Oncofusion

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Analysisstrong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.Q22-phospho-contacts

Enumeration: every phosphorylation-dependent and kinase-regulated contact on KMT2A and its fusions that this beat could find

also called phospho-contact enumeration · Q22 enumeration · kinase enumeration KMT2A

A list of everything we searched for, what we found, what we looked at and rejected, and what we could not read. The point is that someone repeating this next year can tell our decisions from our blind spots.

The same thing, for a scientist

The interactome enumeration declared itself close to saturated for stable complexes and **'not at all' for transient and phosphorylation-dependent contacts**. This is the first pass at that class. **Found: 11 kinase-substrate relations touching KMT2A, AFF1, AFF4, MEN1 or the SEC, of which 3 sit on the fusion-retained fragment, 2 on the lost fragment, 3 on partners, 2 on menin and 1 on a downstream effector.** The headline is that the retained fragment is **not** menin-only: **CK2 phosphorylation of S136/S142 gates the LEDGF/p75 IBM**, with a fusion transformation phenotype. **Rejected with reasons: 9.** **Unreadable after all routes: 3.** **Counted absences: 4.** The retained-fragment class is nowhere near saturated - 51 of 56 observed retained phosphosites have no kinase and no function.

Go deeper
FOUND - kinase-substrate relations, with the retained/lost call
#kinasesubstratesitefate in fusiontested on a fusion?entry
1CK2KMT2AS136, S142RETAINEDyes - S136A/S142A impairs transformation, phosphomimetic rescuesinterface-psip1-ibd-kmt2a-phosphoswitch
2ATRKMT2AS516RETAINEDno - fusions tested as dominant negatives onlymechanism-atr-kmt2a-s516-skp2-switch
3CK2KMT2AT2724, S2726LOSTindirectly - fusion displaced by stabilised wild-typemechanism-ck2-t2724-s2726-licenses-taspase1-cleavage
4unassigned (S-P; CDK9 candidate)AFF1S588RETAINED (partner side, t(4;11) only)partly - complex with MLL-AF4 shown; S588A not tested on fusioninterface-ywhaq-aff1-s588-14-3-3
5CDK9/P-TEFbAFF4unmappedpartner, not in any fusion chainnomechanism-cdk9-phosphorylates-aff4-sec-assembly
6CDK9 (auto)CDK9T186partnernosame
7PKC (upstream)CDK9 activity via AFF1-HEXIM1n/apartnernosame
8PKAMEN1S487n/a (menin is not in the fusion chain)no - beta cells onlymechanism-pka-menin-s487-chromatin-release
9ATM/ATRMEN1unmappedn/ano - melanocytes onlymechanism-atm-atr-men1-stabilisation-dna-damage
10CDK1/2 + ERK1/2MEF2CS222downstream effectoryes for drug response, in T-ALLmechanism-mef2c-s222-predicts-menin-inhibitor-response
11CK2DEKS301/303/306/307not a KMT2A contactyes - MLL-AF9 modelsgene-csnk2a1

Adjacent, non-phospho or indirect, recorded so they are not miscounted: SCF(Skp2) and APC(Cdc20) degrons on the retained region (interface-skp2-kmt2a-n-terminal-degron); Cks1/Cks2 spanning MllN and MllC, fate undetermined (interface-cks1-cks2-kmt2a-cell-cycle-adaptor); SET/PP2A binding both wild-type and fusion with the interface unmapped (mechanism-set-pp2a-phosphatase-arm-kmt2a-r); GSK3, a genotype-specific dependency with no KMT2A substrate (mechanism-gsk3-dependency-kmt2a-r-leukaemia).

REJECTED - found, looked on-beat, judged not to be

Recorded per the bounded-rejection rule: identifier, one-line reason, what would change our mind.

  1. PMID 32792914 - 'Mixed Lineage Leukemia 1 Promoted Neuron Apoptosis ... via Regulating ASK-1/TNF-alpha Complex'. MLL1 transcriptionally drives ASK-1; the 14-3-3 in the abstract belongs to the ASK-1/STRAP/14-3-3 complex, not to a KMT2A contact. Change our mind: a co-immunoprecipitation of MLL1 with 14-3-3 in the figures.
  2. PMID 28094252 - LIM proteins, CHK2, CDC25, 14-3-3 and radiosensitivity. MLL1 appears only as a transcriptional driver of FHL1. Change our mind: evidence MLL1 is in the 14-3-3-CDC25 complex.
  3. PMID 31138789 - ZNF479/MT-1 in hepatocellular carcinoma. 14-3-3-epsilon and the MLL complex are separate links in one signalling chain. Change our mind: a direct interaction experiment.
  4. PMID 26800457 - C. elegans EFF-1/AFF-1 and 14-3-3. Species-level homonym: nematode AFF-1 is a cell-fusogen unrelated to human AFF1. Change our mind: nothing; this is a name collision and is recorded so it is not rediscovered.
  5. PMID 41457983 - CAR Tregs in myocardial infarction. Returned by the alias query and entirely off-topic.
  6. PMID 15459319 / 15221006 (Oliver; Bursen) - SIAH1/SIAH2-mediated degradation of AF4 and protection of AF4.MLL. Genuinely on-beat for AFF1 regulation and a real oncogenic mechanism, but neither abstract reports the degron as phosphorylation-dependent, and the robotic-mouse mutation is a coding change in the Siah-binding region rather than a phosphosite. Change our mind: a phosphosite in the AF4 SIAH degron, or a kinase requirement for SIAH binding. This is the strongest single rejection in the list and the most likely to be overturned.
  7. PMID 12511424 - germline ATM missense in one child with MLL rearrangement. A single patient; concerns ATM function, not an ATM substrate on KMT2A. Change our mind: a cohort, or an ATM site on KMT2A.
  8. PMID 14555522 - HSP90/17-AAG in FLT3-positive leukaemia. Explicitly reports that MLL-AF4 levels were not reduced by 17-AAG, so it is a negative for chaperone-dependent fusion stability and is recorded as such rather than as a contact.
  9. PMID 23471103 - the AFF4-P-TEFb structure. Structural, not phospho-regulatory; used as context in the CDK9 entry rather than as a phospho-contact. Change our mind: a phosphosite in the resolved interface.

Laboratories whose work was read and judged inapplicable: the HIV transcription groups working on SEC phospho-assembly (Kuzmina, Schulze-Gahmen, Zhou) publish repeatedly on exactly the phospho-regulation this beat wants, in a system with no KMT2A fusion. Their results are cited here as mechanism and not as evidence about leukaemia. That judgement would be wrong if anyone showed the SEC assembles differently on a fusion than on Tat.

UNREADABLE - found, on topic, could not be read

Per concept-known-but-unreadable-literature. All three had a PMCID and all three failed both fullTextXML and efetch db=pmc with the body-element test applied.

PMIDPMCIDwhat the title assertsroutes tried
17908926PMC1993870bimodal degradation of MLL by SCFSkp2 and APCCdc20; a circuit lost in leukaemogenic fusionsfullTextXML HTTP 500; efetch db=pmc well-formed but no body, carrying the note 'The publisher of this article does not allow downloading of the full text in XML form'; genesdev.cshlp.org HTTP 403; europepmc.org article page HTTP 403
23648481PMC3700119MEN1 stabilised by ATM/ATR phosphorylation after DNA damagefullTextXML HTTP 500; efetch db=pmc no body
31493143PMC1299431914-3-3 theta binds AF4 Ser588 and complexes with MLL-AF4EPMC core record marks it not open access; full text not retrieved

Nothing in this beat states what those papers 'probably say'. Each is cited at abstract level with the limitation named in its caveats.

Two resources were also unreachable: PhosphoSitePlus (HTTP 403 on both the search endpoint and protein pages) and iPTMnet (HTTP 503 on two attempts). The S821 verdict is bounded by that and says so.

COUNTED ABSENCES
  1. S821 is not a phosphosite in 130 PRIDE/PTMeXchange sites or 25 UniProt sites; nor is T820; nor is any acceptor of the other two canonical 14-3-3 motif matches. Positive controls S516, S136, S142, T2724, S2726 all recovered.
  2. No published direct 14-3-3-KMT2A binding. Alias-expanded query over seven paralogue symbols plus family name and four KMT2A symbols returns 5 records, 4 rejected, 1 about AF4. (KMT2A OR MLL1) AND YWHAG returns 0, for the paralogue reproduced in both interactome datasets.
  3. Zero canonical 14-3-3 motifs in AFF1, AFF4 or MEN1. AFF1 nonetheless has a demonstrated 14-3-3 contact at S588, which is the clearest available demonstration that canonical-motif scanning has poor sensitivity for this family.
  4. No kinase assigned to 51 of the 56 observed phosphosites in the retained fragment. This is the size of the remaining space, and it is the answer to 'how saturated is this class': not at all.

All four are bounded by the exact queries in the entries they come from.

WHAT THE BEAT CONCLUDES ABOUT ITS OWN PREMISE

The dispatch premise was that a phospho-gated contact N-terminal to the breakpoint would be a second, non-menin handle and a direct contradiction of the cycle-28 menin-only finding.

The premise's chosen route - 14-3-3 at S821 - is dead. The residue is not phosphorylated in any queryable dataset and the interaction has never been shown directly.

The premise's conclusion is nonetheless right, by a route nobody in this project had looked at. The LEDGF/p75 IBM at KMT2A 111-160 is phospho-gated at S136/S142 by CK2, is retained by every fusion, and has a published fusion transformation phenotype with phosphomimetic rescue. It has been in the literature since 2018 and appears nowhere in this graph - the existing interface-psip1-ibd-kmt2a entry, which describes the same interface, contains no mention of phosphorylation, CK2, S136 or S142.

That is the beat's finding and also its warning: the second handle was not missing from the literature, it was missing from the graph.

Facts and figures
statusrun
found kinase substrate relations11
on retained fragment with kinase2
on lost fragment1
on partners3
on menin2
on downstream effector1
non kmt2a ck2 route1
adjacent non phospho contacts4
rejected with reasons9
unreadable after all routes3
counted absences4
retained phosphosites without kinase51
result summaryThe retained fragment is not menin-only: CK2 phosphorylation of KMT2A S136/S142 gates the LEDGF/p75 IBM and has a published fusion transformation phenotype. The 14-3-3/S821 route is closed. 51 of 56 retained phosphosites remain without a kinase.
resources unreachable
[
  "PhosphoSitePlus (HTTP 403)",
  "iPTMnet (HTTP 503)"
]
menin mbm1 kd conditions
{
  "_why": "Added by king sweep 2026-09-21 (Q82 corrections-owed #3). This entry quotes the menin-MBM1 53 nM figure. That number is NOT comparable to the KMT2A CXXC-DNA affinities without stating its buffer, and the omission is how a 1,300-fold 'discrepancy' was manufactured.",
  "value": "53 nM",
  "motif": "KMT2A MBM1, residues 4-15, FITC-labelled peptide at 15 nM",
  "method": "fluorescence polarisation",
  "buffer": "50 mM Tris, pH 7.5, 50 mM NaCl, 1 mM DTT",
  "nacl_mM": 50,
  "companion_itc": "72 nM by ITC, 50 mM phosphate pH 7.5, 50 mM NaCl, 1 mM beta-mercaptoethanol, 27 C",
  "source_pmid": "20961854",
  "source_quote": "\"in the FP buffer (50 mM Tris, pH 7.5, 50 mM NaCl, 1 mM DTT) used for binding experiments (Kd measurements)\" -- Grembecka 2010 JBC, Methods, fetched and read from PMC3003368 on 2026-09-21",
  "do_not_compare_naively": {
    "against": "KMT2A CXXC-DNA Kd 44 uM (PMID 20010842, Cierpicki 2010)",
    "because": "that measurement was made at 300 mM NaCl -- verbatim from its Methods, \"50 mM phosphate buffer (pH 7.1), 300 mM NaCl, 2 mM DTT at 25 C\" (PMC2908503, read 2026-09-21) -- SIX-FOLD more salt, on a salt-sensitive protein-DNA contact, against a hydrophobic peptide pocket that is not.",
    "best_matched_pair": "44 uM at 300 mM NaCl vs the 5.7 uM tagless CXXC measurement at 150 mM NaCl gives 108-fold against menin, not ~1,000-fold",
    "see": "ana-the-menin-affinity-comparison-was-made-at-one-sixth-the-salt"
  },
  "added": "2026-09-21"
}
cxxc dna kd conditions
{
  "_why": "Added by king sweep 2026-09-21 (cycle 46). This entry quotes the KMT2A CXXC-DNA 44 uM. That number is a property of five choices, not of the molecule, and quoting it bare is how this graph manufactured a 1,300-fold 'discrepancy' with the menin contact.",
  "value": "44 uM",
  "construct": "isolated CXXC domain, KMT2A 1147-1203, untagged, 15N-labelled",
  "ligand": "DNA pal oligonucleotide, one CpG element",
  "method": "NMR titration",
  "buffer": "50 mM phosphate, pH 7.1, 300 mM NaCl, 2 mM DTT, 25 C",
  "nacl_mM": 300,
  "source_pmid": "20010842",
  "source_quote": "\"Determination of the dissociation constants by NMR for the CXXC domain (MLL 1147-1203) and mutants was performed by titration of 15N-labeled protein with concentrated solutions of DNA pal oligonucleotide. Experiments were carried out in 50 mM phosphate buffer (pH 7.1), 300 mM NaCl, 2 mM DTT at 25 C.\" -- Cierpicki 2010 NSMB, Methods, fetched and read from PMC2908503 on 2026-09-21. The result sentence reads \"The wild type protein yielded a Kd of 44 uM\".",
  "this_is_the_high_salt_end": "300 mM NaCl is roughly twice physiological ionic strength and SIX TIMES the 50 mM the menin-MBM1 53 nM was measured in. The same domain measured tagless at 150 mM NaCl gives 5.7 uM -- 7.7-fold tighter from salt alone.",
  "do_not_compare_naively": {
    "against": "menin-MBM1 53 nM (PMID 20961854), measured at 50 mM NaCl by FP",
    "best_matched_pair": "108-fold, not ~1,000-fold",
    "see": "ana-the-menin-affinity-comparison-was-made-at-one-sixth-the-salt"
  },
  "added": "2026-09-21"
}
Still open
  1. Run the same enumeration on the 3' partners MLLT1, MLLT3, MLLT10 and ELL. This beat covered AFF1 and AFF4 and left the others entirely unsearched.
  2. Query PhosphoSitePlus and iPTMnet when reachable; both were down and both curate low-throughput site-kinase assignments that PRIDE re-analysis does not contain.
  3. Mine the supplementary tables of the KMT2A-rearranged AML phosphoproteomic study for KMT2A, AFF1, AFF4, MEN1 and DOT1L peptides. That single file could double this enumeration without a new experiment.
  4. Is the SIAH-AF4 degron phosphorylation-dependent? It is the rejection in this list most likely to be overturned.
Every source behind this entry (3)
  1. E1strong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.analysis

    This enumeration identified eleven kinase-substrate relations touching KMT2A, AFF1, AFF4, MEN1, the SEC or a menin-pathway effector, four adjacent non-phosphorylation contacts, nine bounded rejections, three papers unreadable after all retrieval routes and four counted absences; two of the eleven lie on the KMT2A fragment retained by all fusions with a named kinase, and fifty-one of the fifty-six observed retained-fragment phosphosites have no assigned kinase.

    What this evidence cannot show

    An enumeration is bounded by its queries. Coverage is PubMed title/abstract searching with alias expansion over retired and Greek-letter symbols, UniProtKB curation, and the EMBL-EBI Proteins API proteomics-PTM resource, run 2026-09-19. It does NOT include supplementary tables, preprints, non-indexed journals, PhosphoSitePlus (HTTP 403) or iPTMnet (HTTP 503). Several of the eleven relations are abstract-level reads. The 3' partners MLLT1, MLLT3, MLLT10 and ELL were not searched at all. Eleven is a floor, not a census.

    Evidence that argues against this (2)
    • The same graph's interactome enumeration judged the stable-complex catalogue nearly saturated and this class 'not at all', and this beat's own count of 51 unassigned kinases among 56 retained phosphosites confirms that judgement rather than overturning it. Eleven relations is a first pass and should not be cited as the size of the field. Casado P et al. 2023
    • A dedicated disease-level phosphoproteomic study of 74 KMT2A-rearranged AML patients located the discriminating signal in DOT1L phosphorylation and CDK1 activity - neither of which is a residue on KMT2A - which suggests a literature-derived enumeration keyed to KMT2A residues may be looking in the wrong place for where the fusion's phospho-regulation actually sits. Casado P et al. 2023
  2. E2established confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.database

    The graph's existing entry interface-psip1-ibd-kmt2a maps the LEDGF contact to KMT2A residues 140-160 with Phe148 and Phe151 as key residues, records kd_nM as null, and contains no mention of phosphorylation, CK2, S136 or S142.

    What this evidence cannot show

    This is an observation about this repository on one date, not about the literature, and it will go stale the moment the entry is corrected. The omission does not make the existing entry wrong - its structural content is accurate and independently sourced - only incomplete on the regulatory layer.

    Evidence that argues against this (1)
    • The existing entry is built on the structural validation of the LEDGF/p75-MLL interface, which predates the phospho-switch result by four years. The omission therefore reflects the state of the literature the entry was assembled from rather than an error of reading, and the same gap would be found in most secondary sources on this interface. Cermáková K et al. 2014
  3. E3strong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.paper

    Structurally conserved IBD-binding motifs on LEDGF/p75 partners are regulated by phosphorylation, permitting switching between low- and high-affinity states, and eliminating the IBM phosphorylation sites on MLL1 disrupts the oncogenic potential of primary MLL1-rearranged leukaemic cells, which the authors describe as a previously unknown oncogenic dependency.

    PMID 29997176DOI
    What this evidence cannot show

    Abstract plus PMC HTML full text (PMC6065015) read 2026-09-19; Europe PMC fullTextXML returned HTTP 500 and efetch db=pmc returned a body-less record for this article. The MLL1 residue assignment to CK2 is UniProt's curation of this paper rather than a directly reported CK2-on-MLL1 experiment; the paper's own CK2 treatment experiment is on POGZ. The transformation readout is a colony assay in murine cells.

    Evidence that argues against this (1)
    • No dissociation constant is reported for the KMT2A IBM against the LEDGF IBD in either phosphorylation state, here or anywhere, while the menin arm of the same composite clamp is measured at 53 nM by fluorescence polarisation and 72 nM by calorimetry. A fold-change on an unmeasured baseline cannot establish that the second arm is in a druggable range. Cermáková K et al. 2014
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