The EGR1 promoter fails its own IgG control — an attractive claim withheld because a pre-declared control said no
IRX1 is supposed to switch on EGR1, so we looked. But the negative control at EGR1 was just as high as the real signal, which means the measurement there is background. We did not report it.
The same thing, for a scientist
Kühn reports IRX1 induction raising EGR1 8-fold, EGR2 2.5-fold and EGR3 4.5-fold in HEK293T, with EGR proteins driving a p21-dependent quiescence program that could explain the iALL-IRX1 stem-like phenotype [E1]. EGR1 was therefore a pre-declared locus of interest in this beat. It reads **4.5-11.1x** mean promoter in KMT2A-N across all seven patients — which looks like strong fusion occupancy — but **its IgG control reads 1.8-14.7x at the same window**, giving KMT2A-N/IgG ratios of **1.11, 3.90, 1.16, 1.13 and 0.50** in the five samples where IgG is non-zero [E2]. The EGR1 promoter is a background hotspot in this assay and **no EGR1 claim is made in this beat.**
The numbers, including the one below 1
| patient | EGR1 IgG | EGR1 KMT2A-N1 | N1/IgG |
|---|---|---|---|
| p279 | 5.84 | 6.50 | 1.11 |
| p318 | 2.63 | 10.27 | 3.90 |
| p454 | 6.59 | 7.64 | 1.16 |
| p179 | 6.08 | 6.86 | 1.13 |
| p247 | 14.71 | 7.35 | 0.50 |
| TB920488 | 1.82 | 4.77 | 2.62 |
| TB051939 | 0.00 | 4.48 | IgG = 0 |
| TB986292 | 0.00 | 7.52 | IgG = 0 |
In p247 the IgG is twice the specific antibody. A locus where the negative control outperforms the antibody is not measuring the antibody's target.
Why this is worth an entry rather than a footnote
Without the IgG column, EGR1 at 4.5-11.1x would have been written up as "the fusion occupies EGR1 in every infant t(4;11) patient" — a clean, quotable, mechanistically satisfying result that connects Kühn's stem-cell-quiescence model to primary patient chromatin. It is an artefact of a promoter that is hot in every CUT&RUN library including the ones with no primary antibody.
EGR1 is a canonical immediate-early gene with a highly accessible, CpG-rich promoter, which is the standard profile of a CUT&RUN and ChIP background hotspot. The general rule: run the IgG at the specific locus you intend to report, not just as a global summary. A genome-wide IgG correlation would have looked fine here; the locus-level one did not.
| locus | EGR1 promoter, chr5:138,461,478-138,469,478 (TSS +/- 4 kb, hg38) |
|---|---|
| verdict | background hotspot; no EGR1 claim made |
kmt2an range
[ 4.48, 11.12 ]
igg range
[ 0, 14.71 ]
n over igg — the numbers behind the chart
{
"p279": 1.11,
"p318": 3.9,
"p454": 1.16,
"p179": 1.13,
"p247": 0.5,
"TB920488": 2.62
}- A quiescence program driven by EGR would predict that IRX1-high blasts are less sensitive to cycle-dependent chemotherapy, which is a testable and clinically consequential claim - and it currently rests on a HEK293T Q-PCR whose own ChIP was negative.
- How many loci in this deposit are background hotspots by the same test? A genome-wide IgG-versus-specific screen would give the list and has not been run.
- Do EGR2 and EGR3, which are much colder (0.00-0.43x in KMT2A-N), clear their own IgG? They are too close to zero for the ratio to be meaningful either way, so the EGR family as a whole is unmeasured here rather than negative.
- E1moderate confidence 3 of 5: One strong study, or several smaller consistent ones.paper
IRX1 induction in HEK293T raises EGR1 8-fold, EGR2 2.5-fold and EGR3 4.5-fold, and EGR proteins control a p21-dependent quiescence program in haematopoietic stem cells.
The IRX1/HOXA connection: insights into a novel t(4;11)-specific cancer mechanism · Kuhn A, Loscher D, Marschalek R · Oncotarget 2016What this evidence cannot show
HEK293T transgene overexpression with Q-PCR; the link from EGR to HSC quiescence is imported from other literature and cited, not measured in that paper. IRX1 ChIP could NOT confirm the EGR genes as direct targets ('we could not confirm this with ChIP experiments (data not shown)').
Evidence that argues against this (1)
- The paper itself reports the EGR genes are not direct IRX1 targets by ChIP, so the 8-fold induction is an indirect effect at least one step removed from IRX1 binding. Kuhn A et al. 2016
- E2strong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.dataset
In GSE252378 KMT2A::AFF1 samples the EGR1 promoter reads 1.8-14.7x mean promoter in IgG, giving KMT2A-N/IgG ratios of 0.50-3.90, so EGR1 fails its own background control.
q145 re-analysis of GSE252378: IRX1 and HOXA oncoprotein occupancy across seven KMT2A::AFF1 patients (analyses/q145-irx1-in-hoxa-negative-infants) · Iqbal Lab oncofusion knowledge graph, beat q145 · internal analysis over deposited GSE252378 bigWigs 2026What this evidence cannot show
IgG libraries in this deposit are shallow (57-94% of promoters read zero), so an IgG value at a hot promoter is estimated from few fragments and is itself noisy. That noise cuts both ways but does not rescue p247, where IgG exceeds the specific antibody two-fold. Every number is a WITHIN-SAMPLE share: an interval's mean coverage divided by that same library's mean promoter coverage, so 1.0 = an average promoter in that library. **GSE252378 has no spike-in and no drug arm**, so no between-sample absolute statement is licensed; the only cross-sample quantity used is a ratio of two loci measured inside one library, where the unknown per-library scale cancels.
Evidence that argues against this (1)
- If the IgG libraries are systematically enriched for open chromatin by a mechanism that does not affect the specific-antibody libraries, the ratio understates true EGR1 occupancy and the withheld claim might be real. Janssens DH et al. 2024
a pre-declared control that failed and suppressed an attractive claim
- related to →IRX1 — Iroquois homeobox 1, the gene that names the HOXA-negative half of infant t(4;11) ALL
EGR1 is Kuhn's indirect IRX1 target and could not be measured here