Thread by @kaitochondria

I'm super excited to share my 1st first-author paper now out in @ScienceMagazine, where we tackle an age-old question in microbiology: How do dormant spores decide to wake up? Turns out that they can count in their sleep using electricity!😴⚡️
science.org/doi/10.1126/sc…

Bacterial spores are a special type of cell that look and function like microscopic plant seeds: a hyper-resilient DNA bunker that aims to weather bad conditions and reemerge in a better future. They are metabolically dormant, and by all practical means, pretty much dead.

Because of their extreme resistance to most stresses (like heat, UV, dryness, antibiotics), gaining a better understanding of spores has huge implications in healthcare, food safety, and biotechnology.

The spore needs to decide to wake up (or "germinate") if the environment is good, while keeping the option to remain dormant. So the question is: how would a dead cell think? Mind you, the dormant spore is dehydrated & energy deprived, so fancy biomolecular synthesis can't happen

So we started by asking: can dormant spores make smart decisions? We can test this by exposing spores to short pulses of nutrients like that induce germination (we call these "germinants"). Would spores randomly respond, or would they make smart choices based on the past?

By combining microscopy and microfluidics, we then recorded the germination response of thousands of dormant spores that experience these germinant pulses. And it became very clear that spores are able to integrate information and make smart decisions! More food = better wake up.

What are the dormant spores using to integrate information in their sleep? We thought: ions!💡
Our lab has been studying the functions of potassium ions in bacteria, and ions are also central to how our brains operate. Additionally, ion flux itself can be very energy-efficient!

Inspired by integration-and-fire models used in neuroscience, my labmate Colleen Weatherwax and collaborator @jgojalvo generated a mathematical model that matches the experimental results, as well as making key predictions in how the spores will behave if we make some changes.

One of the model's assumptions was that a spore's germination propensity is defined by its internal potassium ion (K+) concentration. We tested this by making mutant spores lacking KtrC, a K+ pump, leading to lower internal K+. Sure enough, the mutants are much more sensitive!

We also made mutants without the YugO K+ channel, or with WT spores germinating in the presence of quinine, a K+ channel blocker. Both of these perturbations led to lower potassium efflux, lower information integration capacity, and lower germination sensitivity.

We also didn't forget to measure K+ itself! We used an intracellular K+ indicator to measure the initial K+ content for each strain, and an extracellular K+ indicator to visualize K+ efflux during our experiments.

Now that we've established K+ content and efflux is important in germination propensity, we wanted to go a step further. If spores are releasing positively charged ions in response to the pulses, wouldn't that change its electrochemical potential? Our model thought as much.

Enter Thioflavin-T (ThT). ThT is a positively-charged fluorescent dye that we've been using extensively to visualize bacterial electrochemical potentials. As the spore releases K+, we should be able to see the spore also getting brighter as more ThT attaches to the spore.

Et voila!

And again, this is all happening while the spore is still dormant. (My favorite #datavis of my PhD!)

You might ask: what if ThT is showing something else than electrochemical potential?
We were EXTREMELY lucky to have benzothiazole expert Emmanuel Theodorakis and his student Jamie Lam just upstairs from our lab, who synthesized a charge-neutral ThT for us! BOOM💥

So to wrap up: we found that dormant bacterial spores are able to integrate external information using a potassium ion-based electrochemical potential mechanism! The manuscript with the whole story and lots more data can be found here: science.org/doi/10.1126/sc… #ScienceResearch

I'd like to thank my co-first author @leticia_gala for joining forces with me and pushing this paper over the finish line with her bottomless enthusiasm; Colleen, Ashley, Jamie, ET, and Jordi for the fruitful collab; and Gürol for his tireless mentorship. Thanks for reading!