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Reposted by Travis Fleming

Alexis Caulier @hemagene.bsky.social · 03/04/2025

Out today in @science.org! What if you could chart cells' regulatory programs at unprecedented resolution? In my work with Jorge Martin-Rufino from the @bloodgenes.bsky.social lab, we dissect the genome’s control circuits and find where key genetic variation hides bit.ly/3YhBMoO

bit.ly

Transcription factor networks disproportionately enrich for heritability of blood cell phenotypes

Most phenotype-associated genetic variants map to noncoding regulatory regions of the human genome, but their mechanisms remain elusive in most cases. We developed a highly efficient strategy, Perturb...

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Travis Fleming @tjflemin.bsky.social · 03/01/2025

Thanks so much @dpastoors.bsky.social! Excited to see two great stories converging on the same biology :)

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Reposted by Travis Fleming

Dorien Pastoors @dpastoors.bsky.social · 03/01/2025

Congrats @tjflemin.bsky.social, @bloodgenes.bsky.social and coauthors! I feel like preprint directly ties together #HemeSky 's favorite two transcription factors: MECOM and CEBPA. As to why these two: I'm heavily biased as our complementary work will also be on biorxiv soon (monday!!)

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Reposted by Travis Fleming

Vijay G. Sankaran @bloodgenes.bsky.social · 02/01/2025

Wonderful to have this work led by @tjflemin.bsky.social along with other colleagues in our laboratory, as well as great collaborators, out on @biorxivpreprint.bsky.social! Stay tuned for exciting complementary work from @dpastoors.bsky.social, Ruud Delwel, and co coming soon!

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

Joao Paulo, Steven Gygi, @camimoso.bsky.social, Karen Adelman, Jennifer Perry, Yana Pikman, Kimberly Stegmaier, @nubama.bsky.social, @kmachlus.bsky.social, @hovestadt.bsky.social, Andrea Arruda, Mark Minden, and Richard Voit.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

Just as important, none of this work could have been possible without immense help and mentorship from my good friends, collaborators and co-authors: @mantoszewski.bsky.social, Sander Lambo, Michael Gundry, Riccardo Piussi, Lara Wahlster, Sanjana Shah, Fiona Reed, Kevin Dong...

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

This work is the culmination of my PhD thesis in @bloodgenes.bsky.socials’ lab. I am incredibly grateful for the amazing support and mentorship Vijay has provided me over the years, allowing me to freely pursue my scientific interests.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

I’m incredibly optimistic that these approaches can be extended across other leukemias and solid tumors, enabling therapeutic differentiation of malignant cells by rewiring dysregulated transcriptional activity.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

In summary, we demonstrate how synergistic use of targeted protein degradation, functional genomic perturbations, and high-throughput screens can distill the functional significance of a complex oncogenic signaling network into a single, pivotal gene regulatory node.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

Strikingly, activating this cisRE significantly impaired the engraftment ability of primary AML cells, highlighting the therapeutic potential of reactivating myeloid differentiation programs to disrupt the fitness of stem cell-like leukemia cells.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

Finally, we transplanted these cells into immunodeficient mice to assess how CEBPA cisRE activation impacted leukemia burden and engraftment of modified cells.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

The approach yielded a modest increase in CEBPA expression itself. However, this transient and subtle activation was sufficient to induce significant differentiation phenotypes during ex vivo culture.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

We next assessed if activation of this cisRE alone was sufficient to induce differentiation of these primary AMLs. Here, we delivered CRISPRa mRNA and the same guide RNAs targeting this cisRE.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

Remarkably, across a panel of patient samples, MECOM KO induced significant loss of stem cell-like leukemia cells, while inactivation of the CEBPA cisRE could almost completely rescue this phenotype and maintain cells in more stem cell-like states.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

Nonetheless, we wanted to determine if the functional link between MECOM and this CEBPA cisRE was conserved in primary AML samples. To do so, we knocked out MECOM and co-inactivated this cisRE, hypothesizing that cisRE inactivation should rescue MECOM-KO-induced differentiation.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

These functional screens and validation suggested a previously unappreciated and surprisingly simple regulatory logic underlying MECOM’s role in promoting stem cell-like states in AML through repression of a single critical cis-regulatory element.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

We then performed an orthogonal screen using CRISPRa, asking if in the absence of MECOM perturbation, could activation of any single cisRE be sufficient to induce myeloid differentiation? Surprisingly, the only hit from this screen was the same cisRE linked to CEBPA!

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

We first leveraged a CRISPRi screen to determine if repressing any single cisRE is sufficient to maintain cells in a CD34+ stem cell-like state, even after MECOM degradation. The most significant hit from this screen was a cisRE 42 kb away from the myeloid TF CEBPA.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

But which parts of these MECOM-regulated networks are functionally important? This motivated us to perform functional genomic screens to identify MECOM controlled cisREs that are essential in facilitating MECOM’s ability to block differentiation in stem cell-like leukemia cells.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

We hypothesized that if our gene and cisRE networks are repressed in primary AML, MECOM-driven stem cell-like states would be anti-correlated with their activity. Indeed, analyses of single cell genomics data of a large AML patient cohort confirmed this hypothesis.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

We defined conserved networks of genes and cis-regulatory elements (cisREs) under the direct repression of MECOM. However, we ultimately wanted to determine whether these MECOM-repressed gene and cisRE networks were relevant in primary AMLs.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

We then profiled the direct molecular changes that occur upon MECOM loss. Interestingly, we observed significant increases in gene expression and chromatin accessibility hours after degradation, suggesting MECOM functions predominantly as a transcriptional repressor in AML.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

Notably, MECOM degradation resulted in striking myeloid differentiation phenotypes and eventually cell death, presumably due to the loss of stem-like, self-renewal capabilities.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

To characterize MECOM’s direct function, in the absence of confounding cell-state alterations, we engineered a suite of AML cell lines with an endogenous MECOM-FKBP12F36V degron. These models enable rapid and specific degradation of all MECOM protein in AML cells within minutes!

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

As shown by Ruud Delwel and co, HSC gene expression programs in AML are frequently driven by increased expression of MECOM, a TF that plays a key role in normal HSC maintenance. However, the mechanisms by which MECOM drives stem cell-like, high-risk features remained unclear.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

This prompted us to ask: How can we better understand the molecular features that drive stem cell-like states in AML?

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

For some AMLs, therapies like retinoids and menin inhibitors can induce differentiation and improve patient outcomes. But there is still a significant lack of targeted approaches to induce therapeutic differentiation of some of the most primitive AMLs with the worst prognoses.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

As demonstrated by John Dick/Andy Zeng and other colleagues, HSC gene expression programs in AML drive primitive cell states linked to poor patient prognosis and high risk of relapse.

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Travis Fleming @tjflemin.bsky.social · 02/01/2025

Stem cell-like states confer poor outcomes in blood cancer—but what mechanisms drive this and how can they be therapeutically targeted? In our new preprint, we show how a single TF represses one enhancer to maintain a subset of high-risk leukemias: 🧵👇 www.biorxiv.org/content/10.1...

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Travis Fleming @tjflemin.bsky.social · 11/12/2024

Thank you so much Mike! Great connecting yesterday :)

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Reposted by Travis Fleming

Vijay G. Sankaran @bloodgenes.bsky.social · 11/12/2024

🙏 🙏 Honored to have the work from @tjflemin.bsky.social from our lab featured in the Best of #ASH24!!!

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Travis Fleming @tjflemin.bsky.social · 10/12/2024

Had a blast presenting our work today at #ASH24! Very grateful for my mentors and colleagues for their support. Stay tuned very soon for more on this study on how we’ve functionally dissected high-risk leukemias

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Reposted by Travis Fleming

Vijay G. Sankaran @bloodgenes.bsky.social · 09/12/2024

Please come see @tjflemin.bsky.social present on his groundbreaking work at @ash-hematology.bsky.social #ASH24 that reveals a surprisingly simple regulatory logic for high-risk AMLs ... Happening soon in Manchester Grand Hyatt San Diego Seaport Ballrooms ABCD: ash.confex.com/ash/2024/web...

ash.confex.com

Paper: Targeted Protein Degradation Reveals a Repressive Role of Mecom at the <em>CEBPA</Em> Locus to Prevent Differentiation in High-Risk Acute Myeloid Leukemia

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Reposted by Travis Fleming

Vijay G. Sankaran @bloodgenes.bsky.social · 09/12/2024

#ASH24 Please check out these two abstracts from our laboratory! Travis Fleming will present his talk this afternoon and Alexis Caulier will present his poster this evening!

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Travis Fleming @tjflemin.bsky.social · 05/12/2024

Roll out of the hotel bed at 5pm on Monday and come listen my talk! Our CRISPR screening to elucidate mechanisms of #AML stem cell maintenance is guaranteed to calm your throbbing headache. @ash-hematology.bsky.social

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Travis Fleming @tjflemin.bsky.social · 05/12/2024

Anticipating a hefty hangover from those hot Miami Nights at #ASH24 this weekend? Have no fear! I have the solution. Doctors hate this one trick: 🧵👇

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