iiinova — where your digital self keeps evolving

5) Finally, a panel of other oncogenic RTK fusions shows that only few form condensates, and none show correlation between condensation and signaling at single-cell.

100

4) Unexpectedly, even cytoplasmic *monomers* (const. active) can drive signaling and tumor formation, despite complete lack of multimerization that is essential for condensation.

100

3) If condensates are dispensable, diffuse (non-condensing) ALK fusions should be able to signal and drive tumor growth. Indeed they can! Synthetic, cytoplasmic ALK dimers don’t condense but do signal, and also form tumors in mice.

100

2) There is lots of active oncogene (pALK) in the diffuse phase, and when treated with ALK inhibitor, timescale of decay in the diffuse phase matches that of downstream signals, but decay in condensates is much slower.

100

Evidence of dispensability? 1) EML4-ALK condensation is biphasic (highest at mid levels), but signaling increases monotonically w expression. Thus the strongest signaling is in cells with *no* condensates.

100

The condensate (large foci) model suggests that condensates of fusions (e.g. EML4-ALK) are an essential organizer of signaling in the cyto, in the absence of membrane. Causality is unclear though because perturb's of condensates are often also perturb's of kinase activity.

100

🚨Preprint 2/2: Several studies implicate condensates in RTK fusion onco-signaling. So we were surprised to find that condensates are entirely dispensable😮(!). A study from proteins to mice, by dynamic duo @davidgonzmar.bsky.social and @trmumford.bsky.social. www.biorxiv.org/content/10.6... 👇

12010

Finally, synthetic optogenetic RTK fusions demonstrated the key principles: 1) Grb2 binding and sequestration could be decoupled from fusion signaling, and 2) Grb2 sequestration in the cyto is sufficient to desensitize cells to EGFR stim (without signaling).

101

Targeted therapy (kinase inhibitors) releases Grb2 and relieves suppression. Potential therapeutic importance because drugs permit receptor signaling that promotes tolerance.

100

Mechanism? Fusions sequester adapters like Grb2 in cyto and prevent translocation to active EGFR at membrane. Previously shown for EML4-ALK, below for TPM3-ROS, same for (almost) all fusions tested

100

Main surprise: condensates (large foci) are *not* important for EGFR suppression. For EML4-ALK, condensates co-occur with suppression, but for most fusions -- no condensates. See concurrent preprint for more surprises on the role of condensates in fusions. www.biorxiv.org/content/10.6...

100

Main finding: Across a panel of 10 oncogenic fusions and patient-derived cell lines, active fusions broadly suppressed the cell’s response to EGF.

100

Previously we’d shown that EML4-ALK (onco-fusion) doesn’t only drive cancer signaling—it also suppresses EGFR, and suppression was reversed during therapy. www.nature.com/articles/s41.... Do other fusions also suppress EGFR? Also EML4-ALK formed condensates -- are condensates important?

100

Lukasz Bugaj @bugajlab.bsky.social · 08/12/2025

🚨New preprint(1/2)! We show that RTK fusion oncoproteins broadly suppress EGFR signaling. How? Sequestration of adapters as the shared principle. Led by superb PhD student Carol Gao. www.biorxiv.org/content/10.1... Implications for drug tolerance/resistance, and includes one big surprise🫧.👇

21918

#Cellbio2025 come see the latest from the lab, including on opto/thermogenetics, condensates, RTK fusions, stress granules, biosensors of clustering. Talks and posters today and tomorrow!

186

Lukasz Bugaj @bugajlab.bsky.social · 07/12/2025

#CellBio2025 come check out the workshop on applying optogenetic tools in cell biology! Today (Sunday) at 5:30p in Rm 121. Note that this session does *not* appear in the app for some reason

0104

Lukasz Bugaj @bugajlab.bsky.social · 09/05/2025

Are you interested in synthetic and systems biology approaches to study #spatiotemporal processes in cancer? Join NCI for virtual workshops on May 13, 15, & 20 (next week!) from 12:00–4:30 PM ET: registration: events.cancer.gov/nci/syntheti... #SynSysBio4SpatialCancerResearch

1117

We also developed a "universal" Aviatar that can localize to any GFP-tagged compartment using a weak nanobody, and we used it to localize to stress granules through endogenously-tagged G3BP1

Aviatar regulated actin polymerization at the membrane and revealed compartment-specific differences of RTK fragments commonly mislocalized in cancer

100

Aviatar can target any desired compartment for which a weak binder can be found. We show common targets: plasma membrane, endosomes, Golgi, ER, and microtubules.

In Aviatar (Avidity assisted targeting), a single light-sensitive protein translocates from the cytoplasm to an unmodified subcellular compartment. Light induces clustering, clustering turns a weak binder to a high-avidity assembly, and Aviatar flies to its target location.

130

Lukasz Bugaj @bugajlab.bsky.social · 16/04/2025

Proteins often bind targets using avidity, the combined interaction strength of multiple weak binders. Now we’ve harnessed this principle for compact, *single*-component optogenetic tools for translocation. Lead by phenom PhD student @dennishuang.bsky.social #synbiosky🧵 📝 tinyurl.com/aviatar

1197

New preprint! We found dynamic signal processing (IFFL) in a single protein, which pulses in response to step inputs. New insights for capability of protein computation + implications for new opto and thermo probes www.biorxiv.org/content/10.1... Check out the 🧵 from first author Dennis Huang:

172

and the second www.sciencedirect.com/science/arti...

010

Hmm -- issues with the GIFs above: here is the first! www.sciencedirect.com/science/arti...

We used the thermoPlate to explore stress adaptation to heat shock.We found beautiful stress granule dynamics and memory formation, with differences at each degree of heating. What started as a quick peek at stress granules is now an exciting new avenue of inquiry for the lab.

131

Annoyingly, thermodynamics says heat will not stay confined to individual wells, constraining the complexity of our experiments. But we can predict heat spread dynamics over time with our online app, letting us quickly know if our desired experiment is possible. bugajlab.shinyapps.io/ThermoPlate_...

The thermoPlate addresses the lack of methods for fast/accurate/precise/robust control of temperature for mammalian cells, in high throughput, under a microscope. Like the optoPlate but for temperature, including 96x PID controllers for precision.

Our paper on 96-well control of temperature is now out @cp-cellsystems.bsky.social. Exceedingly proud of this work led by Will Benman, plus Pavan Iyengar, @trmumford.bsky.social, and much of our lab. Come for DIY hardware, stay for the stress granule dynamics and memory🧵 📝 tinyurl.com/thermoPlate

2195

Big congratulations to the phenomenal scientists that made this happen: Will Benman and Dennis Huang @dennishuang.bsky.social, along with assists from many others in lab. Perpetually grateful to work with with such exceptional people. 🙏

030

A major promise of thermal control is the ability for precise, non-invasive communication with cells in vivo. Here, Melt allowed temperature-controlled killing of cancer cells in a mouse, in response to local cooling of the mouse's skin (think ice pack).

100

Melt is highly modular and can endow thermal control over signaling, proteolysis, cell shape, and cell death, among other behaviors, through simple end-to-end fusions. Below, Melt-based induction of cell death in cancer cell lines after cooling by ~10C.

100

Melt is a single protein that both clusters and translocates to the plasma membrane as a function of temperature.

100

Excited and grateful that our initial report on temperature-sensitive protein switches is now published @naturemethods.bsky.social Co-led by PhD students Will Benman and @dennishuang.bsky.social www.nature.com/articles/s41... Brief thread:

2248

000

100

100

Melt is a single protein that both clusters and translocates to the plasma membrane as a function of temperature.

100

000

100

100