Bulk RNA-seq analysis of WT and tristetraprolin (TTP)-knockout basophils treated with antigen/IgE stimulation
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1137305 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR29877454.fastq.gz | Fastqsanger.gz | |||
| SRR29877455.fastq.gz | Fastqsanger.gz | |||
| SRR29877456.fastq.gz | Fastqsanger.gz | |||
| SRR29877457.fastq.gz | Fastqsanger.gz | |||
| SRR29877458.fastq.gz | Fastqsanger.gz |
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Project description:Basophils play crucial roles in type 2 immune responses, such as chronic allergic inflammation and protective immunity against parasites. However, the molecular mechanisms regulating basophil activation and effector molecule production remain poorly understood. To investigate the role of RNA-binding proteins (RBPs), we first analyzed the gene expression of CCCH zinc finger proteins in antigen/IgE-stimulated basophils. Among these proteins, we identified that Zfp36, encoding tristetraprolin (TTP), was the most significantly upregulated gene. Therefore, we focused on the functions of TTP in basophils. We generated TTP-KO mice using the CRISPR-Cas9 method and conducted bulk RNA-seq analysis of antigen/IgE-stimulated basophils from wild-type (WT) and TTP-knockout (TTP-KO) mice. TTP-KO basophils exhibited elevated mRNA expression of pro-inflammatory mediators, such as Il4, Il13, Areg, Ccl3, and Cxcl2, compared to WT basophils. These data suggest that TTP is a key regulator of basophil activation, controlling the mRNA expression of inflammatory mediators.
2024-11-17 | GSE272532 | GEO
Project description:Basophils play crucial roles in type 2 immune responses, such as chronic allergic inflammation and protective immunity against parasites. However, the molecular mechanisms underlying the regulation of proinflammatory molecule production by basophils remain unclear. To investigate the role of RNA-binding proteins (RBPs), we first analyzed the gene expression of CCCH zinc finger proteins in antigen/IgE-stimulated basophils and identified that Zfp36, encoding tristetraprolin (TTP), was the most significantly upregulated gene encoding a CCCH zinc finger protein. To explore the functions of TTP in basophils, we generated TTP-knockout (TTP-KO) mice using the CRISPR-Cas9 method and conducted bulk RNA-seq analysis of antigen/IgE-stimulated basophils from wild-type (WT) and TTP-KO mice. TTP-KO basophils exhibited elevated mRNA expression of pro-inflammatory mediators, such as Il4, Il13, Areg, Ccl3, and Cxcl2, compared to WT basophils, suggesting the key role of TTP in the expression of mRNAs encoding proinflammatory mediators. Since TTP is known to regulate mRNA stability in other immune cell types, we evaluated the mRNA stability of WT and TTP-KO basophils under antigen/IgE stimulation using SLAM-seq [thiol (SH)-Linked Alkylation for the Metabolic sequencing of RNA] analysis. SLAM-seq analysis revealed that the mRNA half-lives of proinflammatory molecules, such as Il4, Il13, Areg, Ccl3, and Cxcl2, were prolonged by TTP deficiency, suggesting the importance of TTP in regulating the stability of mRNAs encoding inflammatory molecules in basophils.
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Project description:Basophils are the least common granulocytes which represent less than 1% of peripheral blood leukocytes. Recent development of analytical tools for basophils enables us to understand their critical roles in allergic reactions and protection from parasitic infections. Nevertheless, the differentiation trajectory of basophils remains unclear. We have identified previously-unappreciated pre-basophil subpopulation which encompasses previously-defined basophil precursors (BaPs) in a series of experiments including scRNA-seq analysis (deposited to GSE206589). In this study, we explored gene expression profiles of pre-basophils and mature basophils activated by antigen/IgE-stimulation or IL-3-stimulation. We have revealed that IL-3 activated pre-basophils displayed distinct gene expression profiles from antigen/IgE- or IL-3-stimulated mature basophils, suggesting the unuque property of pre-basophils.
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Project description:SLAM-seq analysis of WT and TTP-KO basophils treated with antigen/IgE stimulation
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Project description:Tristetraprolin (TTP) binds to specific AU-rich elements in the 3'UTR of certain transcripts and regulates post-transcriptional gene expression by increasing the rate of mRNA turnover. Here, we generated a novel TTP mouse line (TTPΔARE) with a 136 base deletion in the 3'UTR AU-rich region of TTP mRNA, which resulted in a moderate increase in TTP protein expression in many tissues. In this study, we evaluated the effects of the moderate TTP overexpression on the overall pattern of gene expression in spleen, to investigate the effect of this mutation on mRNA targets of TTP under normal conditions. We utilized "TTPΔARE" (136 b deletion in the 3'UTR AU-rich region of Zfp36 mRNA) mice that overexpress TTP and compared the transcriptomic changes to the littermate "WT" mice. Spleen mRNA from four WT and four "TTPΔARE" mice were subjected to mRNA-Seq. All the animals used in this study were males between the ages of 8-12 weeks and were on a C57BL/6 background.
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Project description:Tristetraprolin (TTP) binds to specific AU-rich elements in the 3'UTR of certain transcripts and regulates post-transcriptional gene expression by increasing the rate of mRNA turnover. In this study, we evaluated the effects of TTP deficiency on the overall gene expression of spleen tissue, in order to discover tissue specific targets of TTP under normal physiologic conditions. We utilized "Triple KO" (Zfp36-/-/TNFR1-/-/TNFR2-/-) mice that are deficient in TTP and two TNF receptors and compared the transcriptomic changes to "Double KO" (TNFR1-/-/TNFR2-/-) and WT mice. Spleen mRNA from four WT, four "Double KO", and four "Triple mice" was subjected to RNA-Seq in two phases. All the animals used in this study were males between the ages of 12-14 weeks and were on a mixed (75% C57BL/6NTac, 25% 129/SVEV) background.
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Project description:Gene expression profiles of pre-basophils versus mature basophils stimulated with antigen/IgE- or IL-3-stimulation.
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Project description:Tristetraprolin (TTP) binds to specific AU-rich elements in the 3'UTR of certain transcripts and regulates post-transcriptional gene expression by increasing the rate of mRNA turnover. In this study, we evaluated the effects of TTP deficiency on the overall gene expression of spleen tissue, in order to discover tissue specific targets of TTP under normal physiologic conditions. We utilized "Triple KO" (Zfp36-/-/TNFR1-/-/TNFR2-/-) mice that are deficient in TTP and two TNF receptors and compared the transcriptomic changes to "Double KO" (TNFR1-/-/TNFR2-/-) and WT mice. Spleen mRNA from four WT, four "Double KO", and four "Triple mice" was subjected to RNA-Seq in two phases. All the animals used in this study were males between the ages of 12-14 weeks and were on a mixed (75% C57BL/6NTac, 25% 129/SVEV) background. Examination of splenic gene expression difference between "Triple KO"-WT and "double KO"-WT data sets
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Project description:Controlled decay of cytokine and chemokine mRNAs restrains the time and amplitude of inflammatory responses. Tristetraprolin (TTP) binds to AU-rich elements in 3´ untranslated regions of mRNA and targets the bound mRNA for degradation. We have addressed here the function of TTP in balancing the macrophage activation state by a comprehensive analysis of TTP-dependent mRNA decay in LPS-stimulated macrophages from WT and TTP-deficient mice.
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