Reduction of disulfide bonds in cysteine-containing proteins was performed with dithiothreitol (56°C, 30 min, 10 mM in 50 mM HEPES, pH 8.5). Reduced cysteines were alkylated with 2-chloroacetamide (room temperature, in the dark, 30 min, 20 mM in 50 mM HEPES, pH 8.5). Samples were prepared using the SP3 protocol (18 (link),19 (link)) and trypsin (sequencing grade, Promega) was added in an enzyme to protein ratio of 1:50 for overnight digestion at 37°C. Next day, peptides were recovered in HEPES buffer by collecting the supernatants on a magnet and combining with second elution wash of beads with HEPES buffer. Peptides were labeled with TMT10plex (20 (link)) Isobaric Label Reagent (Thermo Fisher Scientific) according the manufacturer's instructions. For further sample clean up an OASIS® HLB μElution Plate (Waters) was used. Offline high pH reverse phase fractionation was carried out on an Agilent 1200 Infinity high-performance liquid chromatography system, equipped with a Gemini C18 column (3 μm, 110 Å, 100 × 1.0 mm, Phenomenex) (21 (link)). For the interactomes, eight fractions were pooled and for input samples 12 fractions, each fraction was subjected individually to mass spectrometry.