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02/05/2026 14:57

Live broadcast from the cell: the dynamic interaction between protein folding helpers and newly produced proteins

Dr. Christiane Menzfeld Öffentlichkeitsarbeit
Max-Planck-Institut für Biochemie

Protein folding helpers such as TRiC and prefoldin (PFD) are crucial for the correct folding of many essential proteins. In a Nature study, researchers from the Max Planck Institute of Biochemistry investigated for the first time interactions between newly synthesized proteins and TRiC/PFD in their native cellular environment. Using single-particle tracking in living human cells, they showed how TRiC and PFD search for nascent chains during translation, assist their folding, and guide them to the native conformation. The study proposes a virtual folding compartment, the “protected folding zone”, which slows chaperone diffusion and promotes efficient folding.

In a nutshell:
- Protein folding helpers such as prefoldin (PFD) and TRiC are responsible for the correct folding of essential proteins such as actin in human cells.

- For the first time, the real-time interplay between newly synthesized proteins and their interaction with TRiC and PFD was analyzed using single particle tracking on a fluorescence microscope.

- Both protein folding helpers repeatedly come into contact with the emerging protein chain – first through short scanning processes and later through longer and stable interactions.

- The protein folding helpers remain close to the client protein at the ribosome in a newly identified protected folding zone.

- The Nature study helps to understand the dynamics of protein folding and misfolding, which is important for processes such as protein aggregation associated with neurodegenerative diseases.

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Protein folding and its helpers

Proteins are the molecular machines of cells. They are produced in protein factories called ribosomes based on their blueprint, the genetic information. Here, the basic building blocks of proteins, amino acids, are assembled into long protein chains. Like the building blocks of a machine, individual proteins must have a specific three-dimensional structure to properly fullfill their functions. To achieve this, the newly produced protein chains in human cells are folded into their stable and functional form with the help of various protein folding helper proteins, known as chaperones, such as TRiC/PFD, or HSP70/40. The protein folding helpers isolate the amino acid chains, which have different chemical properties depending on the amino acid, from the cellular environment. This prevents the newly produced protein chains from clumping together and causing disease.

F.-Ulrich Hartl, a director at the Max Planck Institute of Biochemistry, has spent decades studying the mechanisms of protein folding. Niko Dalheimer, a scientist in Hartl's department and one of the two lead authors of the current study, explains: “Much of what we know about protein folding has been learned from studies conducted in test tubes. However, it is virtually impossible to faithfully replicate the cellular environment in vitro. Unlike a test tube, a cell is a highly complex environment filled with many different macromolecules, like proteins, nucleic acids and lipids. To fully understand how chaperones work, we examined the protein folding dynamics of TRiC and PFD in its natural environment – intact human cells – using single-particle tracking on a fluorescence microscope, an approach that has only recently become feasible thanks to advances in live-cell fluorescent labeling.”

TRiC and prefoldin

A newly synthesized protein is gradually released as an amino acid chain by the ribosomes through a channel. To prevent the newly synthesized protein from clumping together, the free amino acid residues are captured and protected by prefoldin, or PFD for short, a co-chaperone of TRiC. The co-chaperone then passes the protein on to the chaperonin TRiC for folding.

TRiC is a barrel-shaped protein folding helper and is related to the bacterial GroEL/ES. Rongqin Li, scientist and co-first author of the study, explains: "Although TRiC only helps 10% of the proteins in a cell to fold, many of them are particularly important for the cell, including actin and tubulin, which are building blocks of the cytoskeleton. That's why we looked at this part of protein folding. We used actin as a test protein to understand the folding dynamics in cells."

Single particle tracking sheds light on the unknown
In order to follow the real time interaction of all components involved in protein folding, the researchers labeled TRiC and prefoldin, as well as the actin nascent chain as direct chaperone substrate and ribosomes and mRNAs as proxies for chaperone substrates with two colors, green and magenta, in different settings. If the two components were in close proximity, i.e., less than 500 nanometers apart, the colors overlapped and were visible under the microscope as white dots. Niko Dalheimer explains: "There are approximately 10 million ribosomes in a single cell. To enable us to track individual ribosomes and other components under the microscope, we stained only a small proportion of the ribosomes, rather than all of them and used the TIRF method to track the individual molecules and their interactions with chaperones. It is like a diver exploring a pitch-dark deep sea: by shining a light on just a few spots at a time, the diver can get a glimpse of the hidden dynamic life and activity around them.“

What we saw

The scientists observed that TRiC and PFD, repeatedly probe with the newly synthesized actin protein chain emerging from the ribosome for approximately one second. PFD hold the nacent chain shortly before actin is being released from the ribosome and hands it over to the TRiC champer for folding completion. Rongqin Li adds: "Interestingly, the contact between TRiC and actin mutants, i.e., protein chains into which we introduce errors to disrupt its proper folding, was significantly longer. In contrast to the normal condition, the folding-defective actin undergoes multiple-rounds of attempted folding by the chaperonin system and is ultimately targeted for degradation.

F.-Ulrich Hartl summarizes: “For decades, we and others have studied chaperone-mediated protein folding primarily through biochemical experiments, which have been essential for defining how this process is controlled. With live-cell single-particle tracking, we can now examine these concepts directly in living cells. In doing so, we have confirmed key findings from classical biochemical experiments, while at the same time uncovering features – such as the protected folding zone – that could not have been detected with ensemble-based assays. This is the first time these processes have been visualized at the single-molecule level in living cells. As I often tell my colleagues, ‘seeing is believing’.“

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Glossary

Chaperones: French for “chaperone”; a family of proteins that help newly produced proteins fold correctly.

Chaperonins: large barrel-shaped protein complexes that ensure correct, ATP-dependent protein folding. These include TRiC and GroEL/ES

GroEL: large subunit of the chaperonin complex in bacteria
GroES: “lid” subunit of the chaperonin complex in bacteria

Peptide: a molecule consisting of several amino acids (AA). These are linked together by so-called peptide bonds. Peptides are amino acid chains of up to approximately 100 AA. Proteins have the same structure, but have longer amino acid chains (> 100 AA) and are usually more complex and folded into a spatial form.

Prefoldin, or PFD for short: is a co-chaperone that receives newly formed protein chains near the ribosome and protects them from clumping before they are passed on to the chaperonin TRiC. Its name reflects this ‚pre-folding‘ role, highlighting that it acts prior to folding, preparing proteins for their next step in the folding pathway.

TRiC: Abbreviation for T-complex protein ring complex, also known as CCT (chaperonin containing TCP-1); is an essential large protein complex in our cells that acts as a molecular “folding assistant” and helps to bring certain proteins such as actin and tubulin into their correct three-dimensional shape so that they can perform their tasks inside the cell.

TIRF: Abbreviation for total internal reflection fluorescence microscopy; a microscopy method in which only an extremely thin area directly adjacent to a surface, e.g., a cell membrane, is excited with light. As a result, only molecules at this interface light up, while the rest of the sample remains dark, enabling very sharp images with little background light.

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Contact for scientific information:

Prof. Dr. Franz-Ulrich Hartl
Department of Cellular Biochemistry
Max Planck Institute of Biochemistry
Am Klopferspitz 18
82152 Martinsried

office-hartl@biochem.mpg.de
https://www.biochem.mpg.de/hartl

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Original publication:

Rongqin Li*, Niko Dalheimer*, Martin B. D. Müller, and F. Ulrich Hartl: Single molecule dynamics of the TRiC chaperonin system in vivo, Nature, February 2026
*Shared first authorship

DOI: 10.1038/s41586-025-10073-3
https://www.nature.com/articles/s41586-025-10073-3

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More information:

- Video accompanying the press release
https://www.biochem.mpg.de/live-broadcast-from-the-cell

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Shining light on TRiC-mediated protein folding.
Source: Illustration: Marzia Munafo
Copyright: MPI of Biochemistry

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Journalists, Scientists and scholars, Students, all interested persons
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transregional, national
Research results, Scientific Publications
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