A special type of droplet related to stem cell differentiation could provide a target for new cancer therapies

Stem cells are full of potential
.
Their ability to turn into other cell types is critical to our bodies, both during development and throughout life
.
But if that potential goes wrong, it can cause us to collapse and turn some of our most useful cells into malignant cancers
.

In studying the stem cell differentiation pathway, researchers at the University of California, Santa Barbara, found that all the proteins involved did not form assembly lines or rigid structures, but combined into a single droplet
.
Through modeling and manipulation, the team set out to reveal how cells use this droplet to process and transmit information, and how it malfunctions in cancer
.
Their findings were published in PNAS
.

Senior author Max Wilson, assistant professor in the Department of Molecular, Cell and Developmental Biology, said: "The same process that occurs in cells as dew droplets on tissue spider webs causes this liquid molecular computer to appear or disappear
at the behest of a command.
Almost 100% of colorectal cancers are caused by it and are associated with a large number of other cancers
once they go wrong.
”

An important process

Many mechanisms guide stem cells to differentiate into a specific cell type
.
The most important of these is the Wnt pathway, which takes input information from outside the cell, processes it, and passes instructions to the nucleus
.
This initiates a series of actions that tell the stem cells that it is time to differentiate and what type of cells
to become.
Wilson explained that the Wnt pathway is involved in determining the fate of each stem cell, and that it has not changed
in almost all animals.

Stem cell differentiation is very important
during embryonic development.
However, even as adults, there are stem cells in certain parts of our body, such as our intestines, bone marrow, and skin, among others
.
Differentiation pathways play an important role
in the function of these tissues and organs.

Given its importance, Wilson and his colleagues wanted to understand how the Wnt pathway is physically organized
within the cell.
They focused on two proteins — axonin and APC — two proteins that seem to provide scaffolding for the entire process, coordinating the complex dances
of the other proteins.

The functions are as follows

Understanding a protein requires understanding its structure, as the complex geometry of molecules is often as important
as its composition.
So the team fed the chemical composition of the protein into a program called AlphaFold, a deep learning software developed by Google to predict the protein's structure
.

The program returns "a bunch of random spaghetti — the closest AI gets to shrugging and misinformation," Wilson said
.

This suggests that Axin and APC may be working
in a very unconventional way.
Proteins can function in a more fluid, dynamic way, rather than being like a rigid lock and key
.
"We thought they might form intracellular fluids, like droplets of oil in salad dressing," and the only way to do that was to look
.

So they used CRISPR/Cas9 to add fluorescent tags to all the major proteins they knew were part of the Wnt
pathway.
They then observed through a microscope that the proteins condensed into droplets
in the cytoplasm of the cell.

A special pathway

Most of the Wnt pathway occurs in this droplet, which eventually sends a message to the nucleus telling the cell to differentiate
.
It also triggers changes in gene expression that determine the morphology
of stem cells.
"It's like a little liquid computer, right in the middle of a cell," Wilson said
thoughtfully.
Despite its importance, this is the first time scientists have observed the role of
the Wnt pathway.

What's more, the team observed that droplets always formed
next to the nucleus.
In fact, it is located around the central body, which is the structure
that separates chromosomes as cells divide.
Wilson is still exploring avenues in this regard
.
"In my opinion, this suggests that this may be an intrinsic anti-cancer strategy," he said
.
"This could be a way
to coordinate tissue growth, formation and differentiation.
" We don't know yet
.
”

In addition to the other proteins involved in the tissue, axonin and APC appear to be the most important compounds
for the formation of the droplet itself.
At the same time, the Wnt protein is a major participant
in events that occur within the spots.
The droplets are constantly processing a protein called β-catenin, which is produced
elsewhere.
The droplet reads it, modifies it, and sends it back to the cell
.
In addition, there are many helper proteins that play other roles
in the human body.

To their surprise, the team used numerical models to simulate the droplets
.
They showed that concentrating all of the Wnt protein in one droplet was more effective
than spreading it in the cytoplasm.
This droplet is essentially a small reaction chamber
.

Continuing their study, the team adjusted the ratio of
proteins within the droplets.
They found that increasing axonin causes large numbers of small droplets to form inside the cell, rather than on the
centrosome.
At the same time, increasing the APC makes the initial droplets larger
.
The team is actively investigating ways to do this
.

Axonin and APC aren't the only proteins
with this particular behavior.
Wilson said: "Nucleoli and pressure particles are also examples of
liquid protein condensate.
We're just beginning to describe these so-called 'liquid proteins'
.
”

Relationship with cancer

The Wnt pathway is strongly associated
with coloral cancer.
Intestinal epithelial cells can only survive for about three days
before shedding.
They are supplemented
by stem cells located in protected dents in the lining of the intestine.
When replacement is needed, the stem cells divide and one of the daughter cells becomes an intestinal cell
during upward migration.

Mutations in genes involved in the Wnt pathway can cause cells to become cancerous—growth and proliferation unaffected by surrounding cells or environmental conditions
.
Wilson said: "For example, 95% of colorectal cancer patients are missing a large chunk
of the APC gene.
"

Apparently, altering genes somehow altered the dynamics and structure
of Wnt spots.
While both axon and APC contain droplets, the pathway appears to be more sensitive
to problems with the APC gene.
This is another aspect of the system that the team is currently working on
.

There is a great unmet
demand for the treatment of coloral cancer.
"There's no immunotherapy yet," Wilson said
.
"There is no intervention other than chemotherapy
.
"

In fact, the FDA has not approved any drugs
that target the Wnt pathway.
"The fact in itself suggests that we may have been thinking about the wrong thing all along," Wilson said
.
"This new idea of the fluid coordinating the entire process may help us design new drugs and therapies
.
"

Wilson said: "We know that the Wnt pathway is also related to other cancers
.
" The researchers are studying how cancer mutations affect droplet kinetics and mapping all the proteins
involved in the Wnt pathway.
He explained that an appropriate catalogue should provide a list
of possible targets for new therapies.

Ryan S.
Lach, Chongxu Qiu, Erfan Zeyaei Kajbaf, Naomi Baxter, Dasol Han, Alex Wang, Hannah Lock, Orlando Chirikian, Beth Pruitt, Maxwell Z.
Wilson.
Nucleation of the destruction complex on the centrosome accelerates degradation of β -catenin and regulates Wnt signal transmission.
Proceedings of the National Academy of Sciences, 2022; 119 (36)