Single-molecule imaging of transcription dynamics, RNA localization and fate in human T cells
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Abstract
T cells are critical effector cells counteracting infections and malignancies. To achieve this, they produce pro-inflammatory cytokines, including IFN-γ and TNF. Cytokine production is a tightly regulated process, but the relative contribution of transcriptional and post-transcriptional regulation to mRNA expression remains unknown. We optimized single-molecule FISH for primary human T cells (T-cell smFISH) to simultaneously quantify nascent RNA, levels of mature mRNA, and its localization with single-cell resolution. T-cell smFISH uncovered heterogeneous cytokine mRNA levels, with high cytokine producers displaying biallelic IFNG / TNF RNA transcription activity. Throughout activation, nuclear cytokine mRNAs accumulated, whereas cytoplasmic cytokine mRNA was degraded through translation-dependent decay. Lastly, T-cell smFISH uncovered cytokine-specific regulation by the RNA-binding protein HuR. Thus, T-cell smFISH provides novel insights in the intricate (post)-transcriptional processes in T cells.
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- Authors
- Lattanzio, M. V., Šoštarić, N., Kanagasabesan, N., Popović, B., Bradarić, A., Wardak, L., Guislain, A., Savakis, P., Tutucci, E., & Wolkers, M. C.
- Affiliations
- Sanquin (NL)
- Vrije Universiteit Amsterdam (NL)
- Corresponding author
- Tutucci, Evelina
- Wolkers, Monika C
- ORCID
- Venue
- The EMBO Journal
- Published
- 2025-10-14
- Volume
- 44
- Issue
- 22
- Pages
- 6732-6749
- ISSN
- 0261-4189 (ISSN-L)
- 1460-2075
- PMC
- PMC12624010
- OpenAlex
- W4415141514
- Cited by
- 3
- References
- 86
- Citations by year
- 2026: 3
- FWCI
- 0.90
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Locations
- The EMBO Journal (publishedVersion, cc-by)
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc)
- Europe PMC (PubMed Central) (publishedVersion, cc-by)
- PubMed (publishedVersion)
- Pure Amsterdam UMC (publishedVersion, other-oa)
- VU Research Portal (publishedVersion, cc-by)
- Funders
- EC | European Research Council (817533)
- Oncode Institute (MCW)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Research and Splicing
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Single-cell and spatial transcriptomics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
- Keywords
- T cells, single-molecule FISH, nascent RNA, cytokine mRNA, interferon-gamma, TNF-α, transcriptional regulation, post-transcriptional regulation, mRNA localization, HuR, cytokine gene expression, T cell activation
- MeSH terms
- ELAV-Like Protein 1, Single Molecule Imaging, Gene Expression Regulation, Humans, Interferon-gamma, RNA, Messenger, T-Lymphocytes, Transcription, Genetic, Tumor Necrosis Factor-alpha, Cytokines, In Situ Hybridization, Fluorescence
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
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