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Alternative polyadenylation releases PCBP1-mediated suppression of CFIm25 during macrophage differentiation
María Del Pilar Mendoza-Martín1, Salwa Mohd Mostafa2, Atish Barua2
1Francisco de Vitoria University, Madrid, Spain.
Abstract:
CFIm25, a key component of the cleavage factor Im (CFIm) complex needed for mRNA 3' end processing, shows increased protein expression during monocyte-to-macrophage differentiation despite stable mRNA levels. We demonstrate that poly(C)-binding protein 1 (PCBP1) suppresses CFIm25 translation in monocytes by binding to its long 3' untranslated region (UTR). During differentiation, alternative polyadenylation generates a shorter CFIm25 3'UTR lacking PCBP1 binding sites. RNA immunoprecipitation confirms PCBP1 binding to the long 3'UTR, while ribosome association analysis shows enhanced ribosome recruitment upon PCBP1 depletion. PCBP1 knockdown increases CFIm25 protein in undifferentiated cells and induces macrophage differentiation markers without stimulation. These findings reveal how alternative polyadenylation controls CFIm25 expression during immune cell differentiation by modulating RNA-binding protein interactions and provide insight into post-transcriptional regulation of RNA processing factors. Impact statement This work reveals how a key regulator of mRNA processing is itself controlled through a previously uncharacterized mechanism during immune cell differentiation. Our findings provide insights into the molecular circuits governing macrophage development and identify potential therapeutic targets for inflammatory disorders where myeloid cell differentiation is dysregulated.
Insights
Poly(C)-binding protein 1 (PCBP1) suppresses CFIm25 translation via its 3' untranslated region (UTR). Alternative polyadenylation shortens the CFIm25 3'UTR, enabling increased protein expression during macrophage differentiation.
Area of Science:
- Molecular Biology
- Cell Biology
- Immunology
Background:
- CFIm25 is crucial for mRNA 3' end processing and its protein levels rise during monocyte-to-macrophage differentiation.
- This increase occurs despite stable CFIm25 mRNA levels, suggesting post-transcriptional regulation.
Purpose of the Study:
- To investigate the mechanism controlling CFIm25 protein expression during immune cell differentiation.
- To identify the role of RNA-binding proteins and 3' UTRs in regulating CFIm25 translation.
Main Methods:
- RNA immunoprecipitation (RIP) to detect PCBP1 binding to the CFIm25 3' UTR.
- Ribosome association analysis to assess translation efficiency.
- Analysis of alternative polyadenylation and its impact on 3' UTR length.
- PCBP1 knockdown experiments.
Main Results:
- PCBP1 binds to the long 3' UTR of CFIm25, suppressing its translation in monocytes.
- Alternative polyadenylation generates a shorter 3' UTR lacking PCBP1 binding sites during differentiation.
- PCBP1 depletion increases CFIm25 protein levels and promotes macrophage differentiation markers.
Conclusions:
- Alternative polyadenylation of the CFIm25 3' UTR is a key regulatory mechanism controlling its protein expression during immune cell differentiation.
- This process involves modulating RNA-binding protein interactions (PCBP1) and impacts macrophage development.
- Findings offer insights into post-transcriptional regulation and potential therapeutic targets for inflammatory disorders.
