Fig 1.
STAR mice display normal immune reconstitution with diverse TCRβ chain usage.
A) Overview of method for making STAR mice. B) Representative flow cytometry plots showing CD4 vs CD8, CD4+Foxp3- and CD4+Foxp3+, B cell, and APC cell distribution in STAR mice. C) Pie charts displaying the diversity of TCRVβ and TCRJβ usage in Tconv (left) and Tregs (right). Each color represents a different gene segment and the size of the section is proportional to the frequency of that gene segment.
Fig 2.
TCR sharing by CD4+Foxp3- (Tconv) and CD4+Foxp3+ (Treg) cells.
A) Tabulated results displaying the number of individual TCRβ chain amino acid sequences identified in each mouse, and the number and percent of TCRs shared between Tconv and Tregs within each mouse. B-C) Graphical representations of the percentage of TCR sequences shared (left) and TCR repertoire (right) overlap between Tconv and Tregs within individual STAR mice, after accounting for the frequencies of TCR sequences (n = 5, Mean ± SE).
Fig 3.
Shared sequences are found at higher frequencies in peripheral Tconv and Tregs.
A) The sequences and frequencies of the most abundant TCRs in Tconv and Tregs and whether they are shared (gray) or are not shared (uncolored). B) Sequences that are not shared (grey) are found at significantly lower frequencies, while shared sequences (white) are found at significantly higher frequencies in the repertoires. Frequencies are measured as percentage of entire repertoire. Statistical significance was determined using the student’s t-test (n = 5, Mean ± SE, * p<.05, ** p<.01, *** p<0.0001).
Fig 4.
Limited sharing of high frequency TCRs in the thymus.
A) Tabulated results displaying the number of individual TCRβ chain amino acid sequences identified in the thymus of each mouse, and the number and percent of TCRs shared between Tconv and Tregs within each mouse. B) Comparison of the percentage of individual sequences shared by Tconv and Tregs in the spleen/lymph node and thymus of STAR mice. C) Comparison of the percent of all analyzed CD4+ T cells with TCR sequences that are shared by Tconv and Treg populations within the periphery or the thymus. D) Proportions of shared (white) or unshared (gray) TCR sequences found within particular frequency intervals in the thymus of STAR mice. Frequencies are measured as percentage of the entire CD4+ repertoire. Statistical significance was determined using the student’s t-test (n = 3, Mean ± SE, * p<.05, ** p<.01, *** p<0.0001).
Fig 5.
Analysis of amino acid and DNA TCRβ sequence sharing in the thymus and periphery of STAR mice.
A) Comparison of the total number of TCRβ sequences shared between Tconv and Treg populations at the amino acid level (gray) and the number of shared amino acid sequences also sharing an identical nucleotide sequence (white) within individual mice, or between mice, in the periphery (Spl/LN, left) and thymus (Thy, right). B) Percentage of TCR sequences shared at the amino acid level that also share identical nucleotide sequences by Tconv and Treg populations within an individual mouse or between individual mice in the periphery (n = 5) and thymus (n = 3) of STAR mice. Statistical significance was determined using Fisher’s exact test and student’s t-test (Mean ± SE, *** p<0.0001).