The “secretome” refers to proteins that are secreted by a cell, tissue or organism. In a new study published in Open Biology, USC stem cell scientist Andy McMahon and his collaborators present an elegant new way to label and study the secretome in a living organism.
“The secretome orchestrates the subtle and complex processes of embryonic development, maintains the function of individual organs, and coordinates organ activity through inter-organ communication,” said McMahon, chair of the Department of Stem Cell Biology and Medicine. regenerative at USC. “However, it can be difficult to know which cells are secreting proteins and which cells are being targeted.”
To address this challenge, co-first authors Rui Yang and Amanda S. Meyer at USC and Ilia A. Druzhinin formerly at Harvard Medical School and now at Scripps Research generated mice with an integrated system to label and track the secretome.
“This integrated system is like a ‘passport system in the body, as we identify where proteins are coming from and where they are going,'” said Druzhinin, who is a Scripps Research Fellow and Principal Investigator in the Department of Molecular Medicine.
To establish this system, the research team began by genetically editing mouse embryonic stem cells to encode an enzyme designed in Alice Ting’s lab at Stanford University. The enzyme, BirA*G3, indiscriminately labels neighboring proteins with the vitamin B7 derivative, biotin. The reason biotin is a useful tag is that it binds tightly to the protein streptavidin, so biotin-tagged proteins can be separated from other proteins in the cell and blood using a readily available technology called “affinity purification of streptavidin”.
The next step was to ensure that BirA*G3 specifically labeled only the secretome proteins. These proteins come from a structure inside the cell known as the endoplasmic reticulum, or ER. Some of these proteins remain in the ER, while others are either secreted or incorporated into the surrounding membrane of the cell.
To specifically tag only these proteins, BirA*G3 itself was tagged with four additional protein building blocks, or peptides. These four peptides, known as KDELs, served as a signal for the cell to retain BirA*G3 in the ER. Thus, when biotin was added, BirA*G3 only added biotin markers to proteins traveling through the ER.
Using these genetically modified stem cells, the team then produced transgenic mice. When these mice were given biotin, the scientists were able to demonstrate BirA*G3 labeling and identify secretomes from a number of organs, including the brain, liver and kidneys. The scientists also identified proteins that were secreted anyway, even though they weren’t marked with the typical signal that the cell should secrete them.
These transgenic mice allow scientists to label and study specific cell types and organs, facilitating studies of inter-organ communication.
“Our new mouse model demonstrates improved labeling efficiency of secreted and ER proteins in specific cell types, providing a valuable resource for mapping and profiling the secretome,” said Meyer, who is a PhD student at the McMahon Lab at the ‘Eli and Edythe Broad Center for Regenerative Medicine and Stem Cell Research at USC.
Yang, a postdoctoral researcher at the McMahon Lab, added, “Our mouse model will facilitate a better understanding of the critical role of the secretome in development, as well as healthy and diseased adult states.”
The mice are now in a public repository, so other scientists can benefit from this new technology.
“Given the central role of key secreted proteins such as insulin, there is great interest in identifying novel secreted proteins,” McMahon said. “Genome studies suggest that there are still many new proteins to be characterized. We look forward to a deep dive into this area now that we’ve validated the technology.
Additional co-authors include: Jinjin Guo and Jill A. McMahon of USC; Namrata D. Udeshi, Dominique K. Carey, Charles Xu, and Steven A. Carr of Harvard’s Broad Institute and MIT; Yanhui Hu and David Rocco of Harvard Medical School; Norbert Perrimon of Harvard Medical School and the Howard Hughes Medical Institute; Qiao Fang of the University of Toronto; Jihui Sha and James Wohlschlegel of UCLA; Shishang Qin from Peking University; and Alice Y. Ting of the Chan Zuckerberg Biohub and Stanford University.