WASp Deficiency Selectively Affects the TCR Diversity of Different Memory T Cell Subsets in WAS Chimeric Mice

Background: The T cell receptor (TCR) diversity is essential for effective T cell immunity. Previous studies showed that TCR diversity in Wiskott-Aldrich Syndrome (WAS) patients was severely impaired, especially in the memory T cell populations. Whether this defect was caused by intrinsic WASp deficiency or extrinsic reasons is still unclear.

Methods: We sorted different T cell subsets from the bone marrow chimeric mice model using both magnetic beads and flow cytometry. TCR repertoires of memory T cells, especially CD4⁺ effector memory T (TEM) cells and CD8⁺ central memory T (TCM) cells, were analyzed using the UMI quantitative high-throughput sequencing (HTS).

Results: An average of 5.51 million sequencing reads of 32 samples was obtained from the Illumina sequencing platform. Bioinformatic analyses showed that compared with wild type (WT), WAS knock out (KO)-CD4⁺ TEM cells exhibited increased Simpson index and decreased D50 index (P <0.05); The rank abundance curve of KO-CD4⁺ TEM cells was shorter and steeper than that of WT, and the angle of ^qD and q in KO-CD4⁺ TEM cells was lower than that of WT, while these indexes showed few changes between WT and KO chimeric mice in the CD8⁺TCM population. Therefore, it indicated that the restriction on the TCRVβ repertoires is majorly in KO-CD4⁺ TEM cells but not KO- CD8⁺ TCM cells. Principal Component Analysis (PCA), a comprehensive parameter for TCRVβ diversity, successfully segregated CD4⁺ TEM cells from WT and KO, but failed in CD8⁺ TCM cells. Among the total sequences of TRB, the usage of TRBV12.2, TRBV30, TRBV31, TRBV4, TRBD1, TRBD2, TRBJ1.1, and TRBJ1.4 showed a significant difference between WT-CD4⁺ TEM cells and KO-CD4⁺ TEM cells (P <0.05), while in CD8⁺ TCM cells, only the usage of TRBV12.2 and TRBV20 showed a substantial difference between WT and KO (P <0.05). No significant differences in the hydrophobicity and sequence length of TCRVβ were found between the WT and KO groups.

Conclusion: WASp deficiency selectively affected the TCR diversity of different memory T cell subsets, and it had more impact on the TCRVβ diversity of CD4⁺ TEM cells than CD8⁺ TCM cells. Moreover, the limitation of TCRVβ diversity of CD4⁺ TEM cells and CD8⁺ TCM cells in WAS was not severe but intrinsic.