A high-throughput imaging platform reveals the effect of mutations on protein location

The team developed a high-throughput imaging platform to assess the effect of nearly 3,500 mutations on protein location. They found that about one in six disease-causing mutations cause proteins to end up in the wrong place in the cell.

Technological advances in genetic sequencing have allowed researchers to identify thousands of disease-causing protein mutations. We can now identify these mutations in clinic patients, but we have no idea of ​​their consequences for cellular processes. This study was intended to help address this knowledge gap.”

Jessica Lacoste, co-author of the study and a postdoctoral fellow at the Donnelly Center for Cellular and Biomolecular Research at the University of T.

The study was recently published in the journal Honeycomb.

There are several ways genetic mutations can affect the proteins produced in the cell. For example, they can reduce their overall stability by disrupting their ability to fold, altering their interactions with other proteins, or disrupting their movement to different regions of the cell. While the first two effects have been fairly well studied, much less is known about the third. Improving our understanding of the effect of mutations on protein localization is important for elucidating the critical role of this malfunction in a wide range of human diseases.

The research team used a powerful microscope -; as well as computational analysis to fill in the gaps in their visual analysis -; to compare the cellular journeys made by the mutated proteins with those made by the normal proteins. Through these methods, they learned that mislocalization is much more common than previously thought.

The researchers expected the proteins to be in the wrong place because of disruptions in their interactions with other proteins or because of signaling that would normally direct them to the right place. They were surprised to learn that the main causes of protein mislocalization were, in fact, destabilization of proteins and loss of their ability to integrate into membranes.

“We have created the first large-scale map to visualize the effect of mutations on the localization of a protein in a cell,” said Mikko Taipale, co-principal investigator of the study and professor of molecular genetics at the Donnelly Center and T. Temerti University. Faculty of Medicine. “No one else has investigated the effects of pathogenic missense mutations on such a scale, where we tracked the movement of proteins to different organelles. The patterns of mislocalization we observed help explain the severity of disease caused by certain mutations and improve our understanding of mutations that have been less studied.”

Although protein mislocalization is not as well understood as a general loss of protein stability or altered interactions with other proteins, it occurs almost as frequently. The mutation, most commonly associated with cystic fibrosis, causes the affected protein to enter the cell’s endoplasmic reticulum, where it remains instead of moving to the correct location on the cell surface. Drug therapy that promotes the proper exchange of the mutant protein is now being used in the clinic to address this problem and improve patients’ symptoms.

“We’ve made our protein mislocalization database available as a comprehensive resource that other researchers can use to expand our collective knowledge of how genetic variation affects human disease,” said Ann Carpenter, co-principal investigator of the study and senior director of the Imaging Platform at the Broad Institute. . “One particularly useful application of these data would be to identify compounds that could help mutant proteins to localize correctly for the treatment of rare diseases.”

This research was supported by the Canadian Institutes of Health Research, the Texas Institute for Cancer Prevention and Research, the National Institutes of Health, the Ontario Ministry of Research and Innovation, the Susan G. Komen Foundation, and the Connaught University of Toronto Foundation.