Fig. 7: LCK retains CD4 on the surface of T cells in a kinase-independent manner.

a,b, Relative CD4 surface levels on CD4⁺ T cells isolated from indicated Lck variants were determined by flow cytometry. a, Representative mice; Unst., unstained. b, Normalized CD4 expression (average gMFI of Lck^WT/WT was set as 1 in each experiment). Individual mice and medians are shown; n = 25 Lck^WT/WT, n = 13 Lck^KO/KO, n = 12 Lck^KR/KR, n = 18 Lck^CA/CA and n = 10 Lck^CA/KR mice in 11 independent experiments. The statistical significance was calculated using a Wilcoxon signed-rank test (versus the value of 1). c, Surface CD4 levels on LCK^KO Jurkat cells transduced with indicated LCK variants were determined by flow cytometry (gMFI) and normalized to CD4 levels on untransduced LCK^KO Jurkat cells (=1, indicated by a dashed line). Independent experiments and means are shown; n = 3 independent experiments. The statistical significance was calculated using a one-value t-test. d, LN T cells of indicated Lck-variant strains were incubated with a PKC inhibitor Ro-32-0432 overnight. CD4 expression was analyzed by flow cytometry (gMFI) and normalized to untreated cells from the same mouse (=1, indicated by a dashed line); n = 7 independent experiments/mice for Lck^WT/WT and Lck^CA/CA and n = 6 for Lck^CA/KR. Independent experiments and means are shown. The statistical significance was tested using a Mann–Whitney test; PKCi, PKC inhibitor. e,f, CD4 on LN CD4⁺ T cells from Lck^WT/WT or Lck^CA/CA B3K508 Rag2^KO/KO mice was visualized with gold-labeled antibodies using transmission electron microscopy. e, Representative cells are shown. The arrows indicate CD4 molecules. f, Normalized pair correlation function values for CD4 clustering (clusters of 0–50 nM) were calculated for each cell. Individual values and medians + s.d. are shown; n = 358 Lck^WT/WT and n = 423 Lck^CA/CA cells from three independent experiments. The statistical significance was tested using a Mann–Whitney test.

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