Single-Cell Analysis Reveals Regulatory Gene Expression Dynamics Leading to Lineage Commitment in Early T Cell...

Area of Science:

• Immunology
• Developmental Biology
• Cellular and Molecular Biology

Background:

• Intrathymic T cell development involves transitioning multipotent precursors to committed pro-T cells.
• The precise regulatory steps and gene expression dynamics during this process have remained largely undefined.

Purpose of the Study:

• To elucidate the sequential regulatory events during early T cell development.
• To define the order of gene silencing and induction during T cell lineage commitment.
• To investigate the role of multilineage priming in T cell specification.

Main Methods:

• Single-cell RNA sequencing (scRNA-seq) for comprehensive transcriptome analysis.
• Sequential multiplexed single-molecule fluorescent in situ hybridization (seqFISH) for transcript quantification.
• Single-cell cloning to verify precursor populations and developmental trajectories.

Main Results:

• Identified and distinguished early T cell precursor (ETP) subsets and a separate granulocyte precursor subset.
• Confirmed initial co-expression of progenitor and T cell specification genes.
• Established stage-specific relationships between cell cycle progression and differentiation.
• Developed and validated a pseudotime model of T cell lineage commitment using RNA velocity and transcription factor perturbations.

Conclusions:

• Multilineage priming is a fundamental aspect of T cell specification.
• The study provides a detailed regulatory roadmap of intrathymic T cell development.
• This work clarifies the transition from multipotent precursors to committed T cells.