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Chromatin Immunoprecipitation ChIP in Mouse T-cell Lines
Published on: June 17, 2017
Single-Cell Mononucleotide Microsatellite Analysis Reveals Differential Insertion-Deletion Dynamics in Mouse T Cells
Elli-Mari Aska1, Bulat Zagidullin1,2, Esa Pitkänen2,3,4
1Research Program in Systems Oncology, Faculty of Medicine, University of Helsinki, Helsinki, Finland.
Abstract:
Microsatellite sequences are particularly prone to slippage during DNA replication, forming insertion-deletion loops that, if left unrepaired, result in de novo mutations (expansions or contractions of the repeat array). Mismatch repair (MMR) is a critical DNA repair mechanism that corrects these insertion-deletion loops, thereby maintaining microsatellite stability. MMR deficiency gives rise to the molecular phenotype known as microsatellite instability (MSI). By sequencing MMR-proficient and -deficient (Mlh1 and Mlh1 ) single-cell exomes from mouse T cells, we reveal here several previously unrecognized features of in vivo MSI. Specifically, mutational dynamics of insertions and deletions were different on multiple levels. Factors that associated with propensity of mononucleotide microsatellites to insertions versus deletions were: microsatellite length, nucleotide composition of the mononucleotide tract, gene length and transcriptional status, as well replication timing. Here, we show on a single-cell level that deletions - the predominant MSI type in MMR-deficient cells - are preferentially associated with longer A/T tracts, long or transcribed genes and later-replicating genes.
Insights
Mismatch repair deficiency causes microsatellite instability (MSI). This study reveals deletions are the main MSI type in deficient cells, linked to longer A/T tracts and transcribed genes.
Area of Science:
- Genetics
- Molecular Biology
- Genomics
Background:
- Microsatellites are prone to replication slippage, causing mutations if unrepaired.
- Mismatch repair (MMR) corrects these errors, maintaining microsatellite stability.
- MMR deficiency leads to microsatellite instability (MSI).
Purpose of the Study:
- To investigate the previously unrecognized features of in vivo MSI.
- To analyze mutational dynamics of insertions and deletions in MMR-proficient and -deficient cells.
- To identify factors influencing MSI type and location.
Main Methods:
- Sequencing of single-cell exomes from mouse T cells.
- Comparison of MMR-proficient and MMR-deficient (Mlh1-/-) cells.
- Analysis of microsatellite length, nucleotide composition, gene features, and replication timing.
Main Results:
- Deletions are the predominant MSI type in MMR-deficient cells.
- Deletions are preferentially associated with longer A/T tracts.
- Deletions correlate with long or transcribed genes and later replication timing.
Conclusions:
- MMR deficiency exhibits distinct mutational dynamics for insertions and deletions.
- Specific sequence and genomic features influence the propensity for deletions in MSI.
- Single-cell analysis provides novel insights into in vivo MSI mechanisms.
