GPATCH4 (Q5T3I0) - a nucleolar and Cajal-body helicase cofactor in the pan-MLL-fusion core, with no DHX15 and no G-patch relative beside it

SYSTEM: the AP-MS host is JURKAT, a human T-ALL line, while KMT2A::AFF1 disease is B-lineage infant leukaemia; a partner requiring a B-cell or infant proteome cannot appear here and a T-lineage partner can appear spuriously. The bait is a Strep-HA-tagged, SINGLE-COPY, doxycycline-induced transgene at an FRT/Flp cassette, not the endogenous fusion. Prey came from bulk nuclear lysate (buffer C420, 420 mM NaCl) after a single STREP-Tactin step, so co-purification is NOT evidence of a direct contact. INCLUSION RULE: the paper kept the top 300 interactions per bait by combined p-value rather than applying a p threshold (Methods, verbatim: "All the identified bait-prey pairs were ranked by the combined p-value. The 300 most significant interactions per bait were retained."). Measured by this beat from the cached MOESM4 xlsx on 2026-09-21: 545 of the 2,049 per-bait p-values in the table (26.6%) exceed 0.05; within the MLL-AF4 column alone it is 186 of 291 (63.9%), and that column's maximum p is 0.175. MEASURED BY THIS BEAT, 2026-09-21, from the cached MOESM4 xlsx: across all 42 pan-fusion proteins the MLL-AF4 column carries the LOWEST total spectral counts of the seven baits for 34 of 42, and is sixth or seventh for 40 of 42. That is a bait-level property (transgene expression, purification depth or the DDX5/DDX17 GLM normalisation), not a statement about any one prey, so a low MLL-AF4 count must not be read as a weaker association than the same protein's count under another bait. For GPATCH4 the MLL-AF4 total (17 spectra over two replicates) is rank 6 of the seven baits, against a maximum of 28 under another bait. All seven per-bait values, p (replicate spectral counts): MLL-AF1p 0.0654 (4|4); MLL-AF4 0.0024 (6|11); MLL-AF9 0.0015 (16|12); MLL-CBP 0.0002 (12|11); MLL-EEN 0.0113 (9|15); MLL-ENL 0.0032 (9|12); MLL-GAS7 0.0139 (10|11). SPECIFIC TO THIS PROTEIN: MLL-AF1p at p = 0.0654 is above 0.05 and is present by the rank cutoff alone. Conversely the MLL-AF4 value of 0.0024 is the second-strongest in this tranche and MLL-AF4 is unusually NOT this protein's shallowest column. Nucleolar proteins are abundant in nuclear extract, which raises the prior probability of recovery independent of the bait. SELECTIVITY: the paper's named non-selectivity controls are RFC1, SF3A3 and CCT3 - knockdown inhibited MLL-wild-type K562 and HL-60 as much as MOLM-13 - against MEN1 as the selective positive. Only 40 of the 128 ('the 40 candidate genes with the highest confidence') entered that MLL-r versus MLL-wild-type re-screen, and the paper does not list which 40. This protein is not named among the controls, but that is NOT evidence of selectivity: it is equally consistent with the protein never having reached the second screen. Recorded as unknown, and this entry must not be read as nominating a target. Values re-read from the cached xlsx on 2026-09-21 and cross-checked against analyses/skucha-mll-fusion-interactome/results.json, whose two controls (128 proteins shared by >=5 baits and 42 by all seven, both matching the paper; MEN1 present with all seven) both PASS on today's re-run.

• IntAct has never curated this dataset, so no independent curator has re-examined these calls and no interactome-coverage metric built on IntAct can see them; the raw spectra at ProteomeXchange PXD009338 have not been reanalysed here. The authors themselves removed seven proteins from the shared set by hand as 'either frequently observed contaminants or ... not detected in human hematopoietic cell lines', which concedes that the rank cutoff lets contaminants through - and ACTB survived that filter, sitting in the pan-fusion 42 with a NON-ZERO negative control (3|3). Ten of the 42 carry a non-zero negative control: PSIP1 0|5, MCM2 0|2, ACTB 3|3, CSNK2A1 2|0, RUNX1 0|2, SF3A3 0|3, TCEB3 0|1, CHD4 2|2, SMARCC1 1|0, ZNF512 0|2 - counted by this beat from the xlsx neg_ctrl column, 2026-09-21. ProteomeXchange/PRIDE 2018
• The paper states that 'more than one third of proteins in the network of conserved MLL-fusion protein interaction partners were identified as essential in recent genome-wide screens'. Abundance and essentiality both raise the odds of recovery in an AP-MS experiment, so membership in the conserved core is partly a statement about how much of a protein is present in a nucleus, not only about what it binds. Skucha A et al. 2018

Human cell lines, 2024, Nucleic Acids Research; crosslinking, RiboMeth-seq-type methylation profiling and knockdown. Not haematopoietic, not a fusion system. Read from the abstract. The rDNA-locus crosslinking is the fact this entry leans on and it is an abstract-level statement; the specific rDNA regions were not checked.

• DHX15 is ABSENT from the Skucha 960-protein network at any n, as is every other G-patch protein (checked by this beat from the xlsx, 2026-09-21). So the cofactor was recovered without its helicase, and this paper's own finding that part of GPATCH4's function is DHX15-independent is the only thing that keeps that from being fatal to a functional reading. Skucha A et al. 2018

2021, Biochem Biophys Res Commun, a short-format journal; three authors; imaging and knockdown in cultured cells, not haematopoietic and not a fusion system. This is the ONLY source this beat found for GPATCH4's subnuclear address - UniProt Q5T3I0 records NO subcellular location comment at all, which is the standing rule about a database's silence not being evidence. Read from the abstract.

• This paper finds NO significant effect of GPATCH4 knockdown on pre-rRNA processing, while the 2024 NAR study finds impaired 2'-O-methylation and reduced protein synthesis. The two are reconcilable (methylation is not processing) but they are different pictures of how much GPATCH4 matters, from two small studies, and no third group has weighed in. Kanwal N et al. 2024

Mammalian cells with ectopically expressed MLL-AF4 and a murinized pSer variant, 2022, Cell & Bioscience; gene-expression profiling plus intracellular localisation. This is an OVEREXPRESSION system, not the endogenous t(4;11) fusion in a patient-derived blast, and this project's standing rule is that a punctate or compartment-enriched appearance in an overexpression system is weak evidence about the endogenous protein. Cited here for the CONTEXT - that the index fusion has a reported nucleolar pool - not as proof of it. Read from the abstract.

• If MLL-AF4 really is predominantly nucleolar in these cells, then EVERY abundant nucleolar protein becomes a high-probability co-purification partner for reasons of shared compartment rather than function - which makes the thirteen-protein nucleolar block in this network easier to explain and harder to interpret. The same observation that motivates the GPATCH4 row also supplies its leading null hypothesis. Skucha A et al. 2018