The School of Medicine spotlights a recently published faculty research paper each month. The goals are to highlight the great research happening at OHSU and to share this information across departments, institutes and disciplines.

A new era in cell cartography

The School of Medicine’s Paper of the Month for January 2019 is “VIPER is a genetically encoded peptide tag for fluorescence and electron microscopy” published in Proceedings of the National Academy of Sciences (PNAS). This project was led by Kimberly Beatty, Ph.D., associate professor of biomedical engineering, OHSU School of Medicine, and authors are Julia Doh, Jon White (Longwood U.), Hannah Zane, Dr. Young Hwan Chang, Dr. Claudia Lopez, Dr. Caroline Enns and Dr. Beatty.

Scientists seek to image – or take pictures of – proteins inside cells in order to gain novel insights into their function in both normal processes and disease-associated processes.

The problem? Proteins are generally invisible. How can cellular proteins be labeled or tagged in order to image them using fluorescence and/or electron microscopes (EM)?

This study builds on the ongoing collaboration between the labs of Drs. Enns and Beatty and sought to answer that question by developing peptide tags that could be used to localize individual proteins in their cellular environment.

Previous research has shown this is possible. The Beatty Lab had previously developed a VIP Y/Z tag for fluorescence microscopy. Researchers outside OHSU have also developed genetically encoded EM tags, such as APEX, miniSOG or the tetracysteine tag.

“Those are inherently one-color because they all use DAB-precipitation for contrast,” said Dr. Beatty.

So the team designed a new genetically encoded peptide tag and named it VIPER, or VIP coilE/coilR. VIPER consists of a heterodimeric coiled-coil between a genetically encoded peptide tag (CoilE) and a reporter-conjugated peptide (CoilR). The VIP probe peptide, CoilR, has a handle for conjugation reactions, making it capable of being linked to any color fluorophore or other types of reporter molecules (e.g., gold). This is one of the key aspects that makes the “versatile” aspect of Versatile Interacting Peptides.

The team validated that VIPER could light up cell structures, such as the actin cytoskeleton and the nucleus.  VIPER could also be used to observe iron uptake.  They demonstrated that VIPER could deliver different colors, for fluorescence microscopy, or small nanoparticles (“quantum dots”) for correlated fluorescence and electron microscopy.

“The big step forward here is finding a way to tag these proteins that allows visualization either with fluorescence or electron microscopy,” said Mary Heinricher, Ph.D., associate dean for research, OHSU School of Medicine.

VIPER’s compatibility with various chemical reporters creates a flexibility unmatched by the DAB-based tags. However, the researchers emphasize that VIPER does not replace or supersede other genetically-encoded tags.

“VIPER augments other labeling methods, such as immunolabeling or fluorescent proteins, to enable researchers to tag and track multiple distinct targets at once,” wrote the authors. “We anticipate that this enhanced microscopy toolkit will facilitate the generation of more detailed and informative maps of cellular proteins, ushering in a new era in cell cartography.”

The team is now using the new tags to determine how cells sense and respond to iron.  Additionally, they are working with faculty in the OCSSB to study the HER2 signaling network in breast cancer cells. In both applications of the technology, the goal is to understand the molecular mechanisms in cells that lead to human diseases.  With detailed pictures, there will be new opportunities to define new drug targets or to implement new strategies for treating diseases.

Citation

Doh, J. K.; White, J. D.; Zane, H. K.; Chang, Y. H.; López, C. S.; Enns, C. A.; Beatty, K. E.: VIPER is a genetically encoded peptide tag for fluorescence and electron microscopy. Proc. Natl. Acad. Sci. U.S.A. 2018, 115, 12961-12966.

Related

• Read the paper
• Visit the Beatty Lab
• Visit the Enns Lab