HDI-STARR-seq: Condition-specific enhancer discovery in mouse liver in vivo
Abstract Background STARR-seq and other massively-parallel reporter assays are widely used to discover functional enhancers in transfected cell models, which can be confounded by plasmid vector-induced type-I interferon immune responses and lack the multicellular environment and endogenous chromatin...
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2024-12-01
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| Online Access: | https://doi.org/10.1186/s12864-024-11162-9 |
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| author | Ting-Ya Chang David J. Waxman |
| author_facet | Ting-Ya Chang David J. Waxman |
| author_sort | Ting-Ya Chang |
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| description | Abstract Background STARR-seq and other massively-parallel reporter assays are widely used to discover functional enhancers in transfected cell models, which can be confounded by plasmid vector-induced type-I interferon immune responses and lack the multicellular environment and endogenous chromatin state of complex mammalian tissues. Results We describe HDI-STARR-seq, which combines STARR-seq plasmid library delivery to the liver, by hydrodynamic tail vein injection (HDI), with reporter RNA transcriptional initiation driven by a minimal Albumin promoter, which we show is essential for mouse liver STARR-seq enhancer activity assayed 7 days after HDI. Importantly, little or no vector-induced innate type-I interferon responses were observed. Comparisons of HDI-STARR-seq activity between male and female mouse livers and in livers from males treated with an activating ligand of the transcription factor (TF) CAR (Nr1i3) identified many condition-dependent enhancers linked to condition-specific gene expression. Further, thousands of active liver enhancers were identified using a high complexity STARR-seq library comprised of ~ 50,000 genomic regions released by DNase-I digestion of mouse liver nuclei. When compared to stringently inactive library sequences, the active enhancer sequences identified were highly enriched for liver open chromatin regions with activating histone marks (H3K27ac, H3K4me1, H3K4me3), were significantly closer to gene transcriptional start sites, and were significantly depleted of repressive (H3K27me3, H3K9me3) and transcribed region histone marks (H3K36me3). Conclusion HDI-STARR-seq offers substantial improvements over current methodologies for large scale, functional profiling of enhancers, including condition-dependent enhancers, in liver tissue in vivo, and can be adapted to characterize enhancer activities in a variety of species and tissues by selecting suitable tissue- and species-specific promoter sequences. |
| format | Article |
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| institution | DOAJ |
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| language | English |
| publishDate | 2024-12-01 |
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| spelling | doaj-art-9f060d711c0d40f586c2e3c7cfee68d42025-08-20T02:43:35ZengBMCBMC Genomics1471-21642024-12-0125112910.1186/s12864-024-11162-9HDI-STARR-seq: Condition-specific enhancer discovery in mouse liver in vivoTing-Ya Chang0David J. Waxman1Departments of Biology and Biomedical Engineering, and Bioinformatics Program, Boston UniversityDepartments of Biology and Biomedical Engineering, and Bioinformatics Program, Boston UniversityAbstract Background STARR-seq and other massively-parallel reporter assays are widely used to discover functional enhancers in transfected cell models, which can be confounded by plasmid vector-induced type-I interferon immune responses and lack the multicellular environment and endogenous chromatin state of complex mammalian tissues. Results We describe HDI-STARR-seq, which combines STARR-seq plasmid library delivery to the liver, by hydrodynamic tail vein injection (HDI), with reporter RNA transcriptional initiation driven by a minimal Albumin promoter, which we show is essential for mouse liver STARR-seq enhancer activity assayed 7 days after HDI. Importantly, little or no vector-induced innate type-I interferon responses were observed. Comparisons of HDI-STARR-seq activity between male and female mouse livers and in livers from males treated with an activating ligand of the transcription factor (TF) CAR (Nr1i3) identified many condition-dependent enhancers linked to condition-specific gene expression. Further, thousands of active liver enhancers were identified using a high complexity STARR-seq library comprised of ~ 50,000 genomic regions released by DNase-I digestion of mouse liver nuclei. When compared to stringently inactive library sequences, the active enhancer sequences identified were highly enriched for liver open chromatin regions with activating histone marks (H3K27ac, H3K4me1, H3K4me3), were significantly closer to gene transcriptional start sites, and were significantly depleted of repressive (H3K27me3, H3K9me3) and transcribed region histone marks (H3K36me3). Conclusion HDI-STARR-seq offers substantial improvements over current methodologies for large scale, functional profiling of enhancers, including condition-dependent enhancers, in liver tissue in vivo, and can be adapted to characterize enhancer activities in a variety of species and tissues by selecting suitable tissue- and species-specific promoter sequences.https://doi.org/10.1186/s12864-024-11162-9MPRADNase-seqChromatin accessibilityHydrodynamic tail vein injection |
| spellingShingle | Ting-Ya Chang David J. Waxman HDI-STARR-seq: Condition-specific enhancer discovery in mouse liver in vivo BMC Genomics MPRA DNase-seq Chromatin accessibility Hydrodynamic tail vein injection |
| title | HDI-STARR-seq: Condition-specific enhancer discovery in mouse liver in vivo |
| title_full | HDI-STARR-seq: Condition-specific enhancer discovery in mouse liver in vivo |
| title_fullStr | HDI-STARR-seq: Condition-specific enhancer discovery in mouse liver in vivo |
| title_full_unstemmed | HDI-STARR-seq: Condition-specific enhancer discovery in mouse liver in vivo |
| title_short | HDI-STARR-seq: Condition-specific enhancer discovery in mouse liver in vivo |
| title_sort | hdi starr seq condition specific enhancer discovery in mouse liver in vivo |
| topic | MPRA DNase-seq Chromatin accessibility Hydrodynamic tail vein injection |
| url | https://doi.org/10.1186/s12864-024-11162-9 |
| work_keys_str_mv | AT tingyachang hdistarrseqconditionspecificenhancerdiscoveryinmouseliverinvivo AT davidjwaxman hdistarrseqconditionspecificenhancerdiscoveryinmouseliverinvivo |