Suoet al., Science 376 , eabo0510 (2022) 3 June 2022 10 of 15
TRAV genesTRAJ genes
clone of
proliferating
DP
transition stage
transition stage
Unconventional
T cell
Conventional
T cell
cortex medulla
nearby
cTEC
positive selection
T–T interaction
positive selection
T–TEC interaction
recombination
B
D E
F G
A
Proportion of scVDJseq cells (%)
Conventional T cells Unconventional T cells
cell density
C
TRAV TRAJ
5’ 3’
V J
** ** ** ******** * ** **
Unconventional
T cells
Conventional
T cells
*
*
Unconventional T cells
Conventional T cells
Gene segment preferentially used in
* *
*** *
TRGV TRGJ TRDV TRDD TRDJ
development
genomic position
iPSC
ATO
T cells
Hematopoietic
induction
(2 weeks)
T cell
differentiation
(7 weeks)
Mesoderm
induction
(3.5 days)
Mouse stromal
cells with
human DLL4
D0 wk5 wk7wk3
PLZF
Fig. 6. Deep characterization of human unconventional T cells.(A) Proportions
of cells expressing pairedgdTCR, pairedabTCR, both, or neither. The
proportions were calculated over cells that had both single-cellabTCR and
gdTCR sequencing. Expression of neither pairedabTCR nor pairedgdTCR in
some cells could be due to dropouts in single-cell TCR sequencing because
50% of these contained orphan VDJ or VJ chains ofabTCR orgdTCR
(fig. S28A). (B) Heatmap showing the percentage of eachgdTCR gene segment
present in different T cell subtypes. Differential usage between cell subtypes
was computed using the chi-squared test. Gene segments with Benjamini-
Hochberg–adjustedP values < 0.05 are marked with asterisks. (C) Heatmap
showing the proportion of each TCRagene segment present in different T cell
subtypes. The gene segment usage in unconventional T cells and conventional
T cells was compared using logistic regression, controlling for donors and
organs. Gene segments with Benjamini-Hochberg–adjustedP values < 0.05
are marked with magenta asterisks for preferential usage in unconventional
T cells and green asterisks for preferential usage in conventional T cells.
(D) Principal component analysis plot summarizing TRAV, TRAJ, TRBV, and
TRBJ gene segment usage proportion in different T cell subtypes. Each dot
represents a sample of at least 20 cells, with dot size representing the cell count.
The centroid of each cell type is shown as a filled circle, and 80% confidence
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