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Integrative scATAC-seq and scRNA-seq analyses map thymic iNKT cell development and identify Cbfβ for its commitment.

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Summary

Invariant natural killer T (iNKT) cells differentiate into subsets within the thymus. This study identifies novel transcription factor Cbfβ as crucial for iNKT cell commitment and differentiation.

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Area of Science:

  • Immunology
  • Developmental Biology
  • Transcriptomics

Background:

  • Invariant natural killer T (iNKT) cells are crucial immune cells that undergo terminal differentiation in the thymus.
  • The molecular mechanisms governing the differentiation of iNKT cell subsets (iNKT1, iNKT2, iNKT17) remain largely undefined.

Purpose of the Study:

  • To elucidate the developmental trajectories and molecular programs guiding invariant natural killer T (iNKT) cell subset differentiation in the thymus.
  • To identify novel regulators of early iNKT cell commitment and effector function.

Main Methods:

  • Single-cell RNA sequencing (scRNA-seq) of over 17,000 iNKT cells.
  • Single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) of over 39,000 iNKT cells across four thymic developmental stages.
  • Conditional gene deletion to assess the function of identified transcription factors.

Main Results:

  • iNKT2 and iNKT17 lineage commitment may initiate as early as Stage 0 (ST0) via distinct molecular programs.
  • iNKT1 commitment appears to occur after ST0, with significant heterogeneity observed in iNKT1 and iNKT2 cells, unlike more homogenous iNKT17 cells.
  • A novel transcription factor, Cbfβ, was identified as highly expressed during iNKT progenitor commitment and is essential for iNKT cell development and differentiation.

Conclusions:

  • Distinct molecular programs govern iNKT cell subset differentiation, with early commitment for iNKT2/iNKT17 lineages.
  • Cellular heterogeneity exists within iNKT1 and iNKT2 subsets, while iNKT17 cells are more uniform.
  • The transcription factor Cbfβ is a key regulator of early iNKT cell commitment and effector phenotype development.