Researchers analyze how clusters of molecules meet inside cells

A developing area studies how clusters of molecules gather inside of cells, similar to how oil droplets form and separate from the water in a vinaigrette.

Researchers analyze how clusters of molecules meet inside cells
Dense liquid droplets create squishy circuits in cells, with implications for neurologic diseases. Image Credit: Liam Holt, NYU Langone Health

When similar, big molecules clump together into dense droplets and separate from the more diluted portions of the fluid cell interior, “liquid-liquid phase separation” happens in human cells. Previous research had hypothesized that evolution used these “condensates” organic creation to organize cells, supplying, for instance, isolated regions for the construction of cellular machines.

Furthermore, cells from people with neurological diseases, such as Alzheimer’s disease, almost invariably include aberrant, condensed—also termed “tangled”—groups of molecules in droplets. While the exact cause of such condensates is unknown, a recent idea claims that aging alters the biophysical characteristics of cell interiors, in part because of “molecular crowding,” which involves packing more molecules into the same areas in order to affect phase separation.

Since both identify and compute reactions based on incoming information, researchers compare condensates to microprocessors, computers integrated into circuits.

The field has failed to explain the processes linking phase separation, condensate formation, and computation based on chemical signals, which happen at a much smaller level, researchers claim, despite the anticipated effects of physical changes on liquid processors.

This is because experiments have a hard time separating all of the functions performed by natural condensates.

To overcome this obstacle, scientists from the German Center for Neurodegenerative Diseases and NYU Grossman School of Medicine created an artificial system that demonstrated how the production of condensates alters the molecular behavior of the enzyme kinases, an example of chemical computation. Protein switches known as kinases affect cellular functions by phosphorylating—adding a molecule known as a phosphate group—target molecules.

The latest study, which was made available online on September 14 in Molecular Cell, discovered that the development of artificial condensates during phase separation provided additional “sticky” areas where medically significant kinases and their targets could engage and cause phosphorylation signals.

Our study results show that physical changes like crowding can drive condensate formation that is converted into biochemical signals as if condensates were squishy computers.”

Liam J. Holt, PhD, Study Lead Author and Member, Institute for Systems Genetics, NYU Langone Health

Cyclin-Dependent Kinase 2, which phosphorylates the microtubule-binding protein Tau, was one of the research kinases observed to be more active in a packed, condensed environment. Patients with Alzheimer’s disease commonly have tangled condensates of Tau in their brain tissue.

Our experiments suggest that the formation of more Tau condensates drives more Tau phosphorylation. Whether these mechanisms lead to more brain cell death, and whether reversing them could be a new treatment approach, will be important questions in our upcoming work,” adds Dr. Holt, who is also an associate professor in the Department of Biochemistry and Molecular Pharmacology.

In particular, the study discovered that there was a three-fold acceleration of phosphorylation at a set of Tau sites (the AT8 epitope associated with Alzheimer’s disease) when Tau and Cyclin-Dependent Kinase condensed together into dense droplets.

Engineering a biosensor

The study team investigated various artificial condensates, synthesizing various scaffold molecules to determine which effectively pulled sample kinases—MAPK3, Fus3, and Cyclin-Dependent Kinase 1 (Cdk1)—together with their targets to improve signaling in an effort to create usable versions of these computers.

Scaffold molecules mesh inside droplets to generate condensates. The team discovered that in their model, phosphorylation reactions were hundreds of times faster when big macromolecules congregated into droplets inside yeast, a single-celled living creature.

The study also discovered that condensate production enabled the included kinases to phosphorylate a wider variety of molecules without the typical need for certain molecular structures. According to scientists, this shows that condensates in crowded cells create changed computation types, some of which may be linked to diseases.

The current work in Dr. Holt’s laboratory discovered that a protein complex called mTORC1 regulates molecular crowding by regulating the number of ribosomes, or “machines,” which are used by cells to produce other big proteins. The group will investigate whether substances that block mTORC1 can lessen crowding and Tau phosphorylation.

The research team also expects that their discoveries may improve the development of additional cellular computers that respond to external influences. This may involve implanting designed processors into immune cells, which would activate as they tried to fit into tissue made thick by developing tumors in order to target cancer cells.

Source:
Journal reference:

Sang, D., et al. (2022) Condensed-phase signaling can expand kinase specificity and respond to macromolecular crowding. Molecular Cell. doi.org/10.1016/j.molcel.2022.08.016.

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