The order in which the fusion's programme collapses is a property of the partner, not of KMT2A - so there is no single causal chain
also called partner-dependent ordering · two chains not one
When the cancer protein is removed, one part of its machinery fails first. We propose that which part fails first depends on which partner gene is fused - so the single flow-chart the field draws is really several different flow-charts wearing one name.
The same thing, for a scientist
The KMT2A-fusion causal chain has exactly one measured arrow, in MLL::AF9 [E1]. **Hypothesis: that arrow's direction is partner-encoded.** In KMT2A::MLLT3 the retained module is a 70-residue AHD at which AF4, DOT1L, BCOR and CBX8 bind **mutually exclusively** [E2], so elongation and methylation are alternatives on one surface and the first to fail on fusion removal is whichever was occupying it. In KMT2A::AFF1 the retained partner **is** an SEC component, DOT1L association is indirect, and the two activities are not competing for one site - so the collapse order has no structural reason to be the same. **Prediction: in a KMT2A::AFF1 degron, H3K79me2 loss and the elongation change will be closer together or reversed relative to MLL::AF9**, and the difference will track the partner rather than the lineage.
The argument
- The AHD is a single, mutually exclusive binding site; the four complexes superpose at 1.4-1.9 A over the structured AF9 residues 2. A molecule of MLL-AF9 is therefore doing elongation or methylation, not both.
- Reconstitution agrees: DOT1L-AF9 does not detectably interact with AF9-AF4-P-TEFb 3.
- KMT2A::AFF1 retains an AF4-family protein that is itself an SEC component 4, so the elongation function is intrinsic rather than recruited to a contested surface.
- Therefore the same perturbation - removing the fusion - unloads a different structure in the two cases, and there is no reason to expect the same event to fail first.
What makes this more than a quibble. Every schedule and combination argument in this disease is built on the MLL::AF9 ordering, and the index disease is KMT2A::AFF1. If the ordering is partner-dependent, the field has been generalising a structure-specific result, which is the exact failure mode this project already logs as its commonest synthesis error.
Falsifiers, and what each would mean
- A KMT2A::AFF1 degron shows the same ordering with similar kinetics - the hypothesis is false and the MLL::AF9 import is safe for this claim.
- Ordering differs, but tracks lineage rather than partner - the hypothesis is false in its stated form; a myeloid KMT2A::AFF1 model would then separate the two variables.
- A YEATS-domain or AHD point mutant of MLL-ENL changes the collapse order - strong support, and cheaper than a new degron line, because ENL reader-dead mutants exist 5.
- The AHD turns out not to be occupied one-partner-at-a-time in cells - for instance if two fusion molecules cooperate at one locus - the structural premise fails and the prediction has no basis.
| statement | The temporal order in which the fusion's elongation and H3K79-methylation arms fail after acute fusion removal is determined by the retained partner module rather than by the shared KMT2A N-terminus, so the KMT2A-fusion causal chain has no single internal ordering. |
|---|---|
| origin | data-pattern |
| prior probability | 0.45 |
| chain | mechanism-kmt2a-fusion-causal-chain |
predictions
[ "In a KMT2A::AFF1 degron, the interval between the elongation change and H3K79me2 loss is shorter than in MLL::AF9, or the order is reversed.", "The difference tracks the partner: a myeloid KMT2A::AFF1 model behaves like the lymphoid one, and a lymphoid MLL-AF9 model behaves like the myeloid one.", "An AHD point mutant that removes the DOT1L contact in MLL-AF9 shifts the collapse order toward the AFF1 pattern.", "Menin plus DOT1L combination benefit is larger in AHD-type fusions, where the two arms compete, than in AF4-family fusions, where they do not." ]
experiments
[ "exp-ordering-of-elongation-and-h3k79-loss-in-kmt2a-aff1" ]
analyses
[ "ana-a-forward-reference-is-authored-not-demanded" ]
falsifiers
[ "A KMT2A::AFF1 degron reproduces the MLL::AF9 ordering with similar kinetics.", "Ordering differs but tracks lineage rather than partner when both are varied.", "The AHD is shown to be co-occupied at a locus by multiple fusion molecules, removing the structural premise.", "Combination benefit of menin plus DOT1L inhibition is indistinguishable between AHD-type and AF4-family fusions." ]
- If ordering is partner-dependent, a single pharmacodynamic sampling schedule across a KMT2A-rearranged trial will be well timed for some partners and badly timed for others, and the resulting variance will read as biological heterogeneity.
- A combination timed on the MLL::AF9 interval could give the second agent too late in an AFF1 patient, producing a negative combination result for a scheduling reason.
- Is there any published timecourse, of any perturbation, in a KMT2A::AFF1 line dense enough at early timepoints to test this without a new degron line?
- Does the AFF1 fusion recruit DOT1L at all in the same complex, or only through a second fusion molecule or through the wild-type machinery?
- If ordering is partner-dependent, is there a partner for which neither arm is first because both are constitutive?
- E1established confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.paper
Degron-mediated MLL::AF9 removal changes transcriptional elongation within 15 minutes with loss of the active chromatin landscape occurring subsequently.
MLL::AF9 degradation induces rapid changes in transcriptional elongation and subsequent loss of an active chromatin landscape. · Olsen SN, Godfrey L, Healy JP, Choi YA, et al. · Mol Cell 2022What this evidence cannot show
Abstract read 2026-09-20. The single result this hypothesis proposes is partner-specific. One fusion, one system.
Evidence that argues against this (1)
- The same study reports that combined DOT1L and menin inhibition resembles degradation, which implies the two arms converge on one outcome however they are ordered - so a difference in ordering may have no functional consequence. Dafflon C et al. 2017
- E2strong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.paper
The retained AF9 AHD binds AF4, DOT1L, BCOR and CBX8 at one mutually exclusive site, with the four complexes superposing at 1.4-1.9 A over the structured AF9 residues; an AF9 E531R mutant designed to disrupt the BCOR contact abolishes leukaemogenesis in vivo.
BCOR Binding to MLL-AF9 Is Essential for Leukemia via Altered EYA1, SIX, and MYC Activity. · Schmidt CR, Achille NJ, Kuntimaddi A, Boulton AM, et al. · Blood Cancer Discov 2020What this evidence cannot show
Abstract read 2026-09-20. Solution NMR on peptide complexes; mutual exclusivity is inferred from a shared site rather than measured by competition in cells at endogenous stoichiometry.
Evidence that argues against this (1)
- Structural exclusivity at one site says nothing about the population: a locus occupied by many fusion molecules could carry both activities simultaneously, in which case a single molecule's exclusivity does not produce a difference in collapse order. Biswas D et al. 2011
- E3strong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.paper
In reconstituted minimal complexes DOT1L forms a stable complex with AF9 that does not detectably interact with AF9-AF4-P-TEFb complexes.
Function of leukemogenic mixed lineage leukemia 1 (MLL) fusion proteins through distinct partner protein complexes. · Biswas D, Milne TA, Basrur V, Kim J, et al. · Proc Natl Acad Sci U S A 2011What this evidence cannot show
Abstract read 2026-09-20. Purified components; a negative about a stable association, not about co-occupancy in chromatin.
Evidence that argues against this (1)
- A biochemical non-interaction can coexist with functional coupling through the histone mark rather than through protein contact, and DOT1L-dependent PRC1.1 antagonism is an example of exactly such indirect coupling. Neville D et al. 2026
- E4strong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.paper
A higher-order complex containing AF4-family and ENL-family proteins with P-TEFb is normally recruited to KMT2A-target chromatin, and KMT2A fused to an AEP component forms a constitutive hybrid complex causing sustained target expression and transformation.
A higher-order complex containing AF4 and ENL family proteins with P-TEFb facilitates oncogenic and physiologic MLL-dependent transcription. · Yokoyama A, Lin M, Naresh A, Kitabayashi I, et al. · Cancer Cell 2010What this evidence cannot show
Abstract read 2026-09-20. Establishes the structural asymmetry this hypothesis rests on: an AF4-family partner supplies the elongation activity intrinsically rather than recruiting it to a contested surface.
Evidence that argues against this (1)
- AFF1 and AFF4 form distinct SECs with non-overlapping targets, so even within the AF4-family partners the wiring is not one thing, and a partner-level prediction may be too coarse. Lu H et al. 2015
- E5strong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.paper
ENL YEATS reader-dead point mutants F59A and Y78A, which abolish H3K27ac binding, extend latency and lower leukaemia stem cell frequency without abolishing leukaemogenesis, whereas deletion of the YEATS region abolishes it.
The ENL YEATS epigenetic reader domain critically links MLL-ENL to leukemic stem cell frequency in t(11;19) Leukemia. · Hu H, Saha N, Yang Y, Ahmad E, et al. · Leukemia 2023What this evidence cannot show
Abstract read 2026-09-20. Cited as the cheaper falsification route: separation-of-function mutants exist on the ENL partner and could test whether changing what the partner reads changes the collapse order, without generating a new degron line.
Evidence that argues against this (1)
- A mutant that changes leukaemia stem cell frequency changes the cell population as well as the molecular wiring, so an ordering difference measured in such a line could be a cell-state difference rather than a partner-wiring difference. Erb MA et al. 2017
if true, the chain has no single internal ordering and must be drawn per partner
the decisive experiment for this hypothesis
the mutually exclusive AHD is the structural premise
adds collapse order to the list of things that do not transfer from MLL-AF9