multiDIA

Analysis and figures for the multiDIA manuscript: Spectronaut directDIA reports from three parallel digests (trypsin, Lys-C, Glu-C) of one human PD brain cohort.

Layout

data/         inputs; nothing here is produced by the code
derived/      tables built from data/ and read by the figures
analysis/     all code, and its tests
figures/      the paper's figures: panels, whole figures and PNGs
build.sh      rebuilds derived/ and figures/ from data/
index.md …    the website, https://multidia.slavovlab.net (GitHub Pages: _config.yml, CNAME, *.md)

path	contents
data/search/*-60min-Phospho.parquet	the Spectronaut reports, one per digest, searched with variable phospho. Not in the repository (too large for GitHub); place them here to rebuild.
data/uniprot_sprot_2024-01-01_HUMAN_ISOFORMS.fasta	the search database (UniProt Swiss-Prot human with isoforms, 2024-01-01)
data/metadata.xlsx	the brain bank's neuropathology sheet; ADRC # is the patient id and NPDX1 gives the condition
data/gencode/, data/uniprot/	GENCODE v50 annotation with its splice events (events.tsv), and cached UniProt features. gencode.v50.annotation.gtf.gz is not in the repository; download it from GENCODE to rerun prep_splice_events.py.
data/da_panel.txt	the marker panel for extra_differential.py --panel
derived/counts/	per-sample counts (Fig. 1a–c)
derived/da_iso/	discriminating-region coverage (Fig. 2c)
derived/mods/	the phospho scan and its site table (Fig. 3)
derived/multiDIA.sdrf.tsv	the SDRF-Proteomics sample table for the PRIDE submission, one row per raw file (prep_sdrf.py)

Code

Each file in analysis/ is named for what it produces:

prefix	role	files
fig…_	draws that figure's panels	fig1a_workflow, fig1b_depth, fig1cd_coverage, fig2a_supp1_diagnostic_peptides, fig2b_isoform_strip, fig2cd_isoform_coverage, fig3a_phospho_sites, fig3bc_phospho_atlas
prep_	builds derived/ or the data/ caches	prep_counts, prep_phospho, prep_sdrf, prep_splice_events, prep_uniprot, prep_parquet
lib_	shared by the figures	lib_report (reports, run names, metadata), lib_fasta, lib_palette, lib_svg, lib_compose (stacking panels)
extra_	exploratory; no final figure	extra_differential, extra_isoform_da, extra_isoform_unique, extra_changed_proteoforms, extra_digest_upset, extra_reported_groups, extra_perm_null, extra_why_multienzyme

audit.py checks figures/, test_units.py is the unit tests, make_synthetic.py writes a test report and preview.py renders an SVG to PNG.

Rebuild

With the environment from pyproject.toml (uv sync), from the repository root:

./build.sh                                  # derived/ and every figure, then the checks
.venv/bin/python analysis/test_units.py     # unit tests, no data needed
.venv/bin/python analysis/audit.py          # whole figures and PNGs current, one font