Application

Proteomic labeling (labeling of proteins).

Key Benefits

• Method can be used to express TurboID in any cell type.
• The mouse line can be crossed with any existing Cre line and can be extended to any organ/tissue system.
• Detection of labeled biomarkers can be identified in biofluids as biomarkers of cell specific disease mechanisms.
• Can be used for defining proteomic signatures of tissues, labeling sub-cellular organelle proteins or exosomes, and labeling and detection of proteins in bio-fluids to identify cell-type specific biomarkers.

Technical Summary

Researchers at Emory University have successfully used TurboID, an enzyme capable of biotinylating proteins, to label proteins in adult mice. Proteomic labeling is a valuable research tool for multiple uses including identifying mechanisms of disease, biomarkers, and studying exosomal biology. Utilizing TurboID, the researchers constructed a structure that contained a loxp-STOP site which was fused to a nuclear export sequence to allow for biotin labeling of cytosolic proteins in live mice. The researchers developed a “Cree-flox” strategy and were able to demonstrate its capability by displaying diffuse labeling of proteins in TurboID-NES-flox and Cre transfected HEK293 cells. A targeting vector was then produced to generate TurboID floxed mice, these vectors when injected into mice yielded chimeric mice which were then bred to wild-type C57BL6J mice which produced mice that expressed on copy of the TurboID-NES-floxed allele in the ROSA26 locus. The researchers then injected hSYN1-Cre-AAV9 adenovirus into the hippocampus of the brain, resulting in expression of Cre-recombinase in hippocampal neuron thus confirming that TurboID gene in mice can lead to biotinylation. After a 3-week period the mice were fed water containing biotin and upon brain analysis the researchers found that the mice injected with hSYN1-Cre-AAV9 demonstrated robust biotinylation of the proteome compared to the control mice. Though this method focuses on the brain, the researchers suggest it has wider applications such as identifying cell-type specific biomarkers.

Publication

Shou-Ling Xu, Ruben Shrestha, Sumudu S. Karunadasa, Pei-Qiao Xie, Proximity Labeling in Plants, Annual Review of Plant Biology, 10.1146/annurev-arplant-070522-052132, 74, 1, (285-312), (2023).