Last updated: 2026-09-09

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Knit directory: immgenT-GP-analysis/analysis/

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Note that any generated files, e.g. HTML, png, CSS, etc., are not included in this status report because it is ok for generated content to have uncommitted changes.

These are the previous versions of the repository in which changes were made to the R Markdown (analysis/FigureS3.Rmd) and HTML (docs/FigureS3.html) files. If you’ve configured a remote Git repository (see ?wflow_git_remote), click on the hyperlinks in the table below to view the files as they were in that past version.

File Version Author Date Message
Rmd c233cd8 Ziang Zhang 2026-09-09 Extended Data reorganisation: split the tissue/cluster figure, renumber 3-8
html 1e88d7e Ziang Zhang 2026-09-04 Build site: published captions and titles across all 24 pages
Rmd 0267e5b Ziang Zhang 2026-09-04 Captions from the published manuscript; trim editor notes off the page code
html 19c977f Ziang Zhang 2026-09-02 Build site: Extended Data 5-7 renumbered, Figure S5 page rebuilt
html 9167d8f Ziang Zhang 2026-08-19 Build site: Extended Data Figure 3 page rebuilt
Rmd 7b0f736 Ziang Zhang 2026-08-19 Extended Data Figure 3 intro: drop the repository-scope phrasing
html ae03072 Ziang Zhang 2026-08-19 Build site: six pages rebuilt after the prose cleanup
Rmd adc2327 Ziang Zhang 2026-08-19 Site prose: finish taking internal notes off the pages
html eeca07b Ziang Zhang 2026-08-05 Keep pre-refactor provenance in panel comments off the published pages
Rmd 5651d0e Ziang Zhang 2026-08-05 Extended Data tables: reorder to six, rebuild Table 1, drop internal notes
html cbcec52 Ziang Zhang 2026-07-30 Build site: Extended Data Figure naming
Rmd 66aa029 Ziang Zhang 2026-07-30 Name the Extended Data figures as published on the site
html ac650a0 Ziang Zhang 2026-07-30 Build site: Figure S5 (ex-S6a) and Figure S6 as a-f
html ae21d37 Ziang Zhang 2026-07-28 Build site: republish after the reorder commits
html d538aa2 Ziang Zhang 2026-07-28 Build site: reordered Figures 6 / S6 / S3 and the new Figure 7b page
Rmd 4c07670 Ziang Zhang 2026-07-28 Reorder Figures 6, S6 and S3; make the ex-S5 figure Figure 7b
html 029b0ae Ziang Zhang 2026-07-28 Build site.
Rmd 0f5b5da Ziang Zhang 2026-07-28 Align all figure captions with captions_20260728_final.docx
html 3fc3789 Ziang Zhang 2026-07-27 Republish all 24 pages
html 1390a03 Ziang Zhang 2026-07-27 Republish all 24 pages
html adaef21 Ziang Zhang 2026-07-27 Build site: panel fixes and PDF-derived assets
html 5b19858 Ziang Zhang 2026-07-27 Build site.
Rmd 91ee059 Ziang Zhang 2026-07-26 Select each panel’s code block by name, not by line number
html 91ee059 Ziang Zhang 2026-07-26 Select each panel’s code block by name, not by line number
Rmd fa5a501 Ziang Zhang 2026-07-18 Fig S3c: reproduce collaborator panel (activated CD4/CD8, threshold 0.1)
html fa5a501 Ziang Zhang 2026-07-18 Fig S3c: reproduce collaborator panel (activated CD4/CD8, threshold 0.1)
Rmd b197499 Ziang Zhang 2026-07-16 Restructure main figures (renumber + split Figure 1)
html b197499 Ziang Zhang 2026-07-16 Restructure main figures (renumber + split Figure 1)
html 92021bf Ziang Zhang 2026-07-02 Build site.
html 827c89b Ziang Zhang 2026-07-02 Build site.
Rmd 8ac7f9f Ziang Zhang 2026-07-02 Add data provenance notes to each script; remove conversational
Rmd f9db962 Ziang Zhang 2026-07-02 Simplify layout: drop old code/script folders, rename
html c6e5086 Ziang Zhang 2026-07-02 Build site.
html 5a79883 Ziang Zhang 2026-07-02 Build site.
Rmd 2b0e445 Ziang Zhang 2026-07-02 Fix GitHub source links to point at the new
html cf1d0ac Ziang Zhang 2026-07-02 Build site.
Rmd 06b2461 Ziang Zhang 2026-07-02 Initial commit: immgenT-GP-analysis
html 06b2461 Ziang Zhang 2026-07-02 Initial commit: immgenT-GP-analysis

This single-panel figure is produced by script/FigureS3.R. It shows the final full row-centered matrix, with all 200 GPs and all 107 observed level-2 clusters. The analysis uses healthy non-thymocyte cells (condition_broad == "healthy" and annotation_level1 != "thymocyte"). For each GP, its mean loading across clusters is subtracted from every cluster mean, so a row says where a program is more or less active than its own average rather than how large its loading is. The centered color scale is fixed at -0.2 to 0.2; values outside this range are saturated at the endpoint colors. It is the cluster counterpart of Extended Data Figure 5, which shows the tissue-associated GPs across tissues and lineages, and the mean-loading counterpart of Extended Data Figure 6, which resolves the same clusters cell by cell.

Setup, centering, and ordering

# Figure S3. GP activity across T cell clusters.
#
# One panel (see analysis/FigureS3.Rmd for the caption text):
#   s3   Row-centered mean GP activity across the 107 level-2 clusters, all
#        200 GPs, in healthy non-thymocyte cells.
#
#
# For each GP, its mean loading across clusters is subtracted from every cluster
# mean, so the panel shows where a program is more or less active than its own
# average rather than how large its loading is. The centered color scale is
# fixed at [-0.2, 0.2]; values outside this range saturate at the endpoint
# colors. Level2 columns follow Figure 1's level1 order, with level2 labels
# alphabetized within each level1 block.
#
# Required inputs (data/) -- see code/README.md's "Data provenance" table
# for the full picture:
#   L_pm_filtered.rds                        [code/pipeline/01b_filter_cells.R]
#   igt1_96_..._ADTonly.Rds                  [primary input Seurat object]

suppressPackageStartupMessages({
  library(ComplexHeatmap)
  library(circlize)
  library(grid)
  library(ZemmourLib)
})

if (!file.exists("code/R/setup_data.R")) {
  stop("Run this script from the immgenT-GP-analysis repository root.")
}

source("code/R/setup_data.R")
source("code/R/centered_mean_heatmap.R")

figure_path <- "figures/final-selected/Figure S3"
dir.create(figure_path, recursive = TRUE, showWarnings = FALSE)

# ============================================================
# Setup, centering, and ordering
# ============================================================
gp_data <- load_gp_data()
reference <- healthy_nonthymocyte_reference(gp_data)
L_reference <- reference$L
meta_reference <- reference$meta

centered_color_limit <- 0.2
level1_order <- c("CD8", "CD4", "Treg", "gdT", "CD8aa", "Tz", "DN", "DP")

level2_result <- mean_loading_by_group(L_reference, meta_reference$annotation_level2)
level2_raw <- level2_result$matrix
level2_centered <- center_by_gp_mean(level2_raw)

level2_group_level1 <- level2_to_level1_map(
  meta_reference, colnames(level2_raw), level1_order
)
level2_order <- dominant_group_order(
  level2_raw,
  level2_column_order(colnames(level2_raw), level2_group_level1, level1_order)
)

stopifnot(
  nrow(level2_centered) == 200L,
  ncol(level2_centered) == 107L,
  max(abs(rowMeans(level2_centered))) < 1e-12
)

level2_palette <- palette_for_groups(
  colnames(level2_centered),
  ZemmourLib::immgent_colors$level2,
  "annotation_level2"
)
level1_palette <- ZemmourLib::immgent_colors$level1[level1_order]

Cluster mean loading

render_centered_heatmap(
  level2_centered,
  level2_palette,
  "cluster (annotation_level2)",
  file.path(figure_path, "s3.pdf"),
  level2_order$row_order,
  level2_order$column_order,
  centered_color_limit,
  paste0(
    "all 200 GPs; level2 columns: Figure 1 level1 order ",
    "(CD8, CD4, Treg, gdT, CD8aa, Tz, DN, DP); ",
    "alphabetical within level1; GP rows: dominant-group blocks"
  ),
  group_level1 = level2_group_level1,
  level1_palette = level1_palette
)

summary_dir <- "output/FigureS3"   # build intermediate (not a manuscript panel)
dir.create(summary_dir, recursive = TRUE, showWarnings = FALSE)
write.csv(
  data.frame(
    panel = "s3",
    grouping = "annotation_level2",
    view = "full row-centered mean loading",
    gp_count = nrow(level2_centered),
    group_count = ncol(level2_centered),
    centered_definition = "group mean minus mean across groups for each GP",
    color_min = -centered_color_limit,
    color_mid = 0,
    color_max = centered_color_limit,
    observed_min = min(level2_centered),
    observed_max = max(level2_centered)
  ),
  file.path(summary_dir, "S3_summary.csv"),
  row.names = FALSE,
  quote = FALSE
)

message("Wrote Figure S3 to ", normalizePath(figure_path))

Version Author Date
c233cd8 Ziang Zhang 2026-09-09

Extended Data Fig. 3. Row-centered mean GP activity across the 107 level-2 clusters. Mean GP activity computed in samples at baseline.


sessionInfo()
R version 4.5.1 (2025-06-13)
Platform: aarch64-apple-darwin20
Running under: macOS Sequoia 15.6.1

Matrix products: default
BLAS:   /System/Library/Frameworks/Accelerate.framework/Versions/A/Frameworks/vecLib.framework/Versions/A/libBLAS.dylib 
LAPACK: /Library/Frameworks/R.framework/Versions/4.5-arm64/Resources/lib/libRlapack.dylib;  LAPACK version 3.12.1

locale:
[1] en_CA/en_CA/en_CA/C/en_CA/en_CA

time zone: America/Chicago
tzcode source: internal

attached base packages:
[1] stats     graphics  grDevices utils     datasets  methods   base     

loaded via a namespace (and not attached):
 [1] vctrs_0.7.3     cli_3.6.6       knitr_1.50      rlang_1.2.0    
 [5] xfun_0.55       stringi_1.8.9   otel_0.2.0      promises_1.5.0 
 [9] jsonlite_2.0.0  workflowr_1.7.2 glue_1.8.1      rprojroot_2.1.1
[13] git2r_0.36.2    htmltools_0.5.9 httpuv_1.6.16   sass_0.4.10    
[17] rmarkdown_2.30  evaluate_1.0.5  jquerylib_0.1.4 tibble_3.3.0   
[21] fastmap_1.2.0   yaml_2.3.12     lifecycle_1.0.5 whisker_0.4.1  
[25] stringr_1.6.0   compiler_4.5.1  fs_1.6.6        Rcpp_1.1.1-1.1 
[29] pkgconfig_2.0.3 later_1.4.4     digest_0.6.39   R6_2.6.1       
[33] pillar_1.11.1   magrittr_2.0.5  bslib_0.9.0     tools_4.5.1    
[37] cachem_1.1.0