Scientists at the Center for Molecular Medicine Cologne (CMMC) and the CECAD Cluster of Excellence on Aging Research have demonstrated in a biomedical study that ribosomes, the protein factories of our cells, face an unusually tough challenge when building large, membrane‑embedded proteins. When protein synthesis fails, ribosomes recruit quality control factors to eliminate the resulting defective proteins. These findings shed new light on the physiological roles of protein quality control at the ribosome. The study has now been published under the title 'Principles of ribosome-associated protein quality control during the synthesis of CFTR' in The EMBO Journal.
Proteins are molecular machines that control and execute virtually all cellular processes. A typical human cell synthesizes over 10,000 different proteins; about a quarter of these represent transmembrane proteins that mediate critical cellular processes, including cell signaling, adhesion, and transport of a variety of molecules into and out of the cell. Cells employ thousands of protein quality control factors to build and maintain proteins in the correct shape and to eliminate defective proteins, as the latter tend to form potentially toxic aggregates that are characteristic of many neurodegenerative diseases.
The ribosome-associated quality control (RQC) pathway plays a critical role in maintaining proteins in a healthy, functional state: it is activated when ribosomes get stuck during the decoding of messenger RNAs (mRNAs) into amino acid chains. The resulting truncated proteins are likely to be dysfunctional and could interfere with normal cellular processes. Therefore, the RQC pathway marks these proteins for quick elimination. If this important quality control pathway does not work properly, defective and potentially toxic proteins accumulate. "RQC dysfunction leads to neurodegeneration and contributes to systemic health decline during aging. Although it is fairly clear how the pathway functions, we still lack a comprehensive understanding of the causes of ribosome stalling that trigger RQC," says principal investigator Dr Débora Trentini.
The cellular membrane, which separates the contents of the cell from the surrounding environment, is made of oily lipids. Transmembrane proteins have specific chemical properties that enable them to reside in this lipid environment rather than the watery interior of the cell. "Our research showed that transmembrane proteins are inherently difficult to synthesize: some of the ribosomes translating highly hydrophobic membrane-spanning protein segments become stuck and never finish the job they started. The RQC pathway marks these translation-arrested transmembrane proteins for elimination even before they leave the ribosome, thereby limiting their toxic effects," Trentini added. The new findings reveal an intrinsic pitfall of an essential cellular process and shed light on a novel physiological role of protein quality control at the ribosome: dealing with difficult transmembrane proteins.
This new finding may have implications for the treatment of genetic diseases, as many of them stem from mutations in transmembrane proteins. A notable example is cystic fibrosis, a hereditary condition that causes several organs to malfunction. It is caused by an insufficiency of CFTR, a transmembrane protein that moves chloride ions across epithelial cell membranes. This activity helps to control salt balance, water movement, and mucus hydration in organs like the lungs, pancreas, and intestines. "We were interested in investigating whether the translation process of CFTR is also prone to stalling and, if so, whether disease-associated mutations and cystic fibrosis drug therapies influence this issue," says Dr Tom Oldfield, first author of the study.
To measure how many ribosomes successfully complete CFTR translation, the team engineered an artificial CFTR mRNA containing a green and a red fluorescent protein positioned before and after the CFTR coding sequence. This "reporter" was then introduced into cultured human cells. When a ribosome translates the entire message without interruption, the cell produces equal amounts of green and red fluorescence. But if ribosomes stall during CFTR translation, only the green protein is made, resulting in a stronger green signal than red. Using this method, the team observed that a small portion of CFTR translation events arrest during translation, regardless of the presence of disease-associated mutations or cystic fibrosis medications. This suggests that translation arrest and RQC activation could in principle limit the levels of CFTR chloride channels in the context of disease. The study paves the way for future research into potential interventions that could help ribosomes overcome the challenge of translating membrane-spanning protein segments.
The work was supported by the Fritz Thyssen Foundation, the German Research Foundation (DFG) within the framework of Collaborative Research Centre 1678 'Systems-level consequences of fidelity changes in mRNA and protein biosynthesis', the CMMC, and the Köln Fortune Program.
