Related Experiment Video
Updated: Jul 25, 2025
12:19
Mapping Molecular Diffusion in the Plasma Membrane by Multiple-Target Tracing MTT
Published on: May 27, 2012
17.3K
Dynamic mapping of proteome trafficking within and between living cells by TransitID
Wei Qin1, Joleen S Cheah1, Charles Xu2
1Departments of Biology, Genetics, and Chemistry, Stanford University, Stanford, CA 94305, USA.
Cell
|June 29, 2023
Summary
TransitID enables mapping of thousands of proteins within living cells. This proximity labeling method reveals previously hidden cellular trafficking pathways and functions, like stress granule roles in protecting proteins.
Area of Science:
- Cell biology
- Proteomics
- Molecular biology
Background:
- Mapping endogenous protein trafficking in living cells is crucial for understanding cellular mechanisms.
- Current methods like microscopy and mass spectrometry have limitations in scale and resolution for dynamic proteome trafficking.
Purpose of the Study:
- To develop and validate TransitID, a novel method for unbiased mapping of endogenous proteome trafficking.
- To achieve nanometer spatial resolution in identifying protein movement within living cells.
Main Methods:
- TransitID utilizes two proximity labeling (PL) enzymes, TurboID and APEX, targeted to specific cellular compartments.
- Sequential addition of small-molecule substrates enables tandem PL, followed by mass spectrometry to identify dual-tagged proteins.
Main Results:
- TransitID successfully mapped proteome trafficking between cytosol-mitochondria, cytosol-nucleus, and nucleolus-stress granules.
- The study uncovered a protective role for stress granules in shielding the JUN transcription factor from oxidative stress.
- Intercellular signaling proteins between macrophages and cancer cells were identified.
Conclusions:
- TransitID provides a powerful, high-resolution approach for analyzing endogenous protein trafficking in living cells.
- The method distinguishes protein populations based on their cellular compartment or origin.
- TransitID opens new avenues for discovering previously invisible cellular biology and intercellular communication.
Related Concept Videos
01:19Protein Dynamics in Living Cells
2.2K
Different fluorescence-based techniques are used to study the protein dynamics in living cells. These techniques include FRAP, FRET, and PET.
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...
2.2K
01:45Overview of Protein Sorting and Transport
11.6K
Eukaryotic cells have different membrane-bound organelles with distinct protein requirements. The process by which proteins are targeted to a specific organelle is called protein sorting.
Protein sorting can be of two types: signal-based sorting and vesicle-based trafficking. In signal-based sorting, specific amino acid sequences called sorting signals target proteins to the proper location inside the cell either via gated transport or by protein translocation. In gated transport, folded...
Protein sorting can be of two types: signal-based sorting and vesicle-based trafficking. In signal-based sorting, specific amino acid sequences called sorting signals target proteins to the proper location inside the cell either via gated transport or by protein translocation. In gated transport, folded...
11.6K
01:19Translocation of Proteins into the Mitochondria
3.2K
Mitochondrial precursors are translocated to the internal subcompartments via independent mechanisms involving distinct protein machineries called translocases.
Sorting of outer membrane proteins:
Mitochondrial outer membrane proteins are of two types: the transmembrane, beta-barrel porins, and the membrane-anchored, alpha-helical proteins. Beta-barrel porin precursors are translocated by the TOM complex and inserted into the outer mitochondrial membrane by the SAM complex. In contrast,...
Sorting of outer membrane proteins:
Mitochondrial outer membrane proteins are of two types: the transmembrane, beta-barrel porins, and the membrane-anchored, alpha-helical proteins. Beta-barrel porin precursors are translocated by the TOM complex and inserted into the outer mitochondrial membrane by the SAM complex. In contrast,...
3.2K
01:24Protein Diffusion in the Membrane
4.4K
Proteins show rotational as well as lateral diffusion across the membrane. The lateral diffusion of proteins was confirmed through the cell fusion experiment where mouse and human cells were fused, resulting in hybrid cells. When the human and mouse cells fused, the specific membrane proteins on human and mouse cells were marked with the red and green-fluorescent markers, respectively. Initially, the red and green fluorescence was located on the respective hemisphere of the cell. As time...
4.4K
01:34Protein Transport into the Inner Mitochondrial Membrane
3.8K
Nuclear encoded mitochondrial precursors are imported to the inner membrane in a multistep process involving two separate translocons, TIM22 and TIM23. TIM23 is a cation-selective pore that remains closed by the N terminal segment of the protein. Negative charges on the TIM23 act as a receptor for the incoming precursor, pulling the positively charged matrix-targeting sequence for peptide insertion and translocation.
Transport of mitochondrial precursors across the TIM23 channel is driven by...
Transport of mitochondrial precursors across the TIM23 channel is driven by...
3.8K
01:27Post-translational Translocation of Proteins to the RER
5.8K
A sizable fraction of proteins destined for ER are first synthesized in the cell cytosol and then transported across the ER membrane–a process called post-translational translocation. Similar to cotranslationally translocated proteins, these proteins also use the Sec translocon complex to enter the ER lumen.
Targeting proteins to the ER
Hsp40 and Hsp70 chaperone molecules bind the translated proteins in the cytosol to prevent their folding. The chaperone binding helps to keep the signal...
Targeting proteins to the ER
Hsp40 and Hsp70 chaperone molecules bind the translated proteins in the cytosol to prevent their folding. The chaperone binding helps to keep the signal...
5.8K