plasmid library.
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| Contributor(s) |
Chang T, Waxman DJ |
| Citation(s) |
38915578, 38978599 |
| NIH grant(s) |
| Grant ID |
Grant title |
Affiliation |
Name |
| R01 DK121998 |
Growth Hormone Regulation of Sex Differences in Liver Metabolism |
BOSTON UNIVERSITY CHARLES RIVER CAMPUS |
David J Waxman |
| R01 ES024421 |
Xenobiotic-responsive hepatic long non-coding RNAs |
BOSTON UNIVERSITY CHARLES RIVER CAMPUS |
David J Waxman |
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| Submission date |
May 08, 2024 |
| Last update date |
Aug 08, 2024 |
| Contact name |
David J. Waxman |
| E-mail(s) |
djw@bu.edu
|
| Organization name |
Boston University
|
| Department |
Department of Biology and Bioinformatics Program
|
| Street address |
5 Cummington Mall
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02215 |
| Country |
USA |
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| Platforms (2) |
| GPL17021 |
Illumina HiSeq 2500 (Mus musculus) |
| GPL19604 |
Illumina HiSeq 2500 (synthetic construct) |
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| Samples (12)
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| GSM8258882 |
TCPOBOP_1d_Male_STARRTYC3/7_RNA_replicate1 |
| GSM8258883 |
TCPOBOP_1d_Male_STARRTYC3/7_RNA_replicate2 |
| GSM8258884 |
corn oil_1d_Male_STARRTYC3/7_RNA_replicate1 |
| GSM8258885 |
corn oil_1d_Male_STARRTYC3/7_RNA_replicate2 |
| GSM8258886 |
corn oil_1d_Female_STARRTYC3/7_RNA_replicate1 |
| GSM8258887 |
corn oil_1d_Female_STARRTYC3/7_RNA_replicate2 |
| GSM8258888 |
corn oil_1d_Female_STARRTYC3/7_RNA_replicate3 |
| GSM8258889 |
corn oil_1d_Female_STARRTYC3/7_RNA_replicate4 |
| GSM8258890 |
TCPOBOP_1d_Male_STARRTYC3/7_DNA_replicate1 |
| GSM8258891 |
corn oil_1d_Male_STARRTYC3/7_DNA_replicate1 |
| GSM8258892 |
corn oil_1d_Female_STARRTYC3/7_DNA_replicate1 |
| GSM8258893 |
STARRTYC3/7_plasmid_replicate1 |
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| This SubSeries is part of SuperSeries: |
| GSE267205 |
STARR-seq reporter activity of plasmid libraries delivered to mouse liver by hydrodynamic injection: In vivo MPRA assay for enhancer activity |
|
| Relations |
| BioProject |
PRJNA1109392 |
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